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End-tidal-to-arterial gas gradients during dynamic end-tidal forcing Tymko, Michael Martin 2015

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  END-TIDAL-TO-ARTERIAL GAS GRADIENTS DURING  DYNAMIC END-TIDAL FORCING  by  Michael Martin Tymko  B.HSc., Mount Royal University, 2013  A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF  MASTER OF SCIENCE  in  THE COLLEGE OF GRADUATE STUDIES  (Interdisciplinary Studies)  THE UNIVERSITY OF BRITISH COLUMBIA  (Okanagan)  JULY 2015   © Michael Martin Tymko, 2015   ii Abstract The end-tidal gas partial pressure is often considered a surrogate for the partial pressure of arterial blood gas; however, in healthy humans an end-tidal-to-arterial gradient normally exists for O2 (PET–PaO2) and CO2 (Pa–PETCO2).  We sought to determine if the Pa-PETCO2 affects the measurement of cerebrovascular reactivity (CVR) and the hypercapnic ventilatory response (HCVR), and whether it could be corrected for in subjects with (n=8) and without (n=7) a patent foramen ovale (PFO) during a CO2 reactivity test.  It was hypothesized that (1) the Pa-PETCO2 would be greater in the background of hypoxia compared to normoxia during a CO2 reactivity test, (2) the PET-PaCO2 would be similar while the PET-PaO2 gradient would be greater in those with a PFO, (3) the HCVR and CVR would be lower when plotted against PETCO2 compared to PaCO2, and (4) a PaCO2 prediction algorithm will correct for the Pa-PETCO2.  PETCO2 was controlled by dynamic end-tidal forcing in steady-state steps of -8, -4, 0, +4, and +8mmHg from baseline in normoxia (NX1; PETO2 = 94.3 ± 1.3 mmHg) and hypoxia (HX1; PETO2 = 50.8 ± 0.1 mmHg).  Tests were repeated following correction for the Pa-PETCO2 (NX2 and HX2).  Internal carotid artery blood flow (Q̇ICA), middle cerebral artery blood flow velocity (MCAv) and temperature-corrected end-tidal and arterial blood gases were measured throughout each protocol.  CVR was calculated using linear regression analysis in the hypocapnic and hypercapnic ranges by indexing the percent change in Q̇ICA, and MCAv against PETCO2 and PaCO2.  In both conditions, a Pa-PETCO2 was present in hypercapnia but not hypocapnia, and was unchanged by PFO (P>0.05), however, the PET-PaO2 was greater in PFO+ participants in normoxia (P=0.003).  Relative Q̇ICA CVR, MCAv CVR, and HCVR assessed using PETCO2 were less compared to using PaCO2 during both normoxia and hypoxia (P>0.05).  A previously derived prediction equation minimized the difference between measured and predicted PaCO2 during +4mmHg (NX2: 0.0  0.2 mmHg, P=0.894; HX2: -0.2  0.2 mmHg, P=0.403) and +8mmHg for HX2 (0.0  0.3 mmHg, P=0.860).  In conclusion, care must be taken when indexing reactivity measures to PETCO2 compared to PaCO2.    iii Preface This thesis contains original data collected and analyzed for partial fulfillment of the author’s Master of Science degree.  All protocols were approved by the Clinical Research Ethics Board (UBC number: H14-01128) at the University of British Columbia.  This thesis consists of a review of the literature (Chapter 1: Introduction), and three additional chapters pertaining to the research questions, methodology, results, and discussion.  Appendix A consists of published work conducted at the University of British Columbia (Okanagan) by Mr. Michael M. Tymko, Dr. Philip N. Ainslie, Dr. David B. MacLeod, Dr. Chris K. Willie, and Dr. Glen E. Foster.  All authors were involved in the conception and design of the research, helped perform the experiments, and aided in the interpretation of the experimental results along with editing the drafted manuscript.  I was responsible for data analysis, figure preparation, and writing the drafted manuscript.  Permission to reproduce this material within my thesis was not required by the American Physiological Society.    iv Table of Contents  Abstract .................................................................................................................................... ii Preface ..................................................................................................................................... iii Table of Contents ................................................................................................................... iv List of Tables ........................................................................................................................ viii List of Figures ....................................................................................................................... xiv List of Illustrations ............................................................................................................... xvi List of Symbols and Abbreviations ................................................................................... xvii Acknowledgements ............................................................................................................... xx Dedication ............................................................................................................................. xxi Chapter 1: Introduction ......................................................................................................... 1  Pulmonary gas exchange and the control of arterial blood gases ......................................... 1  Dynamic end-tidal forcing as a method for controlling arterial blood gases independent  of minute ventilation .......................................................................................................................... 3 1.2.1 Methodological considerations with dynamic end-tidal forcing ...................................... 5  Common outcome measurements while using dynamic end-tidal forcing ........................... 5 1.3.1 Cerebrovascular reactivity to CO2 .................................................................................... 6 1.3.2 Ventilatory reactivity to CO2 ............................................................................................ 7  The end-tidal-to-arterial gas gradient .................................................................................... 8  Factors affecting gas gradients ............................................................................................ 10 1.5.1 Diffusion limitation ........................................................................................................ 10 1.5.2 Ventilation-perfusion mismatch ..................................................................................... 11 1.5.3 Intracardiac and intrapulmonary shunting. ..................................................................... 12 1.5.4 High-altitude ................................................................................................................... 14  Dynamic end-tidal forcing vs. the rebreathing technique ................................................... 15  End-tidal vs arterial CO2? ................................................................................................... 16  Research questions and hypothesis ..................................................................................... 18 Chapter 2: Materials and Methods ..................................................................................... 20   v  Ethical approval .................................................................................................................. 20  Participants .......................................................................................................................... 20  Prescreening ........................................................................................................................ 21 2.3.1 Patent foramen ovale screening ...................................................................................... 21 2.3.2 Pulmonary function testing ............................................................................................. 21  Experimental protocol ......................................................................................................... 22 2.4.1 Normoxia CO2 reactivity protocols ................................................................................ 22 2.4.2 Hypoxia CO2 reactivity protocols ................................................................................... 23  Outcome measurements ...................................................................................................... 23 2.5.1 Participant instrumentation ............................................................................................. 24 2.5.2 Respiratory measurements .............................................................................................. 24 2.5.3 End-tidal forcing ............................................................................................................. 25  Cerebral blood flow measurements ..................................................................................... 26 2.6.1 Intracranial blood flow ................................................................................................... 26 2.6.2 Extracranial blood flow .................................................................................................. 26  Arterial blood sampling and blood pressure ....................................................................... 27  Data analysis ....................................................................................................................... 27 2.8.1 Cerebrovascular reactivity in normoxia and hypoxia ..................................................... 28 2.8.2 Hypercapnic ventilatory responses in normoxia and hypoxia ........................................ 28 2.8.3 Intracardiac shunt scoring ............................................................................................... 28 2.8.4 Sample size justification ................................................................................................. 28 2.8.5 Statistics .......................................................................................................................... 29 Chapter 3: Results................................................................................................................. 31  Participants .......................................................................................................................... 31  Research Question One: end-tidal-to-arterial CO2 and O2 gradients .................................. 32 3.2.1 Normoxia protocols ........................................................................................................ 32 3.2.1.1 NX1 CO2 protocol .................................................................................................. 32 3.2.1.2 NX2 CO2 protocol .................................................................................................. 33 3.2.1.3 NX1 vs. NX2 protocols .......................................................................................... 33 3.2.2 Hypoxia Protocols .......................................................................................................... 38 3.2.2.1 HX1 CO2 protocols ................................................................................................ 38 3.2.2.2 HX2 CO2 protocols ................................................................................................ 39 3.2.2.3 HX1 vs. HX2 CO2 protocols .................................................................................. 39 3.2.3 Comparison of normoxia and hypoxia CO2 protocols .................................................... 44   vi  Research Question Two: end-tidal-to-arterial gradients in PFO+ and PFO-   participants .......................................................................................................................... 47 3.3.1 Normoxia protocols ........................................................................................................ 47 3.3.2 Hypoxia protocols ........................................................................................................... 47  Research Question Three: PETCO2 vs PaCO2 cerebrovascular and ventilatory  reactivity........................................................................................................................................... 52 3.4.1 Normoxia reactivity data ................................................................................................ 52 3.4.1.1 Hypercapnic ventilatory reactivity data ................................................................. 52 3.4.1.2 Internal carotid artery reactivity ............................................................................. 53 3.4.2 Hypoxia reactivity data ................................................................................................... 55 3.4.2.1 Hypercapnic ventilatory reactivity data ................................................................. 55 3.4.2.2 Internal carotid artery reactivity ............................................................................. 57 3.4.2.3 Middle cerebral artery reactivity ............................................................................ 58  Research Question Four: performance of our PaCO2 correction algorithm ........................ 59 3.5.1 Comparison between new and old prediction algorithms for PaCO2 ............................. 59 3.5.2 Normoxia and hypoxia data ............................................................................................ 60 3.5.3 Performance PaCO2 correction algorithm in normoxia and hypoxia ............................. 60 Chapter 4: Discussion ........................................................................................................... 67  Summary of main findings .................................................................................................. 67  Research Question One: any differences in the end-tidal-to-arterial gradients between normoxia and hypoxia? .................................................................................................................... 67 4.2.1 The end-tidal-to-arterial CO2 gradient ............................................................................ 67 4.2.2 The end-tidal-to-arterial O2 gradient ............................................................................... 69 4.2.3 Targeting arterial blood gases ......................................................................................... 70  Research Question Two: were there any differences in the end-tidal-to-arterial  gradients between PFO+ and PFO- participants? ............................................................................ 71 4.3.1 End-tidal-to-arterial carbon dioxide gradient ................................................................. 71 4.3.2 End-tidal-to-arterial oxygen gradient.............................................................................. 72 4.3.3 Effect of PFO on hypercapnic ventilatory reactivity ...................................................... 72  Research Question Three: are calculated cerebrovascular reactivity and ventilatory reactivity to CO2 attenuated by using PETCO2 instead of PaCO2? .................................................... 72 4.4.1 Hypercapnic ventilatory response in normoxia and hypoxia .......................................... 73 4.4.2 Middle cerebral artery velocity reactivity in normoxia and hypoxia .............................. 73 4.4.3 Internal carotid artery blood flow reactivity in normoxia and hypoxia .......................... 74   vii  Research Question Four: does our previously derived correction algorithm accurately predict PaCO2? ................................................................................................................................. 74 4.5.1 Performance of previously derived PaCO2 invasive correction algorithms ................... 75 4.5.2 Performance of previously derived PaCO2 non-invasive correction algorithms ............ 75 4.5.3 Performance of newly derived PaCO2 correction algorithms ......................................... 76  Methodological considerations. .......................................................................................... 76 4.6.1 Utility of our PaCO2 invasive correction algorithm ....................................................... 76 4.6.2 The use of transcranial Doppler ultrasound to measure cerebral blood flow ................. 77 4.6.3 Reproducibility of arterial blood gas measurements ...................................................... 77 4.6.4 Nasopharyngeal temperature as a surrogate for core body temperature ......................... 78  Perspective and Significance. ............................................................................................. 78 References .............................................................................................................................. 80 Appendices ........................................................................................................................... 100 Appendix A : Tymko et al. (2015). End-tidal-to-arterial CO2 and O2 gas gradients  at low- and high-altitude during dynamic end-tidal forcing. .......................................... 100 A.1 Abstract ......................................................................................................................... 101 A.2 Introduction .................................................................................................................. 102 A.3 Materials and methods .................................................................................................. 105 A.4 Results .......................................................................................................................... 111 A.5 Discussion ..................................................................................................................... 116 A.6 References .................................................................................................................... 124 Appendix B : University of British Columbia ethics certificate of full  board approval .................................................................................................................... 140 Appendix C : NX1, NX2, HX1, and HX2 CO2 reactivity protocol raw data ................. 142   viii List of Tables   Table 3.1: Pulmonary pre-screening data. .............................................................................. 31 Table 3.2: Ventilatory cardiovascular, and cerebrovascular data during NX1 and NX2   CO2 reactivity tests. .................................................................................................. 36 Table 3.3. Ventilatory and blood gas data during NX1 and NX2 CO2 reactivity tests. ......... 37 Table 3.4: Ventilatory cardiovascular, and cerebrovascular data during HX1 and HX2   CO2 reactivity tests. .................................................................................................. 42 Table 3.5: Ventilatory and blood gas data during HX1 and HX2 CO2 reactivity tests. ......... 43 Table 3.6: Comparison of ventilatory cardiovascular, and cerebrovascular data between   NX1 and HX1 CO2 reactivity tests. .......................................................................... 45 Table 3.7: Comparison of ventilatory and blood gas data during NX1 and HX1 CO2 reactivity tests. .......................................................................................................... 46 Table 3.8: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during NX1 CO2 reactivity test................................................................................. 48 Table 3.9: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during NX2 CO2 reactivity test................................................................................. 49 Table 3.10: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during HX1 CO2 reactivity test................................................................................. 50 Table 3.11: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during HX2 CO2 reactivity test................................................................................. 51 Table 3.12: A comparison of linear regression correction algorithms for PaCO2 derived   from Tymko et al., and the NX1 CO2 reactivity protocol. ...................................... 61 Table 3.13: Summary of the performance of our PaCO2 correction algorithm during normoxia and hypoxia trials. .................................................................................... 66 Table A.1: Ventilatory data during an isooxic CO2 test at LA and HA................................ 130 Table A.2 End-tidal and arterial blood gases during an isooxic CO2 test at LA and HA. .... 131 Table A.3 Ventilatory and arterial blood gas data during an isocapnic O2 test at LA   and HA. ................................................................................................................... 132 Table C.1: Subject demographic raw data for each participant ............................................ 142 Table C.2: Pulmonary function raw data for each participant. ............................................. 143   ix Table C.3: Pulmonary raw data for each participant ............................................................ 144 Table C.4: Subject demographic raw data for each participant ............................................ 145 Table C.5: Subject demographic raw data for each participant ............................................ 146 Table C.6: Minute ventilation (l/min) raw data for each participant .................................... 147 Table C.7: Minute ventilation (l/min) raw data for each participant during the NX2 CO2 reactivity protocol. .................................................................................................. 148 Table C.8: Minute ventilation (l/min) raw data for each participant during the HX1 CO2 reactivity protocol. .................................................................................................. 149 Table C.9: Minute ventilation (l/min) raw data for each participant during the HX2 CO2 reactivity protocol. .................................................................................................. 150 Table C.10. Tidal volume (l) raw data for each participant. NX1 CO2 reactivity   protocol. .................................................................................................................. 151 Table C.11. Tidal volume (l) raw data for each participant during the NX2 CO2   reactivity protocol. .................................................................................................. 152 Table C.12. Tidal volume (l) raw data for each participant during the HX1 CO2   reactivity protocol. .................................................................................................. 153 Table C.13. Tidal Volume (l) raw data for each participant during the HX2 CO2   reactivity protocol. .................................................................................................. 154 Table C.14. Breathing frequency (breaths/min) raw data for each participant ..................... 155 Table C.15. Breathing frequency (breaths/min) raw data for each participant during the   NX2 CO2 reactivity protocol. ................................................................................. 156 Table C.16. Breathing frequency (breaths/min) raw data for each participant during the   HX1 CO2 reactivity protocol. ................................................................................. 157 Table C.17. Breathing frequency (breaths/min) raw data for each participant during the   HX2 CO2 reactivity protocol. ................................................................................. 158 Table C.18. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each   participant ............................................................................................................... 159 Table C.19. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each   participant during the NX2 CO2 reactivity protocol. .............................................. 160 Table C.20. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each   participant during the HX1 CO2 reactivity protocol. .............................................. 161   x Table C.21. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each   participant during the HX2 CO2 reactivity protocol. .............................................. 162 Table C.22. Partial pressure end-tidal oxygen (mmHg) raw data for each participant. ....... 163 Table C.23. Partial pressure end-tidal oxygen (mmHg) raw data for each participant   during the NX2 CO2 reactivity protocol. ................................................................ 164 Table C.24. Partial pressure end-tidal oxygen (mmHg) raw data for each participant   during the HX1 CO2 reactivity protocol. ................................................................ 165 Table C.25. Partial pressure end-tidal oxygen (mmHg) raw data for each participant   during the HX2 CO2 reactivity protocol. ................................................................ 166 Table C.26. Partial pressure of inspired carbon dioxide (mmHg) raw data for each   participant NX1 CO2 reactivity protocol. .............................................................. 167 Table C.27. Partial pressure of inspired carbon dioxide (mmHg) raw data for each   participant during the NX2 CO2 reactivity protocol. .............................................. 168 Table C.28. Partial pressure of inspired carbon dioxide (mmHg) raw data for each   participant during the HX1 CO2 reactivity protocol. .............................................. 169 Table C.29. Partial pressure of inspired carbon dioxide (mmHg) raw data for each   participant during the HX2 CO2 reactivity protocol. .............................................. 170 Table C.30. Core body temperature (C) raw data for each ................................................. 171 Table C.31. Core body temperature (C) raw data for each participant during the NX2   CO2 reactivity protocol. .......................................................................................... 172 Table C.32. Core body temperature (C) raw data for each participant during the HX1   CO2 reactivity protocol ........................................................................................... 173 Table C.33. Core body temperature (C) raw data for each participant during the HX2   CO2 reactivity protocol. .......................................................................................... 174 Table C.34. Oxygen saturation of hemoglobin (%) raw data for each participant. .............. 175 Table C.35. Oxygen saturation of hemoglobin (%) raw data for each participant during   the NX2 CO2 reactivity protocol ............................................................................ 176 Table C.36. Oxygen saturation of hemoglobin (%) raw data for each participant during   the HX1 CO2 reactivity protocol ............................................................................ 177 Table C.37. Oxygen saturation of hemoglobin (%) raw data for each participant during   the HX2 CO2 reactivity protocol. ........................................................................... 178   xi Table C.38. Heart rate (bpm) raw data for each participant. ................................................ 179 Table C.39. Heart rate (bpm) raw data for each participant during the NX2 CO2   reactivity protocol. .................................................................................................. 180 Table C.40. Heart rate (bpm) raw data for each participant during the HX1 CO2   reactivity protocol. .................................................................................................. 181 Table C.41. Heart rate (bpm) raw data for each participant during the HX2 CO2   reactivity protocol. .................................................................................................. 182 Table C.42. Mean arterial pressure (mmHg) raw data for each participant. ........................ 183 Table C.43. Mean arterial pressure (mmHg) raw data for each participant during the   NX2 CO2 reactivity protocol. ................................................................................. 184 Table C.44. Mean arterial pressure (mmHg) raw data for each participant during the   HX1 CO2 reactivity protocol. ................................................................................. 185 Table C.45. Mean arterial pressure (mmHg) raw data for each participant during the   HX2 CO2 reactivity test. ......................................................................................... 186 Table C.46. Middle cerebral artery velocity (cm/s) raw data for each participant ............... 187 Table C.47. Middle cerebral artery velocity (cm/s) raw data for each participant during  the NX2 CO2 reactivity protocol. ........................................................................... 188 Table C.48. Middle cerebral artery velocity (cm/s) raw data for each participant during   the HX1 CO2 reactivity protocol. ........................................................................... 189 Table C.49. Middle cerebral artery velocity (cm/s) raw data for each participant during   the HX2 CO2 reactivity protocol. ........................................................................... 190 Table C.50. Arterial pH raw data for each participant.NX1 CO2 reactivity protocol. .......... 191 Table C.51. Arterial pH raw data for each participant during the NX2 CO2 reactivity protocol. .................................................................................................................. 192 Table C.52. Arterial pH raw data for each participant during the HX1 CO2 reactivity protocol. .................................................................................................................. 193 Table C.53. Arterial pH raw data for each participant during the HX2 CO2 reactivity protocol. .................................................................................................................. 194 Table C.54. Partial pressure of arterial carbon dioxide (mmHg) raw data for each participant.NX1 CO2 reactivity protocol. ............................................................... 195 Table C.55. Partial pressure of arterial carbon dioxide (mmHg) raw data for each    xii  participant during the NX2 CO2 reactivity protocol. .............................................. 196 Table C.56. Partial pressure of arterial carbon dioxide (mmHg) raw data for each   participant during the HX1 CO2 reactivity protocol. .............................................. 197 Table C.57. Partial pressure of arterial carbon dioxide (mmHg) raw data for each   participant during the HX2 CO2 reactivity protocol. .............................................. 198 Table C.58. Partial pressure of arterial oxygen (mmHg) raw data for each participant.   NX1 CO2 reactivity protocol. ................................................................................. 199 Table C.59. Partial pressure of arterial oxygen (mmHg) raw data for each participant   during the NX2 CO2 reactivity protocol. ................................................................ 200 Table C.60. Partial pressure of arterial oxygen (mmHg) raw data for each participant   during the HX1 CO2 reactivity protocol. ................................................................ 201 Table C.61. Partial pressure of arterial oxygen (mmHg) raw data for each participant   during the HX2 CO2 reactivity protocol. ................................................................ 202 Table C.62. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant NX1 CO2 reactivity protocol. ............................................................... 203 Table C.63. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. .............................................. 204 Table C.64. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the HX1 protocol. ...................................................................... 205 Table C.65. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. .............................................. 206 Table C.66. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant   NX1 CO2 reactivity protocol. ................................................................................. 207 Table C.67. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. ................................................................ 208 Table C.68. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. ................................................................ 209 Table C.69. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. ................................................................ 210 Table C.70. Internal carotid artery flow (ml/min) raw data for each participant .................. 211   xiii Table C.71. Internal carotid artery flow (ml/min) raw data for each participant during the NX2 CO2 reactivity protocol. ................................................................................. 212 Table C.72. Internal carotid artery flow (ml/min) raw data for each participant during the HX1 CO2 reactivity protocol. ................................................................................. 213 Table C.73. Internal carotid artery flow (ml/min) raw data for each participant during the HX2 CO2 protocol. ................................................................................................. 214 Table C.74. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2   raw data for each participant NX1 CO2 protocol. .................................................. 215 Table C.75. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2   raw data for each participant during the NX2 CO2 protocol. ................................. 216 Table C.76. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2   raw data for each participant during the HX1 CO2 protocol. ................................. 217 Table C.77. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2   raw data for each participant during the HX2 CO2 protocol. ................................. 218 Table C.78. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2   raw data for each participant NX1 CO2 protocol. ................................................... 219 Table C.79. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2   raw data for each participant during the NX2 CO2 protocol. ................................. 220 Table C.80. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2   raw data for each participant during the HX1 CO2 protocol. ................................. 221 Table C.81. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2   raw data for each participant during the HX2 CO2 protocol. ................................. 222 Table C.82. Internal carotid artery, middle cerebral artery, and HCVR to predicted   arterial CO2 raw data for each participant NX1 CO2 protocol. .............................. 223 Table C.83. Internal carotid artery, middle cerebral artery, and HCVR to predicted   arterial CO2 raw data for each participant during the NX2 CO2 protocol. ............. 224 Table C.84. Internal carotid artery, middle cerebral artery, and HCVR to predicted   arterial CO2 raw data for each participant during the HX1 CO2 protocol. ............. 225 Table C.85. Internal carotid artery, middle cerebral artery, and HCVR to predicted   arterial CO2 raw data for each participant during the HX2 CO2 protocol. ............. 226    xiv List of Figures  Figure 3.1: End-tidal gas tracings from NX1 (A) and NX2 (B) CO2 reactivity trials for   one representative subject. ........................................................................................ 34 Figure 3.2: Bland and Altman plot for agreement between PaCO2 and PETCO2 during   NX1 (A) and NX2 (B) CO2 reactivity protocols. ..................................................... 34 Figure 3.3: Bland and Altman plot for agreement between PaO2 and PETO2 during NX1   (A) and NX2 (B) CO2 reactivity protocols. .............................................................. 35 Figure 3.4: End-tidal gas tracings from HX1 and HX2 CO2 reactivity trials from one representative subject. ............................................................................................... 40 Figure 3.5: Bland and Altman plot for agreement between PaCO2 and PETCO2 during   HX1 (Panel A) and HX2 (Panel B) CO2 reactivity protocols. ................................. 40 Figure 3.6: Bland and Altman plot for agreement between PaO2 and PETO2 during HX1 (Panel A) and HX2 (Panel B) CO2 reactivity protocols. .......................................... 41 Figure 3.7: Hypercapnic ventilatory reactivity data during NX1 (Panel A) and NX2   (Panel B) protocols. .................................................................................................. 52 Figure 3.8: Hypercapnic ventilatory reactivity data during NX1 (Panel A) and NX2   (Panel B) protocols between PFO+ (n=7) and PFO- (n=7) participants. ................. 53 Figure 3.9: ICA hypocapnia and hypercapnia reactivity data during NX1 (Panel A) and  NX2 (Panel B) protocols. ......................................................................................... 54 Figure 3.10: MCA hypocapnia and hypercapnia reactivity data during NX1 (Panel A)   and NX2 (Panel B) protocols. ................................................................................... 55 Figure 3.11: Hypercapnic ventilatory reactivity data during HX1 (Panel A) and HX2   (Panel B) protocols. .................................................................................................. 56 Figure 3.12: Hypercapnic ventilatory reactivity data during HX1 (Panel A) and HX2   (Panel B) protocols between PFO+ (n=8) and PFO- (n=6) participants. ................. 57 Figure 3.13: ICA hypocapnia and hypercapnia reactivity data during HX1 (Panel A) and HX2 (Panel B) protocols. ......................................................................................... 58 Figure 3.14: MCAv hypocapnia and hypercapnia reactivity data during HX1 (Panel A)   and HX2 (Panel B) protocols. ................................................................................... 59 Figure 3.15: Assessment of PaCO2 and PETCO2 relationship of CO2 during NX1 and    xv  NX2 CO2 reactivity protocols. .................................................................................. 62 Figure 3.16: Assessment of PaCO2 and PETCO2 relationship of CO2 during NX2 CO2 reactivity protocols, using new algorithms derived from NX1 CO2 reactivity protocol. .................................................................................................................... 63 Figure 3.17: Assessment of PaCO2 and PETCO2 relationship of CO2 during HX1 and   HX2 CO2 reactivity protocols. .................................................................................. 64 Figure 3.18: Assessment of PaCO2 and PETCO2 relationship of CO2 during HX2 CO2 reactivity protocols, using new algorithms derived from HX1 CO2 reactivity protocol. .................................................................................................................... 65 Figure 4.1: Bland and Altman plot for agreement between two consecutive arterial   blood gas measurements (on the same sample) for CO2 (A) and O2 (B) during eupneic air breathing (n=30). .................................................................................... 78 Figure A.1: Representative tracing of isooxic CO2 test at LA and HA ................................ 135 Figure A.2: Bland and Altman plot of differences between PaCO2 and PETCO2 during   an isooxic CO2 test at LA and HA .......................................................................... 136 Figure A.3: Representative tracing of the isocapnic O2 test at LA and HA ......................... 137 Figure A.4: Bland and Altman plot of differences between PaO2 and PETO2 partial   pressures of O2 during an isocapnic O2 test at LA and HA .................................... 138 Figure A.5: Assessment of PaCO2 and PETCO2 relationship of CO2 during an isooxic   CO2 test at LA and HA ........................................................................................... 139    xvi List of Illustrations  Illustration 1.1: Central (CCR) and peripheral (PCR) respiratory chemoreceptors      traditional negative feedback arcs. ............................................................................. 2 Illustration 1.2: Schematic of our laboratory’s DEF system..................................................... 4 Illustration 1.3: Diagram of the ‘Charged Membrane Hypothesis’ proposed by              Gurtner et al. (1979). .................................................................................................. 9 Illustration 1.4: A schematic representation of blood flow through an intrapulmonary       shunt. ......................................................................................................................... 13 Illustration 1.5: Conceptual display of the potential effect that the end-tidal-to-arterial      CO2 gradient has on hypercapnic HCVR. ................................................................ 16 Illustration 1.6: Conceptual illustration of the potential effect the Pa-PETCO2 gradient         has on CVR to CO2. .................................................................................................. 17   xvii List of Symbols and Abbreviations Symbol Definition  AaCO2 Alveolar-to-arterial carbon dioxide difference AaDO2 Alveolar-to-arterial oxygen difference CaO2 Content of oxygen in arterial blood CcO2 Content of oxygen in pulmonary end-capillary blood CO2 Carbon dioxide CCR Central respiratory chemoreceptor CvO2 Content of oxygen in venous blood CVR Cerebrovascular reactivity to carbon dioxide DEF Dynamic end-tidal forcing DLCO Diffusion capacity of the lung for carbon monoxide transfer ECG Electrocardiogram FEV1 Forced expiratory volume in one second FICO2 Fraction of inspired carbon dioxide FIO2 Fraction of inspired oxygen FVC Forced vital capacity FRC Functional residual capacity HA High-altitude HCVR Hypercapnic ventilatory reactivity HPV Hypoxic pulmonary vasoconstriction HR Heart rate [H+] Hydrogen ion concentration HX-CVR Hypoxic cerebrovascular reactivity to CO2 HX-HCVR Hypoxia hypercapnic ventilatory reactivity HX1 Hypoxic CO2 reactivity protocol 1 HX2 Hypoxic CO2 reactivity protocol 2 HYPER-ICA Hypercapnic relative internal carotid artery blood flow reactivity HYPO-ICA Hypocapnic relative internal carotid artery blood flow reactivity HYPER-MCA Hypercapnic relative middle cerebral artery blood velocity reactivity   xviii HYPO-MCA Hypocapnic relative middle cerebral artery blood velocity reactivity IC Inspiratory capacity ICA Internal carotid artery IPAVA Intrapulmonary arteriovenous anastomoses LA Low-altitude MAP Mean arterial pressure MCA Middle cerebral artery MCAv Middle cerebral artery velocity N2 Nitrogen NX1 Normoxic CO2 reactivity protocol 1 NX2 Normoxic CO2 reactivity protocol 2 O2 Oxygen PaCO2 Arterial partial pressure of carbon dioxide PaO2 Arterial partial pressure of oxygen PCO2 Partial pressure of carbon dioxide PCR Peripheral respiratory chemoreceptor PETCO2 End-tidal partial pressure of carbon dioxide PETO2 End-tidal partial pressure of oxygen PET-PaCO2 End-tidal-to-arterial carbon dioxide gradient PET-PaO2 End-tidal-to-arterial oxygen gradient PFO Patent foramen ovale PICO2 Partial pressure of inspired carbon dioxide PIO2 Partial pressure of inspired oxygen PO2 Partial pressure of oxygen Pred-PaCO2 Predicted arterial partial pressure of carbon dioxide Q̇ Blood flow Q̇ICA Internal carotid artery blood flow Q̇s/Q̇t Shunt fraction RV Residual volume SaO2 Arterial oxyhaemoglobin saturation TCD Transcranial Doppler ultrasound   xix TLC Total lung capacity VC Vital capacity VA Alveolar volume V̇E Minute ventilation V̇/Q Ventilation-to-perfusion ratio VT Tidal volume   xx  Acknowledgements I would like to acknowledge my supervisor, Dr Glen Foster, and the members of my supervisory committee, Drs Philip Ainslie and Craig Steinback for their guidance and scholarly support throughout my studies. I also want to acknowledge Drs Trevor Day and Neil Eves for their mentorship over recent years.    xxi Dedication My parents and sisters, you have provided me with unconditional support throughout my studies, I would not have been able to do this without you. To my friends, each of you has helped me through the long days with your constant activity in “Beauties” chat.  Lastly, to my great grandma, Stanna Falzarano, you gave me 24 years of beautiful memories and with every conversation I gained everlasting happiness and inspiration.     1 Chapter 1: Introduction In this chapter, the physiological mechanisms that control arterial blood gas tensions in humans are briefly reviewed.  In addition, methodology commonly used experimentally to alter and control arterial blood gases including the utility of dynamic end-tidal forcing (DEF) for the assessment of hypercapnic ventilatory reactivity  (HCVR) and cerebrovascular reactivity (CVR) to carbon dioxide are discussed.  Although DEF is a powerful device for human physiology research, multiple considerations should be considered prior to its employment.    Pulmonary gas exchange and the control of arterial blood gases The exchange of oxygen (O2) and carbon dioxide (CO2) between the air that we breathe and pulmonary capillary blood is essential for human life.  During inspiration, the volume of the thoracic cavity expands with the help of the diaphragm and accessory muscles of respiration, creating a pressure gradient for the flow of air high in O2 (fraction of inspired O2 (FIO2) = 0.21) and low in CO2 (FICO2 = 0.0004) from the exterior and into the bronchial tree and ~300-500 million polyhedral air sacs called alveoli.  At the alveoli, inspired fresh gas can equilibrate with the gas contained within pulmonary capillary blood, which contains a lower concentration of O2 and a higher concentration of CO2, through simple diffusion across the alveolar/capillary boundary. This process is called pulmonary ventilation.  Maintaining adequate arterial O2 content and elimination of CO2 from the blood via pulmonary ventilation is tightly regulated by the respiratory chemoreceptors.  Respiratory chemoreceptors are cells that respond to changes in arterial blood gas tensions to modify respiratory rate and depth to control alveolar ventilation to aid in the exchange of gases at the alveolar-capillary interface and to maintain blood gas homeostasis.  Blood gas (i.e. O2 and CO2) homeostasis is coordinated in part through central and peripheral respiratory chemoreceptor negative-feedback arcs, each with their own respective sensitivity (i.e. responsiveness) and onset delay following a change in chemical stimulus (usually PaO2, PaCO2/pH).  The location of the central respiratory chemoreceptors has not been definitively proven, but there is good evidence that neurons and glial cells that act as chemoreceptors for PaCO2/pH are located within the retrotrapezoid nucleus and the raphe in the brain stem (53, 81, 82) and respond more slowly compared to the peripheral chemoreceptors, which are located in the carotid and aortic bodies (46, 57, 86, 117).  The central respiratory chemoreceptors directly respond to changes in hydrogen ion concentration ([H+]) within cerebrospinal fluid, which is largely mediated by changes in the arterial partial pressure of CO2 (PaCO2) (3, 31).  Above eupneic PaCO2 levels, there is a strong linear relationship between pulmonary ventilation and PaCO2 in   2 most individuals, where an increase in [H+] (caused by increases in PaCO2) results in an increase in chemoreflex (central and peripheral) driven ventilation, and vice versa (see Illustration 1, pg 2).  The peripheral respiratory chemoreceptors, also responsive to changes in [H+], respond to changes in both arterial partial pressure of O2 (PaO2) and PaCO2 (31), whereas the central respiratory chemoreceptors are traditionally not thought to be sensitive to PaO2.  However, there is evidence suggesting that central chemoreceptors may respond during severe hypoxia (35).  Because peripheral chemoreceptors respond to both PaO2 and PaCO2/pH, the O2-CO2 stimulus interaction within the peripheral respiratory chemoreceptors results in an augmented CO2 response in hypoxia, and a blunted CO2 response in hyperoxia (45, 47, 68), which is transduced into the respective ventilatory response (28, 84).  It is the combination of neural inputs to the central respiratory rhythm/pattern generator from both the central and peripheral chemoreceptors that plays a role in the regulation of breathing in humans. However, the control of breathing is complex and involves the coordination of many different neural inputs to allow the body to maintain blood gas homeostasis during sleep, exercise and at altitude.     Illustration 1.1: Central (CCR) and peripheral (PCR) respiratory chemoreceptors traditional negative feedback arcs.  Respiratory chemoreceptors are stimulated by changes PaCO2 and PaO2 through minute ventilation (V̇E). The PCR’s detect changes in PaCO2 and PaO2, while the CCR’s detect changes in PaCO2 and [H+] within tissue near the brainstem. Synergistic afferent feedback (solid lines) from these receptors is integrated within the respiratory controller in the medulla oblongata, and efferent feedback (dashed lines) is sent to the respiratory muscles to initiate a change in ventilation (V̇E).  + represents excitatory stimulus.    3    Dynamic end-tidal forcing as a method for controlling arterial blood gases independent of minute ventilation Dynamic end-tidal forcing (DEF) is a servo-mechanism (automatic negative feedback system), and feed-forward system used to control PaO2 and PaCO2 by manipulating the end-tidal partial pressure of O2 (PETO2) and CO2 (PETCO2) independent of minute ventilation (V̇E).  In other words, arterial blood gases can be maintained at a desired level independent of the ventilatory response.  This is an advantage over using fixed fraction administration of gases (i.e. Douglas bag) because it standardizes the chemical stimulus between subjects.  Originating in the early seventies (110-112), DEF was initially developed to quantify the effect of specific pharmaceuticals on the ventilatory responses to CO2 (69).  Dynamic end-tidal forcing is considered the gold standard method for altering arterial blood gases in human participants and has been employed for the purpose of investigating the control of breathing (19, 25, 63), as well as cerebral (88, 89), pulmonary (116), and cardiac reactivity (16) using CO2 and O2 reactivity tests.  This is a powerful method to detect changes in human physiology before and after an intervention (i.e. pharmacological or environmental).  Several different DEF systems have been developed over the past few decades, each with their own strengths and weaknesses (44, 65, 94, 106, 110).  Each of these systems functions based upon the assumption that end-tidal gases accurately reflect arterial blood gases. Robbins and colleagues (94) developed one of the first sophisticated computer controlled fast gas-mixing system which was adapted from the Swanson & Bellville design (110), and was capable of simultaneously clamping PETCO2 and PETO2 over a wide physiological range independent of V̇E.  The design necessitates a high-flow system, which is beneficial because it limits external deadspace allowing for more instantaneous changes in inspired gas content.  However, the required high flow of mixed gases produces a substantial amount of waste-gas increasing financial expense and limiting its use in the field.   More recently, a compact DEF system by Koehle and colleagues (65) has been developed.  This system is portable and cost effective as it uses compressed gases (medical grade CO2, N2 and “Air”) to deliver the desired gas mixture to an inspiratory reservoir bag by the time-dependent control of three solenoid valves (one for each mixing component).  Although effective, the increased external deadspace (from the inspiratory reservoir bag) can lead to a delay in gas delivery and temporary instability in the control system.  Similar to this DEF system, our research   4 laboratory recently developed a DEF system capable of controlling end-tidal gases while being cost effective and portable (see Appendix A, pg 100).  This system mixes three gases (O2, CO2, and N2) together in a 6-liter capacity reservoir bag to achieve the desired inspirate gas (118) (see Illustration 1.2, pg 4).  While the inspiratory reservoir has a 6-liter capacity it only injects a volume of air equal to the participant’s tidal volume (VT) plus an additional reserve of 100 ml for safety.  This volume can be controlled to minimize the deadspace and delay in delivering the required inspirate.  Due to its recent production our DEF system has had limited use, but has proven to effectively control end-tidal gases over a wide ranges of PETCO2 and PETO2 independent of V̇E at low-altitude (LA; 344 m) (48, 90, 118), high-altitude (HA; 5050 m) (118), and with hyperthermic interventions (9).      Illustration 1.2: Schematic of our laboratory’s DEF system. Using feedback information from the participant breathing through a flow sensor and gas analyzer (dashed lines), the DEF software, programmed in LabView (National Instruments, Austin, Tx), triggers three independent solenoid valves (one for CO2, O2, and N2) within a control box. The calculated mixed gas travels through a humidification chamber and into a 6-liter capacity inspiratory reservoir bag for the participant to breathe from, on a breath-by-breath basis.        5 1.2.1 Methodological considerations with dynamic end-tidal forcing Despite the advantages of our DEF system being portable, cost effective due to low gas requirements, and capable of controlling end-tidal gases within narrow limits, it is, similar to other systems is affected by methodological constraints.  (I) Similar to all DEF systems, hypocapnic steps can only be achieved by asking participants to actively hyperventilate, driving their PETCO2 below desired targets where the DEF is then capable of controlling and fine tuning PETCO2 control by adding CO2 into the breathing circuit.  Thus, with hypocapnia end-tidal gas control is dependent upon the ability of the participant to maintain a constant rate of V̇E. (II) Similar to other DEF systems (65), our DEF system functions by mixing inspirate into a reservoir bag on a breath-by-breath basis.  In an ideal world the reservoir bag would be emptied with each breath so that only the inspirate for the next breath is available to the subject at any given time.  However, in order to deal with breath-by-breath variation in VT a small amount of extra air remains in the reservoir bag to prevent its complete emptying.  This safety volume is normally 100 mL more than the current VT but can be controlled by the DEF operator to maintain the reservoir volume as small as possible.  In addition, the algorithm for the system estimates the volume of each gas remaining in the reservoir and takes these volumes into account before injecting the next volume of gases to better target the correct fractions of inspired O2 and CO2.  Nonetheless, inaccuracies in this prediction or inflation in the estimated reservoir volume can lead to a sluggish response or a delay in the systems response to a perturbation.  (III) Our DEF system uses 100% CO2, O2, and N2 compressed gas as source gases to fill our inspirate reservoir bag.  Although avoiding pre-mixed gases is usually less expensive, it does present some potential safety issues as it is possible for the participant to be administered 100% N2 during a severe hypoxic step or with software malfunction.  Therefore, it is crucial to employ a well-trained DEF device operator to recognize these potential issues.  If a problem arises in which the participant’s end-tidal gases are not being controlled safely, the participant can be quickly removed from the breathing apparatus or 100% O2 can be delivered to the participant.  Finally, (IV) similar to all DEF system, our DEF system assumes that end-tidal gases are a surrogate measure for arterial blood gases.   Common outcome measurements while using dynamic end-tidal forcing  Dynamic end-tidal forcing is commonly used to quantify reactivity measures that are responsive to changes in arterial blood gas tensions.  The human body has multiple mechanisms for regulating oxygen delivery to the tissues and these include local changes in blood flow, and changes in   6 alveolar ventilation.  The following two sections highlight the important relationship between cerebral blood flow and alveolar ventilation in response to changes in PaCO2.  1.3.1 Cerebrovascular reactivity to CO2 One of the most common outcome measurements obtained using DEF is cerebrovascular reactivity (CVR).  Cerebrovascular reactivity refers to vasomotor responsiveness of the blood vessels supplying nutrients to brain tissue to changes in PaCO2 and PaO2. The entire cerebrovasculature is sensitive to changes in blood gases; however, the cerebral blood flow response is principally mediated at the arterioles downstream from large conduit vessels (15, 44, 104, 129, 130).  Moreover, recent evidence suggests that large conduit vessels such as the internal carotid and vertebral arteries (ICA and VA, respectively) govern changes in cerebral blood flow as well (130).  The measurement of CVR to CO2 has been used to quantify cerebrovascular function in healthy populations, along with clinical populations such as patients with hypertension (100) and heart failure (134), and CVR impairment has been linked to cerebral ischemic episodes (i.e. stroke) (128).  Changes in both arterial and tissue CO2 and O2 can elicit changes in arteriolar diameter. Specifically, low O2 (hypoxia) and high CO2 (hypercapnia) cause dilatation, increasing the diameter and conductance of cerebral arterioles (6, 66, 77).  Changes in arterial gases can be experimentally induced by increasing or decreasing inspired gas levels, by hyperventilation, or with breath-holding. Changes in tissue gases can result from a change in the arterial-to-venous gas gradient or a change in the metabolic rate of the tissue (129).  Cerebral blood flow is commonly assessed by transcranial Doppler ultrasound (TCD), which measures blood flow velocity (62).  Although the TCD method is controversial, it has been used extensively and has contributed to the development of our current understanding of cerebrovascular physiology.  Transcranial Doppler ultrasound is easy to use, relatively affordable, and non-invasive, making it attractive for human physiology research.  However, the major caveats with TCD are: I) it assumes that the diameter of the cerebral artery remains unchanged, and II) although vessel confirmation tests have been developed, the vessel is insonated “blindly” (4).  It is currently accepted that TCD, if carefully conducted, can be a valuable tool used to measure resting cerebral blood flow in large intracranial conduit vessels (most commonly the middle cerebral artery, MCA; and posterior cerebral artery, PCA), and during mild hypo/hyper-capnia and hypo/hyper-oxia (130).  Recently, there has been some evidence suggesting that cerebral blood flow velocity evaluation using TCD underestimates true cerebral blood flow during modest levels   7 of hyper- and hypo-capnia (i.e. -13 to +9 mmHg PETCO2 from eupneic values) (26, 120).  Collectively, the experimental concern with TCD has resulted in using peripheral duplex ultrasound of large extracranial arteries (i.e. ICA and VA), where any changes in vessel diameter can be accounted for.  Cerebrovascular reactivity to CO2 is most commonly quantified by indexing cerebral blood flow (intracranially using TCD, and/or extracranially using peripheral ultrasound) against either PETCO2 or PaCO2 in hypocapnic and hypercapnic ranges separately, as hypocapnic CVR (HYPO-CVR) to CO2 is less than hypercapnic CVR (HYPER-CVR) to CO2.  There are diverging perspectives with regard to the linearity of CVR to CO2.  Specifically, Battisti-Charbonney and colleagues (15) advanced a non-linear approach to quantify CVR to CO2, whereby the cerebrovascular responses depart from linearity due to increases in arterial pressure (i.e. driving pressure) at extreme levels of CO2, in some, not all, participants (15, 97).  However, it is commonly accepted that linear regression analysis is fitting when quantifying CVR responses to CO2 over the range of PaCO2 normally studied in humans as r2 values are typically high (>0.80) with this relationship (54, 104, 130). Ultimately, to quantify CVR to CO2 responses, we require a consistent measurement of reactivity that warrants discussion on whether indexing cerebral blood flow against PETCO2 instead of PaCO2 is appropriate (see section 1.7, pg 16).    1.3.2 Ventilatory reactivity to CO2 Physiological mechanisms that are responsible for maintaining PaO2 and PaCO2 are amongst the most tightly regulated within the human body as a result of the ventilatory sensitivity to these stimuli.  Due to the linear relationship between V̇E and CO2, HCVR is typically quantified using linear regression analysis methods, and the reported slope between V̇E and CO2 above the participant’s CO2 ventilatory recruitment threshold is measured (i.e. the lowest CO2 stimulus that causes an increase in respiratory efforts).  Importance of measuring HCVR lies within its potential, yet controversial, inverse relationship with CVR reactivity to CO2 (3). It has been previously suggested that individuals with low CVR to CO2 would have a heightened HCVR, and vice-versa, in order to adequately regulate arterial blood gases. The measurement of HCVR has been used to quantify ventilatory function in healthy populations, along with clinical populations where the ventilatory control system may have become unstable in patients with chronic obstructive pulmonary disease (83), sleep apnea (29, 103), and cheyne-stokes respiration (8, 58). In addition to this, the HCVR has been a valuable tool when assessing changes in respiratory sensitivity when acclimatizing to high-altitude (HA) (5, 41).    8 Ventilatory reactivity to hypocapnia is not assessed in healthy individuals since active (i.e. coached) hyperventilation is used in order to achieve this change in PaCO2. In the hypocapnic range, the ventilatory response deteriorates due to a reduced chemoreflex-input ventilatory drive.    The end-tidal-to-arterial gas gradient End-tidal gas is the product of alveolar gas (active gas) and physiological deadspace.  Physiological deadspace contains air that does not participate in gas exchange (inactive gas).  There are two components that make up physiological deadspace: (I) alveolar deadspace from non-perfused ventilated alveoli, and (II) anatomical deadspace from airway structures that do not contribute to gas exchange (i.e. trachea and bronchi).  Upon expiration, alveolar gas involved in gas exchange that contains higher concentrations of CO2 and lower concentrations of O2 mixes with gas within physiological deadspace that is made up of low concentrations of CO2 and high concentrations of O2 (essentially room air). At rest, the end-tidal-to-arterial PCO2 gradient (Pa-PETCO2; calculated as PaCO2-PETCO2) is typically non-existent or positive (20, 93, 127), however, it has also been reported as being negative at LA (87, 118) and HA (118). This gradient can be altered with changes in body position (14), aging (80), exercise (59), low breathing frequencies (56), and with CO2 administration (i.e. hypercapnia) (3, 93, 118).  During hypercapnic conditions with DEF, high concentrations of administered CO2 occupies physiological deadspace during inspiration and mixes with alveolar gas during expiration, inflating PETCO2 to a greater extent than PaCO2 (56, 93, 118).  The same effect can be observed with increases in apparatus deadspace (39), and when administering CO2 from a Douglas bag (87).  There is a vast amount of literature reporting CO2 gradients clinically, but there is less for experimental chemoreceptor respiratory reflex testing (17, 23, 130, 136).  In contrast, there are few reports on the end-tidal-to-arterial PO2 gradient (PET-PaO2; calculated as PETO2-PaO2) in the context of chemoreflex or vascular (e.g., cardiac, pulmonary, cerebral, etc) testing during DEF.  It is well established that the alveolar-to-arterial difference (AaDO2) is greater for O2 compared to CO2, primarily due to diffusion limitation and normal V̇/Q mismatching (30, 107).  This potentially means that chemosensitivity and vascular function tests that manipulate O2 to low levels could be potentially harmful in individuals with abnormally large PET-PaO2 gradients.   A proposed controversial mechanism suggested by Gurtner & Traystman (52) for negative gas gradients observed during steady-state hypercapnia is known as the ‘charged membrane hypothesis’.  It involves the generation of an electrical negative field near the capillary endothelial surface within the lung resulting in a dissociation of weak acids thus elevating local amounts of   9 [H+].  This increase in local [H+] shifts the equilibrium of the bicarbonate buffering relationship toward PCO2 (52) (see Illustration 1.3, pg 9).  However, this hypothesis is based on I) anesthetized dogs during an extreme level of hypercapnia that is not tolerable by healthy humans, and II) measured alveolar-to-arterial gradients, in contrast to end-tidal-to-arterial gradients.  The end-tidal-to-arterial gradient differs from the alveolar-to-arterial gradient since it involves the mixing of alveolar gas with physiological deadspace gas upon expiration.  Although the charged membrane hypothesis supports observed negative gradients during administered hypercapnia, it is better explained by a combination of deadspace mixing (alveolar and anatomical), V̇/Q mismatching, diffusion limitation, and intrapulmonary and/or intracardiac shunting (73, 108).   Illustration 1.3: Diagram of the ‘Charged Membrane Hypothesis’ proposed by Gurtner et al. (1979). Due to a negatively charged endothelium weak acids (i.e. carbonic acid) dissociate, increasing the [H+] concentration, shifting the bicarbonate buffering system resulting in a local increase in CO2 within the pulmonary capillary blood to be exchanged with alveoli.    The issue of a discrepancy between end-tidal and arterial blood gases was addressed in the early 2000’s when a new forcing system called the “RespirAct” was innovated that tried to account for this gradient.  The RespirAct™, considered a sequential gas delivery system rather than a DEF, was developed by Thornhill Research Inc. (106) and is a compact system that can effectively control end-tidal gases without using as much gas as the Robbins DEF system (94).  The RespirAct™ also uses specific mixed gases in a fashion where it is impossible for 100% N2 to be administered to the subject, increasing subject safety.  One of the fundamental features of the RespirAct™ is that the end-tidal-to-arterial gas gradient is minimized by filling physiological deadspace with previously expired gas (considered gas exchange neutral) rather than “fresh” inspirate (56, 106), although this has only been validated in five subjects.  However, because the RespirAct™ aims to control anatomical and alveolar deadspace gas content, it works most effectively during paced ventilation, limiting its utility during exercise testing and during experiments investigating the control of breathing (44).  Regardless of this limitation, the RespirAct™ has proven to be effective in experiments related to vascular function by manipulating   10 end-tidal gases and has even been used in HA field research (85), but one of its principle weaknesses is that its software requires resting arterial blood gas parameters which are often input based upon the measurement of the end-tidal gases, rather than directly obtaining arterial blood samples (49, 50, 98, 119).  In saying this, resting PETCO2 is normally a good estimate of PaCO2 in healthy humans, but has been shown to over/underestimate PaCO2 as discussed throughout this section.  In contrast, PETO2 normally overestimates PaO2 at rest.    Factors affecting gas gradients End-tidal-to-arterial gas gradients are variable amongst healthy individuals.  Over the past few decades, we have gained substantial knowledge on how factors such as pulmonary diffusion limitation and V̇/Q inequality can directly affect the end-tidal-to-arterial gas gradients.  There have also been recent contributions on how anatomical shunting (i.e. intracardiac and intrapulmonary), along with environmental stresses like HA and exercise, may affect end-tidal-to-arterial gas gradients, but many questions regarding these factors and their effect on end-tidal-to-arterial gradients are yet to be answered. In addition, with the continuing advancements in science and technology, portable DEF systems will be more readily available for human physiology researchers interested in conducting their work in more unique locations (such as HA or during magnetic resonance imaging). For this reason, it is prudent to have a complete understanding of the capacity of these devices between different populations of people and in different environments.   1.5.1 Diffusion limitation The exchange of gases at the blood-gas barrier occurs through simple diffusion; and, diffusion capacity of the lung is often quantified through the magnitude of the alveolar-to-arterial gas gradient (34, 122, 124).  Any change in the alveolar-to-arterial gas gradient will contribute to the end-tidal-to-arterial gas gradient.  The lung’s diffusion capacity can be described by Fick’s law of diffusion, which states that the rate of transfer of a gas through a tissue is proportional to the tissue surface area (i.e. contact between alveolus and pulmonary capillary) and the difference in gas partial pressure between the alveoli and capillary (121, 122).  Rate of diffusion is inversely proportional to the thickness of the blood gas interface (122).  For this reason, we rarely see normal gas equilibration in patients with interstitial lung diseases where the tissue and space around the lung alveoli are thickened.  Failure of diffusional equilibration between capillaries and alveoli is   11 also seen in HA related pulmonary edema, a life-threatening condition where fluid accumulates in the lungs upon reaching altitudes typically above 2500 meters (2, 114).  There are two chemical properties of the gases being diffused which can directly affect the diffusion capacity of the lungs.  These are: 1) the solubility of the gas being exchanged, and 2) the molecular weight of the gas being exchanged; such that solubility is inversely related to molecular weight. For these reasons, the diffusion of CO2 is approximately 20 fold greater than O2, and CO2 equilibrates more quickly because it has a much higher solubility but only a modestly greater molecular weight (O2 - 32 g/mol; CO2 - 44.01 g/mol) (122).  Other factors that affect gas diffusion equilibration within the lung are 1) the rate of reaction of gas binding to haemoglobin, 2) haemoglobin’s capacity to carry the gas, and 3) transit time of red blood cells within the pulmonary circulation (124). On average in healthy humans, pulmonary blood spends about 0.75 seconds in the capillary at rest, which is plenty of time to reach alveolar-to-arterial gas equilibration as red blood cells require only about 0.25 seconds for alveolar gas to become fully equilibrated with arterial blood (122, 124). However, failure of equilibration can be seen during exercise at very high cardiac outputs where a large reduction in transit time results in only partial equilibration in the lung, and nearly everyone fails to equilibrate at HA when exercising (124, 126).   1.5.2 Ventilation-perfusion mismatch In the early 1900’s it was assumed that the V̇/Q of the lung was uniformly distributed, resulting in perfect pulmonary gas exchange efficiency (51).  Ventilation and perfusion matching of the lung is essential for efficient gas exchange, however, ventilation and blood flow are often mismatched throughout the lung, leading to impaired gas exchange in some regions of the lung.  The V̇/Q distribution of the lung is generally governed by the following three principles: I) the hydrostatic pressure gradient induced by gravity (i.e. changes in body position), II) the local interaction between alveolar and vascular pressure, and III) the geometry of the vascular tree (varies between individuals) (51). The V̇/Q ratio defines how well a region of the lung’s ventilation is matched to its perfusion.  V̇ defines the ventilation of the lung unit of interest, while Q̇ defines the blood flow to those lung units.  A V̇/Q ratio of 1 (i.e. V̇ = 1 l/min and Q̇ = 1 l/min) indicates the region of the lung is perfectly matched for ideal gas exchange.  Departure from V̇/Q uniformity can occur in the top and bottom of the lung, and is commonly due to changes in gravitational stress on the lung.  However, gas exchange is still occurring in regions of low and high V̇/Q but pulmonary gas   12 exchange efficiency may be hindered.  At one extreme, where there is ventilation but no blood flow to alveoli (V̇/Q = ∞), gas exchange cannot take place.  This is called alveolar deadspace.  At the other extreme, where there is no ventilation and alveolar perfusion is high (V̇/Q = 0), pulmonary gas exchange cannot take place and this region is typically termed a “shunt”.  Although V̇/Q inequality is present at rest, there are specific scenarios where V̇/Q inequality can change such as: body posture (24, 60), cardiopulmonary disease (96, 125), exercise (123), hypoxia (113), and hypercapnia (10, 12, 64, 102).  Ultimately, V̇/Q inequality can lead to an alveolar-to-arterial gradient due to pulmonary blood shunting, additionally resulting in an end-tidal-to-arterial gradient (eg. increasing the Pa-PETCO2 gradient during hypercapnia). The physiological regulatory mechanisms behind V̇/Q matching are unclear; however, pulmonary blood gases play an active role in V̇/Q matching, and hypoxic pulmonary vasoconstriction (HPV) is one important mechanism aimed at improving V̇/Q matching throughout the lung (51, 113, 122).  In response to alveolar hypoxia, HPV acts to redistribute blood from poorly ventilated (poorly oxygenated) to highly ventilated regions in the lung, in order to improve V̇/Q matching heterogeneously, however, this also potentially increases alveolar deadspace (67).  A change in alveolar PCO2 also seems to be a potential regulator of regional V̇/Q matching.  The effect alveolar carbon dioxide has on the pulmonary vasculature is in opposition to alveolar PO2, that is, when alveolar PCO2 is low (i.e. hypocapnia) the smooth muscles within the pulmonary vasculature relax, increasing blood flow, and when alveolar PCO2 is high (i.e. hypercapnia) the smooth muscles within the pulmonary vasculature constrict, decreasing pulmonary blood flow to alveoli (11, 13, 32, 33).   1.5.3 Intracardiac and intrapulmonary shunting. Within the circulatory system there are two main pathways, which could help to acutely drop pulmonary pressure by providing a low resistance pathway for blood.  However, these pathways also result in some blood bypassing the gas exchange sites of the lung, resulting in a reduction in gas exchange efficiency.  One of these pathways is called an intracardiac shunt. There are several different intracardiac shunt pathways, but the most common is the right-to-left shunt pathway through a patent foramen ovale (PFO).  The PFO is a small flap-like opening in the atrial septum, which provides a direct pathway for blood between the right and left atria important in the fetal circulation.  However, with birth and the commencement of eupnea the pressure drop between the pulmonary and systemic circulation normally allows the flap to close and subsequently fuse.  In   13 the general population, a PFO is detected in as many as 35-38% of the general population (38, 78, 133).   The second low-resistance pathway enabling blood to bypass the blood-gas interface is through intrapulmonary arteriovenous anastomoses (IPAVA), which are located within the pulmonary vasculature.  Intrapulmonary arteriovenous anastomoses emerge from small and medium-sized muscular arteries and open up into the venous side of the alveolar septal capillaries (see Illustration 1.4, pg 13) (109).  As a result, IPAVA form a bypass between the arterial and venous pulmonary circulation.  Blood flow through IPAVA is detected at rest in approximately 30% of healthy humans who do not have a concurrent PFO at sea-level, (36, 61, 109) but the prevalence increases to near 100% in healthy humans during exercise and with acute exposure to hypoxia (55, 70, 74). At HA, having a right-to-left shunt pathway, such as, a PFO or IPAVA, could potentially worsen hypoxemia and lead to greater pulmonary hypertension (21).  In this case, alveolar hypoxia would lead to HPV and increased pulmonary artery pressure. In turn, this could further worsen any right-to-left shunt, increase arterial hypoxemia and decrease venous O2 ultimately leading to greater alveolar hypoxia and further enhanced pulmonary hypertension (18, 72). This vicious circle could hinder success at HA by leading to pulmonary hypertension and edema (7, 22).   Illustration 1.4: A schematic representation of blood flow through an intrapulmonary shunt. The second low-resistance pathway enabling blood to bypass the blood-gas interface is through IPAVA’s, which are located within the pulmonary vasculature. As a result they form a bypass between the arterial and venous pulmonary circulation, potentially resulting in a significantly larger difference between end-tidal gases and arterial blood gases. Calculated PaCO2 and PaO2   14 was determined from the following calculation: Q̇s/Q̇t = (CcO2 – CaO2)/(CcO2 – CvO2). The arterial/venous oxygen content difference was assumed at 4.5 ml.dl-1, and haemoglobin concentration at 14.9 g.dl-1. Q̇s/Q̇t, shunt fraction; CcO2, content of oxygen in pulmonary end-capillary blood, CaO2, content of oxygen in arterial blood; CvO2, content of oxygen in venous blood. The PaCO2 was calculated based upon a 5% shunt fraction using the same formula, but CO2 values in place of O2 values.   1.5.4 High-altitude The alveolar-to-arterial gradient has been well documented at HA.  However, very little is known on the end-tidal-to-arterial gas gradient at HA which is particularly important in the context of end-tidal gas control while using DEF.  Due to the compact nature of our DEF system, it was used on a HA research expedition to 5050m at the Ev-K2-CNR pyramid laboratory (118) (see manuscript in Appendix A, pg 100).  To our knowledge, this was the highest elevation an end-tidal gas control device has been used (85), and the only study to quantify end-tidal-to-arterial gas gradients during commonly used CO2 and O2 protocols at such extreme altitude.  Given the recent advancements in DEF forcing systems (65, 85, 106), making them more portable and cost efficient, it is important to consider the potential limitations of their use.  Specifically, whether end-tidal gases accurately reflect arterial blood gases (the true stimulus for HCVR and CVR) in these extreme altitudes for both O2 and CO2.  Surprisingly, there is little literature available with respect to the PET-PaO2 gradient during DEF at LA and HA.  Although experiments investigating the AaDO2 between LA and HA exist, it is difficult to assess the AaDO2 during DEF due to the breath-by-breath fluctuations in inspired fractions of O2 and CO2 making the calculation of metabolic rate (i.e. respiratory exchange ratio) difficult. This necessitates the study of the end-tidal-to-arterial gradients in its place when using DEF. However, literature regarding the AaDO2 is inconsistent suggesting that the AaDO2 can widen or narrow following ascent to HA at rest (67, 91, 115), and has been shown to differ between PFO+ and PFO- participants (37).   Our research group quantified the PET-PaO2 gradient at rest at LA and at HA. Theoretically, this gradient would become smaller at HA due to the decreased difference between atmospheric O2 and venous blood O2 returning to the lung to participate in gas exchange.  Interestingly enough we found no difference in the PET-PaO2 gradient at rest between LA and HA, suggesting that diffusion capacity was hindered at HA (118). The Pa-PETCO2 gradient was also quantified by our research group between LA and HA during an isooxic CO2 reactivity test. The main findings were as follows: I) The Pa-PETCO2 gradient was negative at rest (PETCO2 overestimating PaCO2), II) The Pa-PETCO2 gradient was greater (~two-fold) at HA compared to LA, and III) The Pa-PETCO2 gradient was greater in magnitude during hypercapnia compared to hypocapnia.  The widening of   15 the Pa-PETCO2 gradient between LA and HA could potentially be the result of a diffusion limitation from subclinical mild HA pulmonary oedema (114).  The most likely explanation for the differences in the Pa-PETCO2 gradient between LA and HA during eupneic breathing is due to increased alveolar deadspace (67).  Another explanation for this phenomenon is the heightened ventilatory response that occurs with HA-hypoxia, which necessitates an increased FICO2 delivery from our DEF system in order to maintain a given PETCO2. Ultimately, this effect would increase the FICO2 trapped in physiological and apparatus deadspace and thereby inflate the measured PETCO2.  In addition to chronic hypoxia leading to an increase in alveolar deadspace, there also is evidence demonstrating that administered CO2 can cause small airway dilation and pulmonary vasoconstriction augmenting the HPV response, potentially further increasing physiological deadspace during hypercapnia which could explain the Pa-PETCO2 gradient differences between hypocapnia and hypercapnia (10, 12, 64, 102), in addition to the unchanged PET-PaO2 gradient seen between LA and HA.    Dynamic end-tidal forcing vs. the rebreathing technique Dynamic end-tidal forcing is a technique highly coveted by human physiology researchers due to its ability to accurately control end-tidal gases on a breath-by-breath basis.  However, there are other methods that are used that assess CVR and HCVR, such as the rebreathing technique (41, 54, 104, 105).  The rebreathing method as a means of increasing PaCO2 is beneficial for the following reasons: I) it is inexpensive and usually easy to conduct, II) due to the linear relationship within the HCVR, HYPO-CVR to CO2, and HYPER-CVR to CO2, linear regression can be used to quantify these reactivity measures (41, 54, 104, 105), and III) The gradients between arterial, venous, and tissue compartments are eliminated, thus the end-tidal-to-arterial gradient is substantially minimized (3, 104).  The latter benefit being arguably the most important; however, the rebreathing method seems to greatly underestimate CVR to CO2 compared to steady-state testing, making it difficult to compare to studies that use more sophisticated methods such as DEF (3).  Dynamic end-tidal forcing allows for the assessment of HCVR and CVR to CO2 during controlled steps of CO2, and if desired, in the background of different levels of oxygen (i.e hypoxia, normoxia, or hyperxoia) (118).  This likely makes measurements of cerebral blood flow through both hypocapnia and hypercapnia easier when using peripheral ultrasound, as scanning extracranial vessels continuously has proven to be a very difficult task, especially during periods of high V̇E that can be achieved during hypercapnic steps.  However, during DEF end-tidal-to-  16 arterial gradients are present (see section 1.2.1, pg 5), which have previously been demonstrated to affect the measurement of CVR and HCVR profiles during normoxia (87) at different levels of FICO2 administered with a Douglas bag.  However, it is unclear the extent to which CVR and HCVR profiles are affected by the end-tidal-to-arterial gradient during two DEF protocols: I) normoxic hypo/hypercapnia protocol and, II) hypoxic hypo/hypercapnia protocol.   End-tidal vs arterial CO2? Cerebrovascular reactivity to CO2 is almost always quantified by plotting cerebral blood flow against either PETCO2 or PaCO2. As described in section 1.3.1 and 1.3.2, quantifying CVR and HCVR to PETCO2 during DEF may potentially lead to data misinterpretation as PETCO2 does not uniformly reflect PaCO2.  As presented in Illustration 1.5 (pg 16), the HCVR can be potentially misinterpreted if indexed against PETCO2 as opposed to PaCO2, as PETCO2 overestimates PaCO2 in the hypercapnic range (118).  This same effect can be observed with CVR to CO2 (see Illustration 1.6, pg 17) during hypercapnia.  However, during hypocapnia, PETCO2 seems to be a good estimate of PaCO2, avoiding this potential confound (87, 118).    Illustration 1.5: Conceptual display of the potential effect that the end-tidal-to-arterial CO2 gradient has on hypercapnic HCVR. Panel A, represents the HCVR profiles assuming that PETCO2 is an accurate measure of PaCO2. Panel B, represents the actual observed HCVR profiles, as HCVR is greater when indexed against PaCO2 rather than PETCO2.    17  Illustration 1.6: Conceptual illustration of the potential effect the Pa-PETCO2 gradient has on CVR to CO2.  Panel A, represents the CVR profiles assuming that PETCO2 is an accurate measure of PaCO2 in hypocapnic and hypercapnic ranges. Panel B, represents the actual observed CVR profiles, as CVR is greater when indexed against PaCO2 rather than PETCO2 in hypercapnia, but not hypocapnia.  For this reason, using the rebreathing method for CO2 stimulus is advantageous as the gradient between PETCO2 and PaCO2 is thought to be abolished.  However, having actual PaCO2 measurements during a CO2 reactivity test is beneficial regardless of whether the rebreathing or steady-state method is chosen.  Previous equations have been proposed for predicting PaCO2 during exercise (59), and during CO2 reactivity tests using different levels of FICO2 (87) and the RespirAct™ end-tidal control system (130). We recently derived our own arterial correction algorithms for CO2 for use with our custom DEF system at LA (Kelowna, BC; 344m) and HA (Ev-K2 pyramid laboratory, Nepal; 5050m) using the participant’s end-tidal gases (118).   Low-altitude: (1) PaCO2 = 0.363 + (0.958 * PETCO2)         r2 = 0.95   High-altitude: (2) PaCO2 = 0.143 + (0.861 * PETCO2)  r2 = 0.92   In addition to these correction algorithms, we have also derived correction equations that take into account the participant’s baseline end-tidal-to-arterial gas gradient.  These equations may be particularly advantageous as they may reduce variability in reactivity outcome measures, without having to insert arterial catheters.  However, the reliability of these correction algorithms   18 could be hindered by the ability to reliably obtain a resting baseline end-tidal-to-arterial measurement as subjects may hyperventilate during an arterial blood sample.   Low-altitude: (3) PaCO2 = 0.964 + (0.960 * PETCO2) + (0.331 * Baseline Pa-PETCO2)    r2 = 0.95 High-altitude (4) PaCO2 = 2.899 + (0.861 * PETCO2) + (0.675 * Baseline Pa-PETCO2)    r2 = 0.97   Research questions and hypothesis  (I) Will the end-tidal-to-arterial CO2 gradient be widened during a CO2 reactivity test in the background of acute hypoxia compared with normoxia?  We hypothesize that due to the increase in ventilation and potential increase in alveolar deadspace during acute hypoxia, the end-tidal-to-arterial CO2 gradient will be greater during the hypoxic protocol, due to an increased CO2 administration from our dynamic end-tidal forcing system.   (II) Is the end-tidal-to-arterial gas gradient similar between people that have a patent foramen ovale (PFO+) detectable by agitated saline contrast echocardiography compared to those who do not (PFO-)? We hypothesised that the end-tidal-to-arterial CO2 gradient will be unaffected due to the small arterial-to-venous carbon dioxide gradient, while the end-tidal-to-arterial O2 gradient will be larger in PFO+ compared to PFO- participants due to venous admixture.   (III) Are calculated cerebrovascular reactivity and ventilatory reactivity to CO2 profiles attenuated by using PETCO2 instead of PaCO2 when calculating reactivity to a dynamic end-tidal forcing CO2 test and can this difference be minimized using our PaCO2 prediction equation? We hypothesized that cerebrovascular reactivity and ventilatory reactivity to CO2 will be significantly underestimated when calculating reactivity using PETCO2 compared to PaCO2 since PETCO2 overestimates PaCO2 during hypercapnia.  In   19 addition, there will be no difference between cerebrovascular reactivity and ventilatory reactivity when using the predicted PaCO2.  (IV) Does our previously derived correction algorithm accurately predict PaCO2 in a larger independent sample during dynamic end-tidal forcing? Our correction algorithm will minimize the difference between our predicted PaCO2 and our measured PaCO2, by using PETCO2 and the individual’s baseline end-tidal-to-arterial CO2 gradient in our linear regression algorithm.    20 Chapter 2: Materials and Methods This chapter consists of an overview of the experimental methods used to systematically answer our research questions.  Details are provided on the recruited participants, prescreening measurements, the experimental protocols and procedures, collected dependent variables, and data and statistical analyses.    Ethical approval   All experimental procedures and protocols were reviewed and approved by the Clinical Research Ethics Board at the University of British Columbia and conforms to the Declaration of Helsinki.  All participants provided written informed consent prior to participation in this study (see Appendix B, pg 140).     Participants All experimental sessions were conducted in the Cardiopulmonary Laboratory for Experimental and Applied Physiology at the University of British Columbia’s Okanagan Campus.  Recruited participants (n=18) were required to fill out a health history questionnaire to ensure normal pulmonary and cardiovascular health.  Each participant was pre-screened on two separate days prior to the experimental protocol.  The first pre-screening visit involved an agitated saline contrast echocardiography protocol (see section 2.3.1, pg 21), to detect the presence/absence of a PFO at rest and with provocative manoeuvres.  The second pre-screening visit consisted of a battery of pulmonary function tests including forced vital capacity, slow vital capacity, single breath carbon monoxide test for diffusion capacity, and total lung capacity using a whole body plethysmograph and spirometery system (see section 2.3.2, pg 21).  Participants had to be between the ages of 18-45 years and were excluded from the study if they had abnormal lung function (defined as showing no signs of large or small airway obstruction; and a FEV1/FVC ratio <0.75).  Participants were excluded if they were obese (body mass index >30 kg/m2), pregnant, and hypertensive (i.e. systolic >140 mmHg, diastolic >90 mmHg).  Participants included in mean data analysis (n=15; 3 female) were non-smokers, had no previous history of cardiovascular, cerebrovascular or respiratory diseases, and were not taking any medications during testing (except oral contraceptives).  Three of the 18 subjects screened were excluded from mean data analysis for the following reasons: (1) arterial catheter complications (n=2), and (2) gas analyzer failure (n=1).      21  Prescreening Prior to experimentation, each participant went through the following pre-screening protocols and procedures on two separate occasions.   2.3.1 Patent foramen ovale screening In order to determine the influence of an intracardiac shunt on the end-tidal to arterial gradients all subjects were screened for a PFO by agitated saline contrast echocardiography technique at rest and during the release of a Valsalva maneuver while the patient lie in the supine left lateral decubitus position, as previously described (37, 48).  Briefly, a 20-gauge intravenous catheter was placed into the antecubital fossa of each subject and flushed with 0.9% saline.  A three-way stopcock and two 10 mL syringes were connected.  One syringe contained 0.5 mL of air and the other syringe contained 4 mL of saline.  To create the contrast microbubbles, the saline solution was rapidly flushed back and forth for at least 10 seconds from one syringe to the other.  Immediately following agitation, the saline microbubbles were rapidly injected while recording an apical four-chamber image of the heart by echocardiography using a commercially available ultrasound system (Vivid E9, GE, Fairfield, CT, USA) and the same sonographer.  The ultrasound system uses a broadband M5S 5 MHz probe for 2D imaging.  Images were captured and saved for offline analysis using commercially available software (EchoPAC v.13, GE, Fairfield, CT, USA).  Foreshortening of apical chamber images was avoided by seeking either palpitation of the apex on the chest, or the largest long axis dimension within two consecutive rib spaces.  Immediately following injection, contrast microbubbles are observed in the right side of the heart.  Contrast microbubbles appearing within the left atrium at rest or during a provocative maneuver (release from a Valsalva maneuver) within four cardiac cycles suggests the presence of a PFO (42).  The Valsalva maneuver is performed by requesting the participant to forcefully exhale against a closed glottis, upon release of the Valsalva maneuver a subsequent increase in venous return, thus right atrial pressure, may forcefully open a PFO that is not normally patent at rest.  If the presence/absence of a PFO remained unclear after the first agitated saline injection, a repeated injection was permitted once all microbubbles were absent from the initial injection.  2.3.2 Pulmonary function testing In order to determine whether our participants had normal healthy lung function, we conducted a forced vital capacity (FVC) test to measure lung function, a vital capacity (VC) and inspiratory capacity (IC) maneuver to measure lung volumes, and a single breath carbon monoxide test to   22 quantify diffusion capacity (DLCO) on each individual. All testing procedures were conducted in agreement with the American Thoracic Society and European Respiratory Society’s joint guidelines (1, 43, 76, 79).  For each of these tests, participants sat within the body plethysmography box (V6200, Vmax Sensormedics, Yorba Linda, CA, USA) with a rigid upright posture and their feet flat on the ground, whilst breathing through a spirometer and bacteriological filter with nose clamped.  All pulmonary function measurements were compared against population-based predictions.   Experimental protocol This study was conducted in two parts: (I) normoxia CO2 reactivity protocols (NX1 and NX2) and (II) hypoxia CO2 reactivity protocols (HX1 and HX2).  Normoxia protocols were always conducted first to avoid any potential confounds involving carry-over effects of sympathetic nervous system activation associated with exposure to acute hypoxia (135).  In addition, protocols within normoxia (NX1 and NX2) and hypoxia (HX1 and HX2) were counter-balanced (i.e. protocols were randomly assigned in such a fashion to ensure an equal distribution of protocol order).  Prior to each experiment, all participants abstained from exercise and alcohol for 24h, and caffeine for 12h.  Testing was conducted while participants lay in the supine position on a hospital bed.  Experimental ‘baseline’ measurements took place during a period of quiet eupneic breathing.  2.4.1 Normoxia CO2 reactivity protocols This protocol was selected because it is often used to assess CVR to CO2 (118, 130).  In addition, the hypercapnic portion of the protocol can be used to assess HCVR (54).  Subjects were instrumented and allowed to breathe quietly through a mouthpiece with nose clamp.  The protocol consisted of two baseline periods: baseline 1 (room air breathing; AB) and baseline 2 (resting end-tidal control by DEF; AB-DEF).  Baseline 1 involved 10-minutes of eupnea through our breathing apparatus (apparatus deadspace = ~250 ml).  Immediately following AB (time = 10 minutes) AB-DEF commenced when the DEF system was connected to the inspiratory port of the breathing apparatus, and PETCO2 and PETO2 were controlled at AB baseline values for five-minutes. Following AB-DEF, PETO2 was maintained at AB values (isooxia) while PETCO2 was controlled in a stepwise fashion at -8, -4, 0, +4, and +8 mmHg from individual baseline values.  Targeted PETCO2 in the hypocapnic range was achieved through active hyperventilation and controlled using the DEF system.  Each step change in PETCO2 lasted for approximately 4-minutes after steady-state was achieved and an arterial blood sample and Q̇ICA was collected during the final   23 minute of each stage. The protocol was then repeated after 20-minutes of rest after adjusting the target PETCO2 using a previously derived equation for predicting the PaCO2 (118).  In other words, the PETCO2 target values were adjusted to achieve the desired PaCO2 using the following equation:  (5) PETCO2 = ((PaCO2 – 0.964) – (0.331 * Baseline Pa-PETCO2)) / 0.960   2.4.2 Hypoxia CO2 reactivity protocols This protocol was selected because it can be used to assess the CVR to CO2 in the background of hypoxia (118, 130).  In addition, the hypercapnic portion of the protocol can be used to assess the HCVR in the background of hypoxia (HX-HCVR).  Subjects were instrumented and allowed to relax and breathe normally through a mouthpiece with nose clamp.  The protocol consisted of two baseline periods: baseline 1 (AB) and baseline 2 (AB-DEF).  Baseline 1 involved 10-minutes of eupneic breathing.  Immediately following AB, HX-DEF commenced once our DEF system was connected to the inspiratory port of the breathing apparatus, and PETO2 was controlled at 50 mmHg and PETCO2 was maintained isocapnic (based upon AB values).  Baseline 2 (HX-DEF) in HX trials was longer (10-minutes vs 5-minutes) than baseline 2 (AB-DEF) in NX trials to minimize the influence of the peak hypoxic ventilatory response (92) and to allow the pulmonary vascular pressure response to stabilize (116).  Following HX-DEF, PETO2 was maintained at 50 mmHg while PETCO2 was controlled in a stepwise fashion at -8, -4, 0, +4, and +8 mmHg from individual baseline values.  Targeted PETCO2 in the hypocapnic range was achieved through active hyperventilation and controlled by DEF.  Each step change in PETCO2 lasted for approximately 4-minutes after steady-state was achieved and an arterial blood sample and Q̇ICA was collected during the final minute of each stage.  HX1 and HX2 protocols were identical with the exception that during the HX2 protocol our previously derived correction algorithm (equation 5) was used to correct for PaCO2.    Outcome measurements The following section elaborates on the participant instrumentation procedure, along with detailed information on how we collected respiratory data and used DEF to effectively control end-tidal gases.       24 2.5.1 Participant instrumentation Upon arrival on the experimental day, participants sat down in a semi-recumbent position for 10-minutes.  An arterial catheter was inserted into the left radial artery by an anesthesiologist under ultrasound guidance.  The participant was then instructed to lay supine on a hospital bed where they were instrumented with a lead-II electrocardiogram (ECG) to measure heart rate (HR), nasopharyngeal temperature probe (RET-1, Physitemp Instruments, Clifton, NJ, USA), automated blood pressure (HEM-775CAN, Omron Healthcare, Cannockburn, IL, USA), and TCD headpiece with the ultrasound probe insonating the right MCA (PMD150B, Spencer Technologies, Redmond, WA, USA).  The participant was then instructed to relax and breathe normally for approximately 30 minutes to ensure body positional change in blood volume distribution was complete.   2.5.2 Respiratory measurements   Respiratory measurements have been previously described elsewhere (118). All respiratory parameters were acquired at 200 Hz using an analog-to-digital converter (Powerlab/16SP ML 880; ADInstruments, Colorado Springs, CO, USA) interfaced with a personal computer.  Lab acquisition software was used to collect and analyze ventilatory and cardiovascular variables (LabChart V7.1, ADInstruments, Colorado Springs, CO, USA).  Subjects breathed through a mouthpiece (with nose clamp), bacteriological filter, and a two-way non-rebreathing valve (2600 series, Hans Rudolph, Shawnee, KS, USA).  Respired gas pressures were sampled near the mouth, dried with nafion tubing and desiccant, and analyzed for PETO2 and PETCO2 (ML206, ADinstruments, Colorado Springs, CO, USA).  Inspiratory and expiratory VT were determined using an integral of the respiratory flow signal, breathing frequency (FB) as the amount of breaths taken per minute, and V̇E being the product of these two respiratory measurements.  Gas analyzers were calibrated prior to experimentation with gases of known concentration using the same sample line used in the experiment.  Gas analyzers were checked before and following each protocol (i.e. NX1 and HX1) to ensure accurate respiratory measurements.  Measured PO2 and PCO2 were time-corrected for gas analyzer sample delay and the values corresponding to the end of expiration (i.e. when respiratory flow crossed zero in the positive to negative direction) were identified as the PETO2 and PETCO2.  Respiratory flow was measured near the mouth using a pneumotachograph (HR 800L, HansRudolph, Shawnee, KS, USA) and a differential pressure transducer (ML141, ADinstruments, Colorado Springs, CO, USA), which was calibrated daily with a 3-liter syringe.   25 End-tidal gases were corrected for water vapour pressure.  A clinical temperature probe (RET-1, Physitemp Instruments, Clifton, NJ, USA) connected to a T-type pod (ML312, ADinstruments, Colorado Springs, CO, USA) was calibrated daily, and inserted approximately 10 cm into nasopharyngeal cavity (71).  Nasopharyngeal temperature has been validated as a reliable measurement of core body temperature (95). Using the participants’ nasopharyngeal temperature, water vapour pressure was instantaneously calculated using the Antoine equation:  (6) Log10P = A – (B/(C+T)  Where P is the calculated water vapour pressure (mmHg), A is a constant 8.07131 for water between 1 - 100C, B is a constant 1730.63 for water between 1 - 100C, C is a constant 233.426 for water between 1 - 100C, and T (C) is the measured core body temperature.  Rearranging this formula, water vapour was calculated in real time as:  (7) Water Vapour Pressure = 10^8.07131-((1730.6)/(233.426+BT))  Calculated water vapour pressure was subtracted from the daily atmospheric pressure in order to calculate the partial pressure of respired gases for each individual.   2.5.3 End-tidal forcing PETO2 and PETCO2 were controlled by a portable DEF system as previously described (118).  Briefly, this system uses independent gas solenoid valves for O2, CO2, and N2 and controls the volume of each gas being delivered to the inspiratory reservoir through a mixing and humidification chamber.  PETO2, PETCO2, VT, FB, and V̇E were determined for each breath online using custom software (Airforce V 4.8) programmed in Labview (Version 13.0, National Instruments, Austin, TX, USA).  Using feedback information regarding PETO2, PETCO2, inspired VT, and expired VT the DEF system adjusts the inspirate to bring end-tidal gas to the desired target values.  Feed-forward control of the inspirate is based on estimates of baseline metabolic O2 consumption and CO2 production and employs the alveolar gas equation to determine the required FICO2 and FIO2.  While feedback control is accomplished using a proportional and integral error reduction control system.  This system has been used previously to control end-tidal gases during physiological stressors (1, 15, 36).  End-tidal steady state was achieved when end-tidal gases were   26 within 1 mmHg of the desired end-tidal target for at least 3 consecutive breaths and is normally achieved within 30s.   Cerebral blood flow measurements Below is a detailed description of the methodology used to measure intracranial and extracranial blood flow to the brain. This includes information on the equipment and techniques used to ensure adequate measures of cerebral blood flow.   2.6.1 Intracranial blood flow Unilateral transcranial Doppler ultrasound (TCD) was employed to measure the intracranial cerebral blood flow velocity within the right MCA (MCAv) using previously described search techniques (129) (PMD150B, Spencer Technologies, Redmond, WA, USA).  The transcranial Doppler ultrasound system uses a 2 MHz ultrasound probe connected to a headset, which emit sound waves through the temporal window and registers the frequency of the sound reflected from moving red blood cells within the vessel of interest.  The resulting frequency shift (i.e. Doppler shift) can provide the instantaneous velocity of the moving red blood cells.  Since cerebral blood flow is affected by vessel diameter, it is important for diameter to remain constant in order for blood velocity to reflect the underlying changes in cerebral blood flow.  Cerebral blood flow velocity (via TCD) has been thought to provide an accurate measurement of cerebral blood flow during hypo- and hyper-capnic (-13 mmHg and +9 mmHg PETCO2 from baseline, respectively) conditions (4, 99, 130), however, recent evidence suggests MCA diameter does change within this range (27, 120), suggesting that TCD may not be a useful tool for measuring cerebral blood flow during a CO2 reactivity test.  The same experienced sonographer insonated the right MCA for all subjects.  Mean cerebral blood flow velocity was calculated from the envelope of the velocity tracing.  2.6.2 Extracranial blood flow Continuous diameter and blood flow velocity recordings of the right ICA was obtained using a 10 MHz linear array probe attached to a high-resolution ultrasound machine (Terason 3200, Teratech, Burlington, MA, USA).  Blood flow velocity through the ICA and vessel diameter was measured at least 2 cm from the carotid bifurcation, whilst ensuring there was no evidence of turbulent or retrograde flow.  Simultaneous diameter and blood flow velocity measurements were collected and provide the input data required to calculate blood flow through these vessels assuming that the   27 vessel is circular.  The same experienced sonographer insonated the right ICA for all subjects.  Vessel image was optimized by insonating the vessel at an ideal angle (60 degrees) through two methods: 1) changing the angle of the incident beam, and 2) angling the ultrasound probe using “heel and toe” maneuvers.  Ultrasound recordings were screen captured and saved for offline analysis.  Blood flow analysis of the ICA was performed using edge-detection software, which allows for the integration of synchronous diameter and velocity measurements to determine the mean beat-to-beat flow at 30 Hz independent of investigator bias (132).  Mean blood flow was determined as half the time-averaged maximum velocity (40), multiplied by the cross-sectional luminal area for a minimum of 12 cardiac cycles.   Arterial blood sampling and blood pressure    After local anaesthesia (2% lidocaine), a 20-gauge catheter (Radial artery catheter, Arrow International, Reading, PA, USA) was placed transcutaneously into the radial artery using ultrasound guidance and a modified Seldinger technique (118).  The catheter was connected to a commercially available arterial blood sampling kit (VAMP Adult, Edwards Lifescience, Irvine, CA, USA), allowing for repeated sampling and flushing with 0.9% saline and measuring beat-by-beat arterial blood pressure.  Before sampling, the deadspace volume was withdrawn and then an arterial sample (3 ml) was carefully collected into pre-heparanized syringes (safePICO syringes, Radiometer, Copenhagen, Denmark).  Air bubbles were immediately evacuated from the syringe, the syringe was capped, and blood gas analysis was performed within 30-seconds of sampling with a gas analyzer (ABL90 FLEX, Radiometer, Copenhagen, Denmark).  The gas analyzer aspirates the blood into a chamber containing ion selective electrodes, specific to our measured variables.  After each measurement, the blood is automatically expelled into an internal waste container. The blood gas analyzer was calibrated every 8 hours using manufacturer’s standard internal quality checks and external ampoule-based quality checks were routinely performed to confirm internal calibrations.  Reported variables that were calibrated and analyzed included: PaO2, PaCO2, pH, and arterial oxyhaemoglobin (SaO2).  Arterial blood was temperature corrected offline to nasopharyngeal temperature using previously derived constants and logarithmic equations as described previously by JW Severinghaus in 1966 (101) (see section 2.5.2, pg 24).    Data analysis The following subsections detail the specific methodology used in our data analysis.  This includes how we analyzed our cardiovascular, respiratory and arterial blood data.  In addition the   28 quantification of CVR and HCVR are provided, along with a scoring system used to subjectively quantify the magnitude of a PFO in participants that tested positive.   2.8.1 Cerebrovascular reactivity in normoxia and hypoxia We quantified relative HYPO- and HYPER-CVR to CO2 during both normoxia and hypoxia protocols by indexing MCAv and Q̇ICA against (I) PETCO2, (II) PaCO2, and (III) Pred-PaCO2.  Linear regression analysis was used to quantify slope, an index of reactivity, for each participant.  A Pearson correlation coefficient of >0.7 was considered linear, and any relationships below this were considered non-linear and omitted from analysis.  Mean HYPO- and HYPER-CVR to CO2 slopes indexed against PETCO2, PaCO2, and Pred-PaCO2 were compared between and within hypoxia and normoxia trials.  2.8.2 Hypercapnic ventilatory responses in normoxia and hypoxia We quantified absolute HCVR and Hx-HCVR by indexing V̇E against (I) PETCO2, (II) PaCO2, and (III) Pred-PaCO2 and used linear regression analysis, an index of reactivity, separately for each participant.  A Pearson correlation coefficient >0.7 was considered a linear response, and any relationships below this were considered non-linear and omitted from analysis.  Mean HCVR slopes indexed against PETCO2, PaCO2, and Pred-PaCO2 were compared within hypoxia and normoxia trials.   2.8.3 Intracardiac shunt scoring A scoring system has previously been established to determine the severity of blood flow through PFO based on the greatest density and spatial distribution of microbubbles in the left ventricle of a single cardiac cycle during the subsequent four cardiac cycles (75).  The 0–5 scoring system assigns: ‘0’ for no microbubbles; ‘1’ for 1–3 microbubbles; ‘2’ for 4–12 microbubbles; ‘3’ for greater than 12 microbubbles bolus; ‘4’ for greater than 12 microbubbles heterogeneously distributed; and a ‘5’ for greater than 12 microbubbles homogeneously distributed.  This scaling system is reproducible between independent blinded observers (70).  All agitated saline contrast echocardiograms were reviewed and scored by two individuals.   2.8.4 Sample size justification A sample size calculation was conducted based on our previous research (118) indicating that a 2.1 ± 0.5 mmHg (mean ± SE) difference in the end-tidal-to-arterial PCO2 gradient during a CO2   29 reactivity test at sea-level was present.  In order to determine a significant difference between the gradient quantified and the assumption with DEF that there is no gradient (i.e. Pa-PETCO2 = 0) we will need to study a minimum of five subjects.  However, as our research question also considers the impact of an intracradiac shunt on end-tidal-to-arterial gas gradients we have selected a sample size of 18.  Since a PFO is normally prevalent in 35-38% of the population (38, 78, 133) this sample size should be adequate to address our research questions.    2.8.5 Statistics All data were analyzed using SigmaStat V11.5 (Systat, Chicago, IL, USA).  During all CO2 reactivity protocols (NX1, NX2, HX1, and HX2), measurements were averaged over a 1-minute period at the end of each baseline in normoxia (i.e. AB and AB-DEF), hypoxia (AB and HX-DEF), and with each CO2 step (-8, -4, 0, +4, and +8 mmHg PETCO2), just prior to arterial blood sampling.    Two-way repeated measures analysis of variance was used to compare differences (P<0.05) in ventilatory, cardiovascular, cerebrovascular and arterial blood measurements for each protocol, including analysis between PFO+ and PFO- participants.  Two-way non-repeated measures analysis of variance was used to compare differences (P<0.05) in ventilatory, cardiovascular, cerebrovascular and arterial blood measurements between the normoxia and hypoxia CO2 reactivity protocols.  To determine differences (P<0.05) between each reactivity measure when indexed against PETCO2, PaCO2 and pred-PaCO2 a one-way repeated measures analysis of variance was used. When significant F-ratios were detected, all post-hoc comparisons were made using Tukey’s HSD.   End-tidal-to-arterial gradients for both O2 and CO2 were determined by comparing end-tidal and arterial gas values using a paired t-test, significance was set at P<0.05. Bland-Altman relationships were used to assess the agreement between end-tidal and arterial blood gases, along with demonstrating the reproducibility of the arterial blood gas machine used.  The 95% limits of agreement were calculated by using 1.96 x standard deviation (SD) of the bias (9).  HCVR, and hypo- and hyper-capnic relative ICA and MCAv reactivity to CO2 were quantified using linear regression analysis in both normoxia and hypoxia trials.  Multiple forward stepwise regression analysis was used to determine which variables could accurately predict arterial blood gases, based upon the current study data.  The following independent variables were included in regression analysis: V̇E, VT, FB, FIO2, FICO2, PETO2, PETCO2, the baseline PET-PaO2 gradient, and the baseline Pa-PETCO2 gradient.  Inclusion of   30 independent variables in the multiple forward stepwise regression model was determined with a P-value <0.05. All data are expressed as means  SEM.    31 Chapter 3: Results   Participants All participants (n=15) included in normoxia and hypoxia protocol data analysis had a mean  SEM age of 25.8  1.5 years, height of 174.2  2.1 cm, weight of 72.9  3.7 kg, body mass index of 23.9  0.9 kg/m2, and the right MCA was insonated at a depth of 5.3  1.1 cm.  Of the participants studied in the mean data analysis, eight tested positive for a PFO (two with a shunt score of ‘1’; six with a shunt score of ‘2’).  However, all PFO’s detected were only with release from Valsalva and suggest that none of our subjects had a resting PFO.  Error! Reference source not found. displays resting pulmonary function data for all participants. Participants were normotensive (systolic blood pressure = 116  2.2 mmHg, diastolic blood pressure = 68  2.4 mmHg), non-smokers, had no previous history of cardiovascular, cerebrovascular or respiratory diseases, and were not taking any medications during testing (except oral contraceptives).   Table 3.1: Pulmonary pre-screening data. Variable Mean  SEM  (% predicted)  Variable Mean  SEM  (% predicted) FVC (l) 5.3  0.2 (108  3) FRC (l) 3.0  0.1 (95  4) FEV1 (l) 4.2  0.2 (101  4) RV (l) 1.1  0.1 (80  6) FEV1/FEV 79.2  1.4 DLCO  (ml/min/mmHg) 31.9  1.6 (94  3) TLC (l) 6.5  0.3 (101  3) DLCO/VA (ml/min/mmHg/l) 5.2  0.2 (99  4) IC (l) 3.5  0.2 VA (l) 6.2  0.2 (96  2) Definition of abbreviations: DLCO, diffusion capacity of the lung for carbon monoxide transfer; DLCO/VA, DLCO corrected for alveolar volume; FEV1, forced expiratory volume in one second; FVC, forced vital capacity; FRC, functional residual capacity; IC, inspiratory capacity; RV, residual volume; TLC, total lung capacity; VA, alveolar volume.     32  Research Question One: end-tidal-to-arterial CO2 and O2 gradients  3.2.1 Normoxia protocols Of the 15 participants included in this study, one participant was completely excluded from the normoxic trial due to an arterial catheter failure in the middle of protocol (n=14).  3.2.1.1 NX1 CO2 protocol Table 3.2 displays ventilatory, cardiovascular, and cerebrovascular data for both NX1 and NX2 isooxic CO2 reactivity tests (see Error! Reference source not found., pg 34 for end-tidal gas profiles from a representative subject).  As expected, V̇E and VT, were elevated in hypocapnic and hypercapnic ranges compared to AB-DEF (DEF controlled baseline), while participants FB was elevated only at the +8 mmHg PETCO2 step.  The partial pressure of inspired carbon dioxide (PICO2) was less during AB and -8 mmHg PETCO2 step, while it was elevated during hypercapnia.  Heart rate and MAP were elevated in hypercapnia, but not hypocapnia, compared to AB-DEF. Absolute MCAv was 5.0  0.9% lower during AB compared to AB-DEF, and absolute MCAv and Q̇ICA were 20.9  1.1% and 27.0  1.7% lower at -8 PETCO2 mmHg, and 11.1  1.0% and 16.0  1.9% lower at -4 PETCO2 mmHg from AB-DEF, respectively.  During hypercapnia, absolute MCAv and Q̇ICA were 11.8  1.7% and 20.8  5.0% higher at +4 PETCO2 mmHg, and 30.8  1.8% and 53.7  6.0% higher at +8 PETCO2 mmHg from AB-DEF, respectively.   Table 3.3 displays end-tidal and arterial blood parameters for both NX1 and NX2 isooxic CO2 reactivity tests.  The PETCO2 was lower during AB compared to AB-DEF, while PETCO2 and PaCO2 were both lower during hypocapnia and elevated during hypercapnia. A positive Pa-PETCO2 gradient (i.e. PaCO2 > PETCO2) was present during AB, whilst a negative Pa-PETCO2 gradient (i.e. PETCO2 > PaCO2) was observed at +4 and +8 mmHg stages (see Figure 3.2, pg 34).  Throughout all stages of the NX1 CO2 reactivity protocol, PETO2 was not different from AB-DEF, however, PaO2 was elevated during +8 mmHg, compared to AB-DEF.  The PET-PaO2 gradient was present at all protocol stages, and was reduced by 1.5  0.9 mmHg compared to AB-DEF at +8 mmHg PETCO2 step (see Figure 3.3, pg 35).  SaO2 was increase by 0.8  0.2% during hypocapnia (-8 mmHg PETCO2), but was similar across all other stage compared to AB-DEF.  Finally, pH increased during hypocapnia and decreased during hypercapnia compared to AB-DEF.     33 3.2.1.2 NX2 CO2 protocol Table 3.2 displays ventilatory, cardiovascular, and cerebrovascular data for both NX1 and NX2 isooxic CO2 reactivity tests.  As expected, V̇E and VT, were elevated compared to AB-DEF (DEF controlled baseline) in hypo- and hyper-capnic ranges, while participants FB was only elevated at the +8 mmHg PETCO2 step.  The PICO2 was lower during AB and the -8 mmHg PETCO2 step, while it was elevated during hypercapnia.  Heart rate and MAP were elevated in hypercapnia, but not hypocapnia, compared to AB-DEF.  Absolute MCAv was 6.4  1.3% lower during AB compared to AB-DEF, and absolute MCAv and Q̇ICA were 22.3  1.2% and 27.6  2.6% lower at -8 PETCO2 mmHg, and 12.4  0.9% and 12.4  3.4% lower at -4 PETCO2 mmHg from AB-DEF, respectively. During hypercapnia, absolute MCAv and Q̇ICA were elevated by 13.4  1.6% and 26.4  4.7% at +4 PETCO2 mmHg, and elevated by 30.1  1.8% and 59.1  5.5% at +8 PETCO2 mmHg from AB-DEF, respectively.  Table 3.3 displays respiratory and arterial blood data for both NX1 and NX2 isooxic CO2 reactivity tests. The PETCO2 was lower during AB compared to AB-DEF, while PETCO2 and PaCO2 were both lower during hypocapnia and elevated during hypercapnia. A positive Pa-PETCO2 gradient was present during AB and a negative Pa-PETCO2 gradient was present during hypercapnia (see Figure 3.2 pg, 34).  Throughout all stages of the NX1 CO2 reactivity protocol, PETO2 was not different from AB-DEF, however, PaO2 was elevated during the +8 mmHg PETCO2 step compared to AB-DEF.  The PET-PaO2 gradient was present at all protocol stages, and was reduced by 4.7  1.0 mmHg compared to AB-DEF at +8 mmHg PETCO2 step (see Figure 3.3, pg 35).  SaO2 was increased by 0.8  0.2% during hypocapnia (-8 mmHg PETCO2), but was similar across all other stage compared to AB-DEF, and as expected, pH increased during hypocapnia and decreased during hypercapnia compared to AB-DEF. 3.2.1.3 NX1 vs. NX2 protocols Table 3.2 and Table 3.3 display ventilatory, cardiovascular, cerebrovascular, respiratory and arterial blood gas data during NX1 and NX2 CO2 reactivity protocols. As expected, all outcome variables were similar between protocols with the exception of PETCO2, PICO2, and PaCO2, which were elevated in NX2 compared to NX1. In addition, a main effect was detected with nasopharyngeal temperature, which was 0.14  0.04 C lower at +8 mmHg compared to AB-DEF.   34 Figure 3.1: End-tidal gas tracings from NX1 (A) and NX2 (B) CO2 reactivity trials for one representative subject.  Open circles represent PETO2; end-tidal partial pressure of oxygen; closed circles represent PETCO2; end-tidal partial pressure of carbon dioxide.  Each data point represents a 15-second average.  AB, air breathing baseline; AB-DEF, air-breathing baseline controlled using DEF.     Figure 3.2: Bland and Altman plot for agreement between PaCO2 and PETCO2 during NX1 (A) and NX2 (B) CO2 reactivity protocols.  (), -8 mmHg; (▼), -4 mmHg; (∆), 0 mmHg; (■), +4 mmHg; (□), +8 mmHg.  Dotted lines represent the 95% confidence intervals, and the continuous lines represent the mean bias.  Time (s)0 500 1000 1500 2000 2500PETCO2 and PETO2 (mmHg)0102030405060708090100110Time (s)0 500 1000 1500 2000 2500 30000102030405060708090100110A. B.AB AB-DEF-8-40+4+8AB AB-DEF-8-40+4+8A. B.PaCO2 and PETCO2 mean (mmHg)25 30 35 40 45 50 55 60PaCO2 - PETCO2 (mmHg)-4-2024PaCO2 and PETCO2 mean (mmHg)25 30 35 40 45 50 55 60-4-2024  35   Figure 3.3: Bland and Altman plot for agreement between PaO2 and PETO2 during NX1 (A) and NX2 (B) CO2 reactivity protocols.  (), -8 mmHg; (▼), -4 mmHg; (∆), 0 mmHg; (■), +4 mmHg; (□), +8 mmHg Dotted lines represent the 95% confidence intervals, and the continuous lines represent the mean bias. PaO2 and PETO2 mean (mmHg)80 85 90 95 100 105PETO2 - PaO2 (mmHg)-10-5051015PaO2 and PETO2 mean (mmHg)80 85 90 95 100 105-10-5051015A. B. 36  Table 3.2: Ventilatory cardiovascular, and cerebrovascular data during NX1 and NX2 CO2 reactivity tests.Variable Trial AB AB-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E NX1 12.4 ± 0.5 16.9 ± 1.0 22.2 ± 1.3* 20.6 ± 1.1 17.7 ± 0.8* 26.5 ± 2.2* 38.9 ± 3.3* (l/min) NX2 13.1 ± 0.5 16.5 ± 0.7 23.0 ± 1.5* 21.9 ± 1.2 18.6 ± 1.0* 27.4 ± 1.6* 41.1 ± 3.1*  NX1 vs NX2: P=0.102; Stage: P<0.001; Interaction: P=0.725 VT NX1 0.9 ± 0.0 1.3 ± 0.1 1.7 ± 0.2* 1.6 ± 0.2* 1.3 ± 0.1 1.7 ± 0.1* 2.2 ± 0.1* (l) NX2 0.9 ± 0.0  1.2 ± 0.1 1.8 ± 0.2* 1.8 ± 0.2* 1.3 ± 0.2 1.7 ± 0.1* 2.2 ± 0.1*  NX1 vs NX2: P=0.437; Stage: P<0.001; Interaction: P=0.252 FB NX1 14.3  1.0 14.2  1.1 15.8  1.7 15.8  2.1 14.8  1.1 17.3  2.0 18.9  2.0* (/min) NX2 15.2  0.8 14.2  0.8 15.3  2.1 15.4  2.3 15.7  1.3 17.4  1.6 20.0  1.9*  NX1 vs NX2: P=0.666; Stage: P=0.040; Interaction: P=0.745 PICO2 NX1 3.8 ± 0.7* 19.1 ± 2.1 12.3 ± 1.7* 19.1 ± 1.3 22.1 ± 1.2 33.1 ± 1.1* 40.7 ± 1.0* (mmHg) NX2 4.0 ± 0.7* 20.4 ± 1.3 14.6 ± 1.2* 20.8 ± 1.4  23.4 ± 1.4  34.7 ± 1.0* 41.5 ± 0.8*  NX1 vs NX2: P=0.043; Stage: P<0.001; Interaction: P=0.903 HR  NX1 61.3 ± 2.5 63.7 ± 2.5 66.8 ± 2.2 65.1 ± 1.9 64.1 ± 2.5 69.6 ± 2.9* 74.5 ± 2.7* (/min) NX2 61.8 ± 2.3  64.4 ± 2.6 65.7 ± 2.1 64.2 ± 2.2  63.5 ± 2.5 68.1 ± 2.7* 73.9 ± 2.8*  NX1 vs NX2: P=0.620; Stage: P<0.001; Interaction: P=0.529 MAP  NX1 83.9 ± 2.1 85.2 ± 1.8 85.2 ± 1.7 86.1 ± 1.7 87.4 ± 1.6 91.8 ± 2.2* 95.3 ± 2.1* (mmHg) NX2 84.8 ± 1.3  86.7 ± 1.4  85.8 ± 1.5 87.1 ± 1.5  87.5 ± 1.4 91.2 ± 1.5* 95.1 ± 1.6*  NX1 vs NX2: P=0.708; Stage: P<0.001; Interaction: P=0.570 MCAv  NX1 61.1 ± 2.7* 64.5 ± 3.1 50.8 ± 2.1* 57.1 ± 2.5* 65.6 ± 3.1  72.3 ± 3.9* 84.2 ± 3.9* (cm/s) NX2 62.5 ± 2.9* 67.0 ± 3.2 51.8 ± 2.2* 58.5 ± 2.7* 67.3 ± 3.2 76.1 ± 3.9* 86.9 ± 4.0*  NX1 vs NX2: P=0.123; Stage: P<0.001; Interaction: P=0.116 Q̇ICA  NX1 265.5 ± 15.8 277.7 ± 11.9 203.5 ± 12.1* 234.3 ± 13.8* 289.3 ± 14.7 334.9 ± 19.7* 428.7 ± 28.2* (ml/min) NX2 277.9 ± 10.2 274.3 ± 16 196.6 ± 9.7* 238.3 ± 14.5* 287.7 ± 12.4 344.9 ± 21.9* 434.4 ± 25.4*  NX1 vs NX2: P=0.525; Stage: P<0.001; Interaction: P=0.659 Nasal Temp NX1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1* (C) NX2 36.5 ± 0.1 36.6 ± 0.0 36.5 ± 0.1 36.5 ± 0.1 36.6 ± 0.1 36.5 ± 0.1 36.4 ± 0.1*  NX1 vs NX2 P=0.959; Stage: P=0.021; Interaction: P=0.243 Definition of Abbreviations: V̇E, minute ventilation; VT, tidal volume; FB, breathing frequency; PICO2, partial pressure of inspired carbon dioxide; HR, heart rate; MAP, mean arterial pressure; MCAv, middle cerebral artery blood flow velocity; Q̇ICA, internal carotid artery blood flow; Nasal Temp, nasal pharyngeal temperature.  *P<0.05, main effect of CO2 stage vs AB-DEF. Bolded NX2, main effect between NX1 and NX2, bolded value is significantly larger. Values are mean  SEM.   37     Table 3.3. Ventilatory and blood gas data during NX1 and NX2 CO2 reactivity tests.  Variable Trial AB AB-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg PETCO2 NX1 39.5 ± 0.7* 41.0 ± 0.7 32.8 ± 0.7* 37.1 ± 0.7* 41.1 ± 0.7 45.1 ± 0.7* 49.1 ± 0.7* (mmHg) NX2 39.5 ± 0.7* 41.5 ± 0.7 32.8 ± 0.7* 37.2 ± 0.7* 41.5 ± 0.7  45.7 ± 0.7*‡  49.9 ± 0.7*‡   NX1 vs NX2: P=0.014; Stage: P<0.001; Interaction: P=0.029 PaCO2 NX1 40.2 ± 0.7 40.7 ± 0.7 33.3 ± 0.9* 36.9 ± 0.9* 41.4 ± 0.7 44.4 ± 0.7* 47.9 ± 0.8* (mmHg) NX2 40.4 ± 0.7  41.3 ± 0.7‡ 32.9 ± 0.9* 37.3 ± 0.8* 41.5 ± 0.8 44.9 ± 0.8* 47.9 ± 0.8*  NX1 vs NX2: P=0.041; Stage: P<0.001; Interaction: P=0.199 Pa-PETCO2 NX1 0.7 ± 0.3†* -0.3 ± 0.2 0.5 ± 0.2  -0.2 ± 0.3 0.3 ± 0.2 -0.7 ± 0.2† -1.1 ± 0.2†* (mmHg) NX2 0.8 ± 0.3†* -0.3 ± 0.3 0.1 ± 0.4 0.1 ± 0.3 0.0 ± 0.3 -0.8 ± 0.2† -1.9 ± 0.3†*  NX1 vs NX2: P=0.225; Stage: P<0.001; Interaction: P=0.063 PETO2 NX1 95.6 ± 1.1 94.3 ± 1.3  94.2 ± 1.1 93.3 ± 1.2 93.5 ± 1.1 94.4 ± 1.0  94.3 ± 1.0 (mmHg) NX2 95.0 ± 0.9 94.0 ± 1.1 94.3 ± 1.1  93.3 ± 1.3  93.7 ± 1.0  94.0 ± 1.2  94.3 ± 1.1  NX1 vs NX2: P=0.596; Stage: P=0.005; Interaction: P=0.969 PaO2 NX1 88.5 ± 1.4 90.8 ± 1.4  90.1 ± 1.4 89.8 ± 1.4  88.8 ± 1.8  91.5 ± 1.1  92.3 ± 2.1* (mmHg) NX2 88.4 ± 1.1  88.4 ± 1.2 91.2 ± 1.6  90.4 ± 1.1  88.9 ± 1.4 90.4 ± 1.3 93.3 ± 1.5*  NX1 vs NX2: P=0.795; Stage: P<0.001; Interaction: P=0.326 PET-PaO2 NX1 7.0 ± 0.8† 3.5 ± 0.9† 4.1 ± 0.9† 3.5 ± 0.8† 4.7 ± 0.9† 2.9 ± 0.6† 2.1 ± 0.4†* (mmHg) NX2 6.6 ± 0.9† 5.7 ± 0.7† 3.1 ± 1.0† 2.9 ± 1.0† 4.8 ± 0.8† 3.6 ± 0.7† 1.0 ± 0.8*  NX1 vs NX2: P=0.960; Stage: P<0.001; Interaction: P=0.449 SaO2 NX1 97.7 ± 0.2 97.7 ± 0.3 98.2 ± 0.2* 97.9 ± 0.3 97.6 ± 0.3 97.6 ± 0.2 97.6 ± 0.2 (%) NX2 97.7 ± 0.3 97.7 ± 0.3 98.5 ± 0.2* 98.2 ± 0.2 97.8 ± 0.2 97.6 ± 0.2 97.7 ± 0.2  NX1 vs NX2: P=0.293; Stage: P<0.001; Interaction: P=0.270 pH NX1 7.42  0.00 7.42  0.01 7.49  0.01* 7.45  0.01* 7.41  0.00 7.39  0.00* 7.37  0.00*  NX2 7.42  0.00 7.41  0.00 7.49  0.01* 7.45  0.01* 7.41  0.00 7.39  0.00* 7.37  0.01*  NX1 vs NX2: P=0.308; Stage: P<0.001; Interaction: P=0.214 Definition of Abbreviations: PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference; SaO2, oxygen saturation of hemoglobin. *P<0.05, main effect of CO2 stage vs AB-DEF. Bolded NX2, main effect between NX1 and NX2, bolded value is significantly larger. ‡P<0.05, interaction effect. Values are mean  SEM. †P<0.05, presence of an end-tidal-to-arterial gradient.   38 3.2.2 Hypoxia Protocols Of the 15 participants included in normoxia and hypoxia protocol data analysis, one participant was completely excluded due to incomplete data in the hypoxia trials.  Baseline 1 (AB) values are the same between HX1 and HX2 trials, due to the nature of our protocol, all participants only had one unforced room air baseline (baseline 1 = AB), prior to both hypoxia protocols.  3.2.2.1 HX1 CO2 protocols The end-tidal gas profiles for a single representative subject during HX1 and HX2 is illustrated in Figure 3.4.  Table 3.4 displays ventilatory, cardiovascular, and cerebrovascular data for both HX1 and HX2 isooxic CO2 reactivity tests.  As expected, V̇E and VT were lower during AB compared to HX-DEF when hypoxia was administered, and were elevated compared to HX-DEF (DEF controlled baseline) in the hypercapnic ranges, while participants’ FB was only elevated at the +8 mmHg PETCO2 step.  The PICO2 was lower during AB and hypocapnia, while it was elevated during hypercapnia.  Heart rate was lower during AB and hypocapnia, but elevated during the +8 mmHg PETCO2 step from HX-DEF, while MAP was lower during AB and the -8 mmHg PETCO2 step, but elevated during hypercapnia compared to HX-DEF. Absolute MCAv was reduced by 8.9  2.0% during AB compared to HX-DEF, and absolute MCAv and Q̇ICA were reduced by 21.6  1.3% and 26.0  2.6% at -8 PETCO2 mmHg, and 9.7  1.5% and 13.2  2.8% at -4 PETCO2 mmHg from HX-DEF, respectively.  During hypercapnia, absolute MCAv and Q̇ICA were increased by 20.8  2.8% and 30.2  3.8% at +4 PETCO2 mmHg, and 39.0  3.7% and 64.5  5.5% at +8 PETCO2 mmHg from HX-DEF, respectively.   Table 3.5 displays respiratory and arterial blood data for both HX1 and HX2 isooxic CO2 reactivity tests.  The PETCO2 was lower during AB compared to HX-DEF, while PETCO2 and PaCO2 were both lower during hypocapnia and elevated during hypercapnia. A positive Pa-PETCO2 gradient was present during AB, whilst a negative Pa-PETCO2 gradient was present during both +4 and +8 mmHg PETCO2 steps (see also Figure 3.5, pg 40).  Throughout all stages of the HX1 CO2 reactivity protocol, PETO2 and PaO2 were not different from HX-DEF with the exception of AB where the subject was breathing room air. The PET-PaO2 gradient was present and consistent throughout all protocol stages (see also Figure 3.6, pg 41). SaO2 was   39 elevated during AB and -8 mmHg PETCO2, and lower during +4 and +8 mmHg PETCO2 compared to HX-DEF. 3.2.2.2 HX2 CO2 protocols Table 3.4 displays ventilatory, cardiovascular, and cerebrovascular data for both HX1 and HX2 isooxic CO2 reactivity tests.  Minute ventilation and VT were lower during AB compared to HX-DEF, and as expected, were elevated compared to HX-DEF (DEF controlled baseline) in the hypercapnic ranges, while participants FB was only elevated at the +8 mmHg PETCO2 step.  The PICO2 was lower during AB and hypocapnia, while it was elevated during hypercapnia.  Heart rate was lower during AB and hypocapnia, but elevated during the +8 mmHg PETCO2 step from HX-DEF, while MAP was lower during AB and -8 mmHg PETCO2, but elevated during hypercapnia compared to HX-DEF.  Absolute MCAv was -10.9  1.6% during AB compared to HX-DEF, and absolute MCAv and ICA blood flow were -24.0  1.2% and -30.4  2.5% at -8 PETCO2 mmHg, and -10.3  0.9% and -15.6  2.4% at -4 PETCO2 mmHg from HX-DEF, respectively.  During hypercapnia, absolute MCAv and Q̇ICA were 16.0  2.3% and 23.8  3.6% at +4 PETCO2 mmHg, and 37.0  4.0% and 60.7  7.0% at +8 PETCO2 mmHg from HX-DEF, respectively.  Table 3.5 displays respiratory and arterial blood data for both HX1 and HX2 isooxic CO2 reactivity tests.  The PETCO2 was lower during AB compared to HX-DEF, while PETCO2 and PaCO2 were both lower during hypocapnia and elevated during hypercapnia.  A positive Pa-PETCO2 gradient was present during AB, while a negative Pa-PETCO2 gradient was present during +4 and +8 mmHg PETCO2 steps (see also Figure 3.5, pg 40). Throughout all stages of the HX1 CO2 reactivity protocol, PETO2 and PaO2 were not different from HX-DEF with the exception of AB where the subject was breathing room air.  The PET-PaO2 gradient was present and consistent throughout all protocol stages (see Figure 3.6, pg 41).  SaO2 was elevated during AB and -8 mmHg PETCO2, and lower during +4 and +8 mmHg PETCO2 compared to HX-DEF.  3.2.2.3 HX1 vs. HX2 CO2 protocols Table 3.4 and Table 3.5 display ventilatory, cardiovascular, cerebrovascular, respiratory and arterial blood gas data during HX1 and HX2 CO2 reactivity protocols.  As expected, all outcome variables were similar, with the exception of PETCO2 and PaCO2, which were elevated   40 in HX2 compared to HX1.  In addition, there were no difference in between PFO+ and PFO- participants between both HX1 and HX2 protocols.  A main effect was detected with nasopharyngeal temperature which was significantly increased during AB compared to HX-DEF by 0.16  0.01 C.    Figure 3.4: End-tidal gas tracings from HX1 and HX2 CO2 reactivity trials from one representative subject. Open circles () represent PETO2, end-tidal partial pressure of oxygen; closed circles () represent PETCO2, end-tidal partial pressure of carbon dioxide. Each data point represents a 15-second average.  AB, air-breathing baseline; HX-DEF, isocapnic hypoxia breathing controlled using DEF.  Figure 3.5: Bland and Altman plot for agreement between PaCO2 and PETCO2 during HX1 (Panel A) and HX2 (Panel B) CO2 reactivity protocols.  (), -8 mmHg; (▼), -4 mmHg; (∆), 0 mmHg; (■), +4 mmHg; (□), +8 mmHg.  Dotted lines represent the 95% confidence intervals and the continuous lines represent the mean bias. Col 1 vs PO2 Col 1 vs PCO2 Time (s)0 1000 2000 3000 4000 5000PETCO2 and PETO2 (mmHg)010030405060708090100110AB HX-DEF-8-40+4+8HX-DEF-8-40+4+8HX1 HX2PaCO2 and PETCO2 mean (mmHg)25 30 35 40 45 50 55PaCO2 - PETCO2  (mmHg)-4-202PaCO2 and PETCO2 mean (mmHg)25 30 35 40 45 50 55-4-2024A. B.  41  Figure 3.6: Bland and Altman plot for agreement between PaO2 and PETO2 during HX1 (Panel A) and HX2 (Panel B) CO2 reactivity protocols.  (), -8 mmHg; (▼), -4 mmHg; (∆), 0 mmHg; (■), +4 mmHg; (□), +8 mmHg.  Dotted lines represent the 95% confidence intervals and the continuous lines represent the mean bias.   PaO2 and PETO2 mean (mmHg)44 46 48 50 52PETO2 - PaO2 (mmHg)02468101214PaO2 and PETO2 mean (mmHg)44 46 48 50 5202468101214A. B.  42    Table 3.4: Ventilatory cardiovascular, and cerebrovascular data during HX1 and HX2 CO2 reactivity tests.  Trial AB HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  HX1 12.8 ± 0.5* 27.4 ± 1.5 22.2 ± 1.2 22.2 ± 1.3 26.7 ± 1.3 40.5 ± 2.6* 56.4 ± 4.3* (l/min) HX2 12.8 ± 0.5* 28.7 ± 1.5 21.5 ± 1.1 20.7 ± 1.2 27.6 ± 1.4 43.0 ± 2.9* 58.4 ± 4.4*  HX1 vs HX2: P=0.315; Stage: P<0.001; Interaction: P=0.417 VT  HX1 0.9 ± 0.0* 1.7 ± 0.1 1.7 ± 0.2 1.6 ± 0.2 1.7 ± 0.2 2.3 ± 0.1* 2.6 ± 0.1* (l) HX2 0.9 ± 0.0*  1.9 ± 0.1 1.7 ± 0.2 1.6 ± 0.2 1.8 ± 0.2 2.3 ± 0.1* 2.6 ± 0.1*  HX1 vs HX2: P=0.221; Stage: P<0.001; Interaction: P=0.579 FB  HX1 15.1  0.9 17.0  1.3 15.5  1.9 16.0  1.9 17.0  1.2 17.9  1.3 22.0  1.7* (/min) HX2 15.1  0.9 16.5  1.2 14.5  1.6 14.3  1.3 16.3  1.2 19.3  1.7 22.6  1.9*  HX1 vs HX2: P=0.505; Stage: P<0.001; Interaction: P=0.082 PICO2  HX1 4.2 ± 0.6* 27.9 ± 0.7 12.5 ± 1.4* 20.7 ± 1.2* 28.8 ± 0.7 36.5 ± 0.5* 41.3 ± 0.5* (mmHg) HX2 4.2 ± 0.6* 29.4 ± 0.6 11.5 ± 1.5*  20.4 ± 1.3*  29.9 ± 0.7  37.0 ± 0.6* 41.7 ± 0.4*  HX1 vs HX2: P=0.301; Stage: P<0.001; Interaction: P=0.239 HR  HX1 61.5 ± 2.4* 76.0 ± 2.7 73.2 ± 2.6*  70.5 ± 2.9* 72.8 ± 2.6 77.1 ± 2.8 83.4 ± 3.3* (/min) HX2 61.5 ± 2.4* 76.2 ± 3.0 71.3 ± 2.7* 71.3 ± 2.7* 72.9 ± 2.9 77.9 ± 2.7 84.5 ± 2.9*  HX1 vs HX2: P=0.852; Stage: P<0.001; Interaction: P=0.405 MAP  HX1 86.5 ± 1.7* 92.1 ± 2.2 87.8 ± 2.1*  90.5 ± 2.0 93.0 ± 2.0 95.9 ± 2.0* 101.9 ± 2.3* (mmHg) HX2 86.5 ± 1.7* 91.9 ± 2.4  89.4 ± 2.5* 91.3 ± 2.2 92.0 ± 2.5 96.9 ± 2.7* 103.4 ± 2.7*  HX1 vs HX2: P=0.606; Stage: P<0.001; Interaction: P=0.227 MCAv  HX1 61.8 ± 3.0* 68.1 ± 2.8 53.3 ± 2.0* 61.1 ± 2.4* 69.6 ± 2.8 81.9 ± 3.3* 94.5 ± 4.1* (cm/s) HX2 61.8 ± 3.0* 69.4 ± 3.0 52.6 ± 2.2* 62.0 ± 2.4* 69.7 ± 2.9 80.1 ± 3.2* 94.6 ± 4.3*  HX1 vs HX2: P=0.983; Stage: P<0.001; Interaction: P=0.710 Q̇ICA  HX1 278.1 ± 14.4 312.1 ± 15.6 229.4 ± 11.0* 268.2 ± 11.6 326.5 ± 18.0 407.0 ± 26.4* 513.0 ± 31.3* (ml/min) HX2 278.1 ± 14.4 324.6 ± 17.5 223.4 ± 9.6* 271.6 ± 11.3 332.7 ± 20.4 398.9 ± 19.2* 517.2 ± 30.1*  HX1 vs HX2: P=0.765; Stage: P<0.001; Interaction: P=0.798 Nasal Temp  HX1 36.5 ± 0.1*  36.3 ± 0.1 36.3 ± 0.1 36.3 ± 0.1 36.4 ± 0.1 36.3 ± 0.1 36.2 ± 0.1 (C) HX2 36.5 ± 0.1*  36.4 ± 0.1 36.3 ± 0.1 36.4 ± 0.1 36.4 ± 0.1 36.3 ± 0.1 36.3 ± 0.1  HX1 vs HX2: P=0.267; Stage: P<0.001; Interaction: P=0.366 Definition of Abbreviations: V̇E, minute ventilation; VT, tidal volume; FB, breathing frequency; PICO2, partial pressure of inspired carbon dioxide; HR, heart rate; MAP, mean arterial pressure; MCAv, middle cerebral artery blood flow velocity; Q̇ICA, internal carotid artery blood flow.  *P<0.05, main effect of CO2 stage vs HX-DEF. Values are mean  SEM.    43 Table 3.5: Ventilatory and blood gas data during HX1 and HX2 CO2 reactivity tests.   Trial AB HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg PETCO2  HX1 39.1 ± 0.6* 40.2 ± 0.6 31.9 ± 0.6* 36.2 ± 0.6* 40.3 ± 0.6 44.2 ± 0.6* 48.2 ± 0.6* (mmHg) HX2 39.1 ± 0.6* 40.8 ± 0.6‡ 31.8 ± 0.6* 36.5 ± 0.6* 40.6 ± 0.5 44.7 ± 0.6‡* 48.8 ± 0.5‡*  HX1 vs HX2: P=0.008; Stage: P<0.001; Interaction: P<0.001 PaCO2  HX1 40.0 ± 0.6 40.0 ± 0.7 32.3 ± 0.7* 36.4 ± 0.7* 40.1 ± 0.6 43.5 ± 0.7* 47.3 ± 0.7* (mmHg) HX2 40.0 ± 0.6 40.4 ± 0.7  32.3 ± 0.7* 36.4 ± 0.7* 40.5 ± 0.6 44.2 ± 0.6‡* 48.2 ± 0.7‡*  HX1 vs HX2: P=0.010; Stage: P<0.001; Interaction: P=0.024 Pa-PETCO2  HX1 0.9 ± 0.3†* -0.2 ± 0.3 0.4 ± 0.3 0.2 ± 0.2 -0.2 ± 0.2 -0.7 ± 0.3†* -0.9 ± 0.3†* (mmHg) HX2 0.9 ± 0.3†* -0.4 ± 0.3 0.5 ± 0.2 -0.1 ± 0.3 -0.1 ± 0.2 -0.5 ± 0.3†* -0.7 ± 0.3†*  HX1 vs HX2: P=0.745; Stage: P<0.001; Interaction: P=0.425 PETO2  HX1 94.4 ± 0.6* 50.8 ± 0.1 50.6 ± 0.3 50.8 ± 0.1 50.8 ± 0.1 50.9 ± 0.1 50.9 ± 0.1 (mmHg) HX2 94.4 ± 0.6* 50.6 ± 0.1 51.4 ± 0.2‡ 50.5 ± 0.3‡ 50.9 ± 0.1 50.8 ± 0.1 50.9 ± 0.1  HX1 vs HX2: P=0.660; Stage: P<0.001; Interaction: P<0.001 PaO2  HX1 87.4 ± 1.1* 44.1 ± 0.5 45.0 ± 0.6 44.4 ± 0.3 44.1 ± 0.7 45.0 ± 0.4 44.9 ± 0.5 (mmHg) HX2 87.4 ± 1.1* 45.1 ± 0.6 45.3 ± 0.6 43.9 ± 0.7 44.4 ± 0.6 44.5 ± 0.6 45.5 ± 0.5  HX1 vs HX2: P=0.422; Stage: P<0.001; Interaction: P=0.262 PET-PaO2  HX1 6.9 ± 1.0† 6.7 ± 0.5† 5.6 ± 0.5† 6.4 ± 0.3† 6.7 ± 0.7† 5.9 ± 0.4† 6.1 ± 0.6† (mmHg) HX2 6.9 ± 1.0† 5.5 ± 0.6† 6.0 ± 0.7† 6.5 ± 0.7† 6.4 ± 0.6† 6.3 ± 0.6† 5.5 ± 0.6†  HX1 vs HX2: P=0.426; Stage: P=0.394; Interaction: P=0.129 SaO2  HX1 97.9 ± 0.2* 83.5 ± 0.6 87.1 ± 0.4* 84.7 ± 0.5 83.0 ± 0.5 82.1 ± 0.6* 81.5 ± 0.5* (%) HX2 97.9 ± 0.2* 83.7 ± 0.6 87.2 ± 0.4* 84.0 ± 0.5 82.9 ± 0.6 82.0 ± 0.6* 81.3 ± 0.7*  HX1 vs HX2: P=0.366; Stage: P<0.001; Interaction: P=0.508 pH HX1 7.42  0.00 7.42  0.00 7.50  0.00* 7.46  0.00* 7.43  0.00 7.40  0.00* 7.37  0.00*  HX2 7.42  0.00 7.42  0.00 7.49  0.01* 7.46  0.01* 7.42  0.00 7.39  0.00* 7.36  0.00*  HX1 vs HX2: P=0.058; Stage: P<0.001; Interaction: P=0.499 Definition of Abbreviations: PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference; SaO2, oxygen saturation of hemoglobin.  *P<0.05, main effect of CO2 stage vs HX-DEF. ‡P<0.05, interaction effect. Bolded HX2, main effect between HX1 and HX2, bolded value is significantly larger. †P<0.05, end-tidal-to-arterial gradient present. Values are mean  SEM.     44 3.2.3 Comparison of normoxia and hypoxia CO2 protocols We compared the results between normoxia (NX1) and hypoxia (HX1) CO2 reactivity protocols.  Table 3.6 displays ventilatory, cardiovascular, and cerebrovascular data for both NX1 and HX1 isooxic CO2 reactivity tests.  Minute ventilation and VT were lower during NX1 compared to HX1 trial, while participants FB was the same.  The PICO2 was significantly lower overall during NX1 trial compared to HX1, but only had an interaction effect at +4 mmHg PETCO2 during hypercapnia, there was no interaction effect in PICO2 during the +8 mmHg PETCO2 step, which was unexpected.  There was a main effect for HR, MAP, MCAv, and Q̇ICA, where they were significantly higher during HX1 compared to NX1, but no interactions for each of these variables.  Nasopharyngeal temperature was lower during NX1 compared to HX1 CO2 reactivity protocol. Table 3.7 displays respiratory and arterial blood data for both NX1 and HX1 CO2 reactivity tests.  The PETCO2 was higher during NX1 compared to HX1 CO2 reactivity protocols, while PaCO2 and the Pa-PETCO2 gradient remained unchanged.  As expected there was a main effect with variables PETO2 and PaO2, as they were significantly higher during NX1 compared to HX1 CO2 reactivity protocol.  There was a significantly larger (more positive) PET-PaO2 gradient during the HX1 trial compared to NX1, however there were no interaction effects.  SaO2 was higher, while pH was lower during the NX1 protocol compared to the HX1 CO2 reactivity protocol.   45 Table 3.6: Comparison of ventilatory cardiovascular, and cerebrovascular data between NX1 and HX1 CO2 reactivity tests.  Trial AB AB/HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  NX1 12.4 ± 0.5 16.9 ± 1.0 22.2 ± 1.3 20.6 ± 1.1 17.7 ± 0.8 26.5 ± 2.2* 38.9 ± 3.3* (l/min) HX1 12.8 ± 0.5* 27.4 ± 1.5‡ 22.2 ± 1.2 22.2 ± 1.3 26.7 ± 1.3 40.5 ± 2.6‡* 56.4 ± 4.3‡*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P<0.001 VT  NX1 0.9 ± 0.0* 1.3 ± 0.1 1.7 ± 0.2* 1.6 ± 0.2 1.3 ± 0.1 1.7 ± 0.1* 2.2 ± 0.1* (l) HX1 0.9 ± 0.0* 1.7 ± 0.1 1.7 ± 0.2 1.6 ± 0.2 1.7 ± 0.2 2.3 ± 0.1* 2.6 ± 0.1*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P=0.107 FB  NX1 14.3  1.0 14.2  1.1 15.8  1.7 15.8  2.1 14.8  1.1 17.3  2.0 18.9  2.0* (/min) HX1 15.1  0.9 17.0  1.3 15.5  1.9 16.0  1.9 17.0  1.2 17.9  1.3 22.0  1.7*  NX1 vs HX1: P=0.123; Stage: P=0.013; Interaction: P=0.916 PICO2  NX1 3.8 ± 0.7* 19.1 ± 2.1 12.3 ± 1.7* 19.1 ± 1.3 22.1 ± 1.2 33.1 ± 1.1* 40.7 ± 1.0* (mmHg) HX1 4.2 ± 0.6* 27.9 ± 0.7‡ 12.5 ± 1.4* 20.7 ± 1.2* 28.8 ± 0.7 36.5 ± 0.5‡* 41.3 ± 0.5*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P<0.001 HR  NX1 61.3 ± 2.5* 63.7 ± 2.5 66.8 ± 2.2 65.1 ± 1.9 64.1 ± 2.5 69.6 ± 2.9 74.5 ± 2.7* (/min) HX1 61.5 ± 2.4* 76.0 ± 2.7 73.2 ± 2.6  70.5 ± 2.9* 72.8 ± 2.6 77.1 ± 2.8 83.4 ± 3.3*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P=0.463 MAP  NX1 83.9 ± 2.1 85.2 ± 1.8 85.2 ± 1.7 86.1 ± 1.7 87.4 ± 1.6 91.8 ± 2.2 95.3 ± 2.1* (mmHg) HX1 86.5 ± 1.7* 92.1 ± 2.2 87.8 ± 2.1  90.5 ± 2.0 93.0 ± 2.0 95.9 ± 2.0 101.9 ± 2.3*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P=0.890 MCAv  NX1 61.1 ± 2.7 64.5 ± 3.1 50.8 ± 2.1* 57.1 ± 2.5 65.6 ± 3.1  72.3 ± 3.9* 84.2 ± 3.9* (cm/s) HX1 61.8 ± 3.0 68.1 ± 2.8 53.3 ± 2.0* 61.1 ± 2.4 69.6 ± 2.8 81.9 ± 3.3* 94.5 ± 4.1*  NX1 vs HX1: P=0.004; Stage: P<0.001; Interaction: P=0.688 Q̇ICA  NX1 265.5 ± 15.8 277.7 ± 11.9  203.5 ± 12.1* 234.3 ± 13.8 289.3 ± 14.7 334.9 ± 19.7* 428.7 ± 28.2* (ml/min) HX1 278.1 ± 14.4 312.1 ± 15.6 229.4 ± 11.0* 268.2 ± 11.6 326.5 ± 18.0 407.0 ± 26.4* 513.0 ± 31.3*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P=0.555 Nasal Temp  NX1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1* (C) HX1 36.5 ± 0.1*  36.3 ± 0.1 36.3 ± 0.1 36.3 ± 0.1 36.4 ± 0.1 36.3 ± 0.1 36.2 ± 0.1  NX1 vs HX1: P<0.001; Stage: P=0.676; Interaction: P=0.806 Definition of Abbreviations: V̇E, minute ventilation; V̇T, tidal volume; FB, breathing frequency; PICO2, partial pressure of inspired carbon dioxide; HR, heart rate; MAP, mean arterial pressure; MCAv, middle cerebral artery blood flow velocity; Q̇ICA, internal carotid artery blood flow.  *P<0.05, main effect of CO2 stage vs AB/HX-DEF. ‡P<0.05, interaction effect. Bolded HX1 or NX1, main effect between NX1 and HX1, bolded value is significantly larger. Values are mean  SEM.    46     Table 3.7: Comparison of ventilatory and blood gas data during NX1 and HX1 CO2 reactivity tests.   Trial AB AB/HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg PETCO2  NX1 39.5 ± 0.7 41.0 ± 0.7 32.8 ± 0.7* 37.1 ± 0.7* 41.1 ± 0.7 45.1 ± 0.7* 49.1 ± 0.7* (mmHg) HX1 39.1 ± 0.6 40.2 ± 0.6 31.9 ± 0.6* 36.2 ± 0.6* 40.3 ± 0.6 44.2 ± 0.6* 48.2 ± 0.6*  NX1 vs HX1: P=0.027; Stage: P<0.001; Interaction: P=1.00 PaCO2  NX1 40.2 ± 0.7 40.7 ± 0.7 33.3 ± 0.9* 36.9 ± 0.9* 41.4 ± 0.7 44.4 ± 0.7* 47.9 ± 0.8* (mmHg) HX1 40.0 ± 0.6 40.0 ± 0.7 32.3 ± 0.7* 36.4 ± 0.7* 40.1 ± 0.6 43.5 ± 0.7* 47.3 ± 0.7*  NX1 vs HX1: P=0.066; Stage: P<0.001; Interaction: P=0.991 Pa-PETCO2  NX1 0.7 ± 0.3†* -0.3 ± 0.2 0.5 ± 0.2  -0.2 ± 0.3 0.3 ± 0.2 -0.7 ± 0.2† -1.1 ± 0.2† (mmHg) HX1 0.90 ± 0.3†* -0.2 ± 0.3 0.4 ± 0.3 0.2 ± 0.2 -0.2 ± 0.2 -0.7 ± 0.3† -0.9 ± 0.3†  NX1 vs HX1: P=0.738; Stage: P<0.001; Interaction: P=0.750 PETO2  NX1 95.6 ± 1.1 94.3 ± 1.3‡  94.2 ± 1.1‡ 93.3 ± 1.2‡ 93.5 ± 1.1 94.4 ± 1.0‡  94.3 ± 1.0‡ (mmHg) HX1 94.4 ± 0.6* 50.8 ± 0.1 50.6 ± 0.3 50.8 ± 0.1 50.8 ± 0.1 50.9 ± 0.1 50.9 ± 0.1  NX1 vs HX1: P=<0.001; Stage: P<0.001; Interaction: P<0.001 PaO2  NX1 88.5 ± 1.4 90.8 ± 1.4‡ 90.1 ± 1.4‡ 89.8 ± 1.4‡  88.8 ± 1.8  91.5 ± 1.1‡  92.3 ± 2.1‡* (mmHg) HX1 87.4 ± 1.1* 44.1 ± 0.5 45.0 ± 0.6 44.4 ± 0.3 44.1 ± 0.7 45.0 ± 0.4 44.9 ± 0.5  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P<0.001 PET-PaO2  NX1 7.0 ± 0.8† 3.5 ± 0.9† 4.1 ± 0.9† 3.5 ± 0.8† 4.7 ± 0.9 2.9 ± 0.6† 2.1 ± 0.4†* (mmHg) HX1 6.9 ± 1.0† 6.7 ± 0.5† 5.6 ± 0.5† 6.4 ± 0.3† 6.7 ± 0.7 5.9 ± 0.4† 6.1 ± 0.6†  NX1 vs HX1: P<0.001; Stage: P=0.002; Interaction: P=0.117 SaO2  NX1 97.7 ± 0.2 97.7 ± 0.3‡ 98.2 ± 0.2‡ 97.9 ± 0.3‡ 97.6 ± 0.3 97.6 ± 0.2‡ 97.6 ± 0.2‡ (%) HX1 97.9 ± 0.2* 83.5 ± 0.6 87.1 ± 0.4* 84.7 ± 0.5 83.0 ± 0.5 82.1 ± 0.6 81.5 ± 0.5*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P<0.001 pH NX1 7.42  0.00 7.42  0.01 7.49  0.01* 7.45  0.01* 7.41  0.00 7.39  0.00* 7.37  0.00*  HX1 7.42  0.00 7.42  0.00 7.50  0.00* 7.46  0.00* 7.43  0.00 7.40  0.00* 7.37  0.00*  NX1 vs HX1: P<0.001; Stage: P<0.001; Interaction: P=0.902 Definition of Abbreviations: PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference; SaO2, oxygen saturation of hemoglobin.  *P<0.05, main effect of CO2 stage vs AB/HX-DEF. ‡P<0.05, interaction effect. Bolded HX1 or NX1, main effect between NX1 and HX1, bolded value is significantly larger. †P<0.05, presence of an end-tidal-to-arterial gradient. Values are mean  SEM.    47  Research Question Two: end-tidal-to-arterial gradients in PFO+ and PFO- participants  3.3.1 Normoxia protocols Table 3.8 and Table 3.9 display ventilatory and arterial blood data for PFO+ (n=7) and PFO- (n=7) participants during the NX1 and NX2 CO2 reactivity protocols.  During both NX1 and NX2 CO2 reactivity protocols there were no differences in V̇E, PETCO2, PaCO2, the Pa-PETCO2 gradient, PETO2, PaO2 and nasopharyngeal temperature found between PFO+ and PFO- participants. However, during the NX1 protocols PFO+ participants presented with a greater PET-PaO2 gradient compared to PFO- participants (5.0  1.2 mmHg vs 2.9  1.2 mmHg, P=0.003). This difference in the PET-PaO2 gradient between PFO+ and PFO- participants was not observed during the NX2 protocol, but it was trending towards significance (P=0.070).   3.3.2 Hypoxia protocols Table 3.10 and Table 3.11 display ventilatory and arterial blood gas data for PFO+ (n=8) and PFO- (n=6) participants during the HX1 and HX2 CO2 reactivity protocol.  During both hypoxia CO2 reactivity protocols, no differences were found in V̇E, PETCO2, PaCO2, the Pa-PETCO2 gradient, PETO2, PaO2, the PET-PaO2 gradient, and body temperature between PFO+ and PFO- participants.   48           Table 3.8: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during NX1 CO2 reactivity test. NX1 AB AB-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  PFO+ 12.5  0.8 16.2  1.4 21.6  2.0 20.3  1.6 16.8  0.7 25.1  2.1* 34.5  3.0* (l/min) PFO- 12.3  0.7 17.7  1.4 22.8  1.7 20.9  1.5 18.5  1.3 27.8  3.9* 43.3  5.3*  PFO+ vs PFO-: P=0.363; Stage: P<0.001; Interaction: P=0.343 PETCO2  PFO+ 39.3 ± 0.8* 40.7 ± 0.8 32.4 ± 1.0*  36.8 ± 0.8* 41.0 ± 0.8 44.8 ± 0.8* 48.8 ± 0.8* (mmHg) PFO- 39.7 ± 1.2* 41.3 ± 1.0 33.2 ± 1.0* 37.3 ± 1.1* 41.3 ± 1.1 45.3 ± 1.0* 49.3 ± 1.1*  PFO+ vs PFO-: P=0.727; Stage: P<0.001; Interaction: P=0.845 PaCO2  PFO+ 40.2 ± 0.9 40.6 ± 0.8 32.6 ± 1.0*  36.6 ± 1.1* 41.5 ± 0.8 44.1 ± 0.9* 47.8 ± 0.8* (mmHg) PFO- 40.2 ± 1.2  40.7 ± 1.2 34.0 ± 1.3* 37.1 ± 1.4* 41.3 ± 1.1 44.6 ± 1.1* 48.1 ± 1.3*  PFO+ vs PFO-: P=0.824; Stage: P<0.001; Interaction: P=0.151 Pa-PETCO2  PFO+ 0.8 ± 0.5†* 0.0 ± 0.2 0.2 ± 0.3  -0.2 ± 0.4 0.6 ± 0.4 -0.7 ± 0.2† -1.0 ± 0.2† (mmHg) PFO- 0.6 ± 0.4†* -0.6 ± 0.3 0.8 ± 0.3 -0.2 ± 0.4 0.0 ± 0.2 -0.7 ± 0.3† -1.3 ± 0.3†  PFO+ vs PFO-: P=0.619; Stage: P<0.001; Interaction: P=0.372 PETO2  PFO+ 95.1 ± 1.2 93.3 ± 1.2 93.8 ± 1.1 92.4 ± 0.9 92.7 ± 0.8 93.7 ± 0.7 93.6 ± 0.8 (mmHg) PFO- 96.0 ± 1.7 95.4 ± 2.2 94.5 ± 1.9 94.2 ± 2.2 94.3 ± 1.9 95.2 ± 1.9 95.1 ± 1.8  PFO+ vs PFO-: P=0.516; Stage: P=0.053; Interaction: P=0.962 PaO2  PFO+ 86.9 ± 1.7 88.0 ± 1.3 88.4 ± 1.5 88.0 ± 1.4 86.5 ± 1.6 90.6 ± 0.9 91.0 ± 0.9 (mmHg) PFO- 90.2 ± 2.1 93.6 ± 2.0 91.8 ± 2.0 91.7 ± 2.3 91.1 ± 2.9 92.4 ± 2.0 93.5 ± 2.0  PFO+ vs PFO-: P=0.143; Stage: P=0.043; Interaction: P=0.816 PET-PaO2  PFO+ 8.2 ± 1.1†* 5.3 ± 1.0† 5.4 ± 1.2† 4.4 ± 1.0† 6.2 ± 1.0 3.1 ± 0.4† 2.5 ± 0.7† (mmHg) PFO- 5.8 ± 0.9†* 1.8 ± 1.0† 2.7 ± 1.2† 2.5 ± 1.1† 3.2 ± 1.3 2.7 ± 1.1† 1.6 ± 0.5†  PFO+ vs PFO-: P=0.003; Stage: P<0.001; Interaction: P=0.780 Nasal Temp  PFO+ 36.4 ± 0.2 36.6 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.6 ± 0.0 36.6 ± 0.1 36.5 ± 0.1 (C) PFO- 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.4 ± 0.1  PFO+ vs PFO-: P=0.665; Stage: P=0.285; Interaction: P=0.732 Definition of Abbreviations: V̇E, minute ventilation; PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference. *P<0.05, main effect of CO2 stage vs AB-DEF. Bolded PFO+, main effect between PFO+ and PFO-, bolded value is significantly larger. †P<0.05, presence of an end-tidal-to-arterial gradient. Values are mean  SEM.     49              Table 3.9: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during NX2 CO2 reactivity test.   NX2 AB AB-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  PFO+ 13.5  0.8 16.4  1.0 23.2  2.6* 21.3  1.5 19.0  1.6 27.7  2.2* 38.1 2.8* (l/min) PFO- 12.8  0.6 16.7  1.1 22.8  1.6* 22.6  1.9 18.2  1.2 27.0  2.5* 44.0  5.3*  PFO+ vs PFO-: P=0.780; Stage: P<0.001; Interaction: P=0.491 PETCO2  PFO+ 39.0 ± 0.6* 41.0 ± 0.9 32.4 ± 0.9*  36.6 ± 0.8* 41.0 ± 0.8 45.3 ± 0.8* 49.2 ± 0.9* (mmHg) PFO- 40.1 ± 0.9* 42.0 ± 1.1 33.4 ± 1.1* 37.8 ± 1.0* 42.0 ± 1.1 46.1 ± 1.1* 50.5 ± 1.0*  PFO+ vs PFO-: P=0.463; Stage: P<0.001; Interaction: P=0.924 PaCO2  PFO+ 40.3 ± 0.6 41.0 ± 0.9 32.8 ± 1.4*  36.9 ± 1.1* 41.3 ± 0.9 44.9 ± 0.9* 47.5 ± 1.1* (mmHg) PFO- 40.5 ± 1.3  41.6 ± 1.2 33.1 ± 1.2* 37.6 ± 1.2* 41.8 ± 1.3 44.9 ± 1.2* 48.4 ± 1.3*  PFO+ vs PFO-: P=0.780; Stage: P<0.001; Interaction: P=0.936 Pa-PETCO2  PFO+ 1.2 ± 0.4†* -0.1 ± 0.5 0.4 ± 0.6  0.3 ± 0.4 0.3 ± 0.3 -0.4 ± 0.3† -1.8 ± 0.3†* (mmHg) PFO- 0.4 ± 0.4†*  -0.5 ± 0.3 -0.3 ± 0.4 -0.2 ± 0.3 -0.3 ± 0.4 -1.2 ± 0.1† -2.1 ± 0.5†*  PFO+ vs PFO-: P=0.156; Stage: P<0.001; Interaction: P=0.991 PETO2  PFO+ 94.9 ± 1.1 93.2 ± 0.8 93.5 ± 0.7 92.3 ± 1.6 92.7 ± 1.0 92.7 ± 0.9 93.5 ± 0.8 (mmHg) PFO- 95.0 ± 1.5 94.8 ± 1.9 95.1 ± 2.0 94.3 ± 2.0 94.7 ± 1.7 95.4 ± 2.0 95.1 ± 1.9  PFO+ vs PFO-: P=0.462; Stage: P=0.201; Interaction: P=0.525 PaO2  PFO+ 87.0 ± 1.0 85.7 ± 0.7 89.7 ± 2.2 88.3 ± 0.9 87.7 ± 1.2 87.9 ± 1.1 92.2 ± 1.2* (mmHg) PFO- 89.8 ± 1.8 91.0 ± 1.7 92.7 ± 2.4 92.5 ± 1.6 90.0 ± 2.3 92.9 ± 1.8 94.4 ± 2.6*  PFO+ vs PFO-: P=0.124; Stage: P<0.001; Interaction: P=0.719 PET-PaO2  PFO+ 8.0 ± 1.0† 7.5 ± 1.0† 3.8 ± 1.7† 4.0 ± 1.7† 5.0 ± 0.7 4.8 ± 0.8† 1.3 ± 0.9†* (mmHg) PFO- 5.1 ± 1.2† 3.8 ± 0.6† 2.5 ± 1.1† 1.8 ± 0.7† 4.6 ± 1.5 2.4 ± 1.1† 0.7 ± 1.4†*  PFO+ vs PFO-: P=0.070; Stage: P<0.001; Interaction: P=0.753 Nasal Temp PFO+ 36.5 ± 0.1 36.6 ± 0.1 36.5 ± 0.1 36.6 ± 0.1 36.7 ± 0.0 36.6 ± 0.0 36.5 ± 0.1* (C) PFO- 36.5 ± 0.1 36.5 ± 0.1 36.4 ± 0.1 36.4 ± 0.1 36.5 ± 0.1 36.4 ± 0.1 36.3 ± 0.2*  PFO+ vs PFO-: P=0.237; Stage: P=0.007; Interaction: P=0.412 Definition of Abbreviations: V̇E, minute ventilation; PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference. *P<0.05, main effect of CO2 stage vs AB-DEF. †P<0.05, presence of an end-tidal-to-arterial gradient. Values are mean  SEM     50         Table 3.10: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during HX1 CO2 reactivity test.   HX1 AB HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  PFO+ 12.7  0.6* 27.8  2.5 21.4  1.6 22.6  1.6 28.5  1.7 41.7  3.7* 55.0  5.0* (l/min) PFO- 13.0  0.8* 26.9  1.4 23.1  1.8 21.7  2.2 24.4  1.4 39.1  3.4* 58.3  7.5*  PFO+ vs PFO-: P=0.884; Stage: P<0.001; Interaction: P=0.829 PETCO2  PFO+ 38.6 ± 0.6* 39.9 ± 0.7 31.4 ± 0.7*  35.8 ± 0.7* 39.8 ± 0.7 43.8 ± 0.7* 47.8 ± 0.7* (mmHg) PFO- 39.9 ± 1.0*  40.7 ± 0.9 32.5 ± 1.0* 36.8 ± 1.0* 40.9 ± 1.0 44.7 ± 0.9* 48.9 ± 1.0*  PFO+ vs PFO-: P=0.416; Stage: P<0.001; Interaction: P=0.564 PaCO2  PFO+ 39.9 ± 0.7 39.6 ± 0.8 31.8 ± 1.0*  35.8 ± 0.8* 39.7 ± 0.7 43.2 ± 0.9* 47.1 ± 0.9* (mmHg) PFO- 40.2 ± 1.2 40.6 ± 1.1 33.0 ± 1.0*  37.2 ± 1.1* 40.6 ± 1.0 43.8 ± 1.0* 47.7 ± 0.9*  PFO+ vs PFO-: P=0.550; Stage: P<0.001; Interaction: P=0.469 Pa-PETCO2  PFO+ 1.3 ± 0.4†* -0.3 ± 0.3 0.3 ± 0.4  0.0 ± 0.2 -0.1 ± 0.3 -0.6 ± 0.4† -0.7 ± 0.5† (mmHg) PFO- 0.4 ± 0.4†*   -0.1 ± 0.4 0.5 ± 0.5 0.4 ± 0.4 -0.3 ± 0.2 -0.9 ± 0.4† -1.2 ± 0.4†  PFO+ vs PFO-: P=0.674; Stage: P<0.001; Interaction: P=0.326 PETO2  PFO+ 94.2 ± 0.6* 50.9 ± 0.1 50.5 ± 0.5 51.0 ± 0.2 50.9 ± 0.1 50.8 ± 0.1 50.9 ± 0.1 (mmHg) PFO- 94.7 ± 1.2* 50.7 ± 0.2 50.7 ± 0.3 50.6 ± 0.4 50.7 ± 0.2 51.0 ± 0.1 51.0 ± 0.2  PFO+ vs PFO-: P=0.908; Stage: P<0.001; Interaction: P=0.905 PaO2  PFO+ 86.7 ± 0.7* 43.7 ± 0.7 45.2 ± 0.8 44.8 ± 0.2 43.6 ± 0.7 44.5 ± 0.5 44.3 ± 0.7 (mmHg) PFO- 88.5 ± 2.4* 44.6 ± 0.6 44.7 ± 1.1 43.8 ± 0.4 44.7 ± 1.1 45.7 ± 0.6 45.7 ± 0.6  PFO+ vs PFO-: P=0.402; Stage: P<0.001; Interaction: P=0.561 PET-PaO2  PFO+ 7.5 ± 0.7† 7.1 ± 0.7† 5.4 ± 0.4† 6.2 ± 0.3† 7.3 ± 0.7 6.3 ± 0.5† 6.6 ± 0.8† (mmHg) PFO- 6.2 ± 2.0† 6.1 ± 0.6† 6.0 ± 1.0† 6.8 ± 0.6† 6.0 ± 1.3 5.3 ± 0.5† 5.3 ± 0.8†  PFO+ vs PFO-: P=0.492; Stage: P=0.482; Interaction: P=0.533 Nasal Temp  PFO+ 36.6 ± 0.1* 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.4 ± 0.2 36.3 ± 0.1 (C) PFO- 36.3 ± 0.2* 36.2 ± 0.2 36.1 ± 0.2 36.2 ± 0.2 36.2 ± 0.2 36.2 ± 0.2 36.1 ± 0.2  PFO+ vs PFO-: P=0.133; Stage: P<0.001; Interaction: P=0.504 Definition of Abbreviations: V̇E, minute ventilation; PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference. *P<0.05, main effect of CO2 stage vs AB-DEF.  †P<0.05, presence of an end-tidal-to-arterial gradient. Values are mean  SEM     51            Table 3.11: Ventilatory and arterial blood gas data between PFO+ and PFO- participants during HX2 CO2 reactivity test.   HX2 AB HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg V̇E  PFO+ 12.7  0.6* 30.2  2.4 22.1  1.6 19.9  1.2 29.9  1.9 45.7  3.8* 56.3  3.8* (l/min) PFO- 13.0  0.8* 26.8  0.7 20.6  1.5 21.9  2.3 24.6  1.2 39.4  4.0* 61.2  8.7*  PFO+ vs PFO-: P=0.650; Stage: P<0.001; Interaction: P=0.438 PETCO2  PFO+ 38.6 ± 0.6* 40.3 ± 0.7 31.2 ± 0.7*  36.1 ± 0.7* 40.2 ± 0.7 44.2 ± 0.7* 48.4 ± 0.7* (mmHg) PFO- 39.9 ± 1.0*  41.5 ± 1.0 32.6 ± 1.0* 37.0 ± 1.0* 41.3 ± 0.8 45.4 ± 0.9* 49.4 ± 0.8*  PFO+ vs PFO-: P=0.339; Stage: P<0.001; Interaction: P=0.695 PaCO2  PFO+ 39.9 ± 0.7 40.1 ± 0.7 31.8 ± 0.7*  36.2 ± 0.7* 40.2 ± 0.8 43.9 ± 0.8* 47.9 ± 0.7* (mmHg) PFO- 40.2 ± 1.2 40.8 ± 1.2 33.1 ± 1.2* 36.8 ± 1.4* 41.0 ± 1.0 44.7 ± 1.0* 48.5 ± 1.0*  PFO+ vs PFO-: P=0.608; Stage: P<0.001; Interaction: P=0.809 Pa-PETCO2  PFO+ 1.3 ± 0.4†* -0.2 ± 0.3 0.6 ± 0.2*  0.0 ± 0.3 0.0 ± 0.2 -0.3 ± 0.3† -0.4 ± 0.3† (mmHg) PFO- 0.4 ± 0.4†* -0.7 ± 0.4 0.5 ± 0.4* -0.2 ± 0.6 -0.3 ± 0.4 -0.7 ± 0.4† -1.0 ± 0.5†  PFO+ vs PFO-: P=0.312; Stage: P<0.001; Interaction: P=0.881 PETO2  PFO+ 94.2 ± 0.6* 50.6 ± 0.1 51.2 ± 0.2 50.4 ± 0.2 50.9 ± 0.1 50.8 ± 0.1 51.0 ± 0.2 (mmHg) PFO- 94.7 ± 1.2* 50.6 ± 0.2 51.6 ± 0.3 50.6 ± 0.6 50.8 ± 0.2 50.7 ± 0.2 50.9 ± 0.2  PFO+ vs PFO-: P=0.656; Stage: P<0.001; Interaction: P=0.966 PaO2  PFO+ 86.7 ± 0.7* 44.6 ± 0.6 44.7 ± 0.5 43.2 ± 0.9 43.9 ± 0.7 43.5 ± 0.6 44.8 ± 0.8 (mmHg) PFO- 88.5 ± 2.4* 45.7 ± 1.0 46.2 ± 1.3 45.0 ± 1.0 45.1 ± 0.9 45.8 ± 0.8 46.3 ± 0.4  PFO+ vs PFO-: P=0.110; Stage: P<0.001; Interaction: P=0.994 PET-PaO2  PFO+ 7.5 ± 0.7† 6.0 ± 0.7† 6.5 ± 0.6† 7.2 ± 1.0† 7.0 ± 0.7 7.3 ± 0.7† 6.1 ± 0.9† (mmHg) PFO- 6.2 ± 2.0† 4.9 ± 1.0† 5.4 ± 1.3† 5.6 ± 0.7† 5.7 ± 1.0 5.0 ± 0.8† 4.6 ± 0.5†  PFO+ vs PFO-: P=0.208; Stage: P=0.308; Interaction: P=0.976 Nasal Temp  PFO+ 36.6 ± 0.1 36.4 ± 0.1 36.4 ± 0.1 36.5 ± 0.1 36.5 ± 0.1 36.4 ± 0.1 36.3 ± 0.1 (C) PFO- 36.3 ± 0.2 36.3 ± 0.1 36.2 ± 0.1 36.3 ± 0.1 36.3 ± 0.2 36.2 ± 0.2 36.2 ± 0.2  PFO+ vs PFO-: P=0.246; Stage: P=0.001; Interaction: P=0.789 Definition of Abbreviations: V̇E, minute ventilation; PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide difference; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure of oxygen; PET-PaO2, end-tidal-to-arterial oxygen difference. *P<0.05, main effect of CO2 stage vs AB-DEF. †P<0.05, presence of an end-tidal-to-arterial gradient. Values are mean  SEM   52  Research Question Three: PETCO2 vs PaCO2 cerebrovascular and ventilatory reactivity   3.4.1  Normoxia reactivity data 3.4.1.1 Hypercapnic ventilatory reactivity data Figure 3.7 displays the comparison of normoxia HCVR when plotted against PETCO2, PaCO2 and Pred-PaCO2 (calculated using our previously derived correction algorithm, see equation 5).  For the NX1 trial the mean R2 values for HCVR was 0.96  0.02 for PETCO2 HCVR, 0.94  0.00 for PaCO2 HCVR, and 0.95  0.02 for Pred-PaCO2 HCVR.  During the NX1 reactivity protocol, PaCO2 HCVR was greater than PETCO2 HCVR by 0.5  0.1 l/min/mmHg and Pred-PaCO2 HCVR by 0.2  0.1 l/min/mmHg (P<0.001 and P=0.027, respectively), and the Pred-PaCO2 HCVR was greater compared to PETCO2 HCVR by 0.3  0.0 l/min/mmHg (P=0.006).   For the NX2 trial the mean R2 values for HCVR were 0.97  0.02 for PETCO2 HCVR, 0.96  0.02 for PaCO2 HCVR, and 0.97  0.02 for Pred-PaCO2 HCVR.  During the NX2 reactivity protocol, PaCO2 HCVR was greater than PETCO2 HCVR by 0.9  0.3 l/min/mmHg and Pred-PaCO2 HCVR by 0.5  0.2 l/min/mmHg (P=0.001 and P=0.05, respectively), and there was no difference between Pred-PaCO2 HCVR and PETCO2 HCVR (P=0.266).  Hypercapnic ventilatory response to PaCO2 was also compared between PFO+ and PFO- participants (see  Figure 3.8, pg 53), we found no differences in HCVR between PFO+ and PFO- participants during NX1 (P=0.437) and NX2 (P=0.383) CO2 reactivity protocols.   Figure 3.7: Hypercapnic ventilatory reactivity data during NX1 (Panel A) and NX2 (Panel B) protocols. *P<0.05, PaCO2 HCVR vs. PETCO2 and Pred-PaCO2 HCVR. †P<0.05, Pred-PaCO2 HCVR vs. PETCO2 HCVR.  NX1 Ventilatory Reactivity (l/min/mmHg)01245NX2 Ventilatory Reactivity (l/min/mmHg)012345**†A. B.PETCO2PaCO2Pred-PaCO2PETCO2PaCO2Pred-PaCO2  53   Figure 3.8: Hypercapnic ventilatory reactivity data during NX1 (Panel A) and NX2 (Panel B) protocols between PFO+ (n=7) and PFO- (n=7) participants.  3.4.1.2 Internal carotid artery reactivity Figure 3.9 displays relative HYPO-ICA and relative HYPER-ICA reactivity data during NX1 and NX2 CO2 reactivity protocols. Of the 14 participants included in normoxia mean data, two were excluded from the ICA analysis due to incomplete data (n=12).  For the NX1 trial the mean R2 values for HYPO-ICA CVR was 0.93  0.03 for PETCO2 CVR, 0.93  0.03 for PaCO2 CVR, and 0.93  0.03 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-ICA CVR was 0.89  0.02 for PETCO2 CVR, 0.90  0.02 for PaCO2 CVR, and 0.89  0.02 for Pred-PaCO2 CVR.  During the NX1 protocol (Panel A), there was no difference between PaCO2, PETCO2, and Pred-PaCO2 ICA reactivity in the hypocapnic range (P=0.131).  In the hypercapnic range, PaCO2 ICA reactivity was greater by 1.5  0.3 ml/min/mmHg compared to PETCO2 ICA reactivity (P<0.001), and by 0.8  0.2 ml/min/mmHg compared to Pred-PaCO2 ICA reactivity (P=0.011).  In addition, Pred-PaCO2 ICA reactivity was greater compared to PETCO2 ICA reactivity by 0.7  0.1 ml/min/mmHg (P<0.022).  For the NX2 trial the mean R2 values for HYPO-ICA CVR was 0.93  0.03 for PETCO2 CVR, 0.94  0.03 for PaCO2 CVR, and 0.94  0.03 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-ICA CVR was 0.96  0.02 for PETCO2 CVR, 0.95  0.02 for PaCO2 CVR, and 0.96  0.02 for Pred-PaCO2 CVR.  During the NX2 protocol (Panel B), there was no difference between PaCO2, PETCO2, and Pred-PaCO2 ICA reactivity in the hypocapnic range (P=0.480).  In the hypercapnic range, PaCO2 ICA reactivity was greater by 1.4  0.3 ml/min/mmHg compared to PETCO2 ICA reactivity (P<0.001) and by 0.8  0.2 ml/min/mmHg compared to the Pred-PaCO2 NX1 Ventilatory Reactivity (l/min/mmHg)0123456NX2 Ventilatory Reactivity (l/min/mmHg)0123456A. B. PFO + PFO- PFO + PFO-  54 ICA reactivity (P=0.007).  In addition, the Pred-PaCO2 ICA reactivity was greater by 0.7  0.1 ml/min/mmHg compared to the PETCO2 ICA reactivity (P=0.014).  HYPO-ICA PaCO2 CVR and HYPER ICA PaCO2 CVR was compared between PFO+ and PFO- participants, no differences during both NX1 and NX2 CO2 reactivity protocols for HYPO- (P=0.292 and P=0.460) and HYPER- (P=0.839 and P=0.575) ICA PaCO2 CVR were found between these two groups.  Figure 3.9: ICA hypocapnia and hypercapnia reactivity data during NX1 (Panel A) and NX2 (Panel B) protocols.  (black bar), ICA reactivity indexed against PETCO2; (white bar), ICA reactivity indexed against PaCO2; (grey bar), ICA reactivity indexed against Pred-PaCO2; calculated using our previously derived algorithm. †P<0.05, PaCO2 hypercapnic ICA reactivity vs. PETCO2 and Pred-PaCO2 hypercapnic ICA reactivity. *P<0.05, Pred-PaCO2 hypercapnic ICA reactivity vs PETCO2 ICA reactivity.   Middle cerebral artery reactivityFigure 3.10 displays relative HYPO-MCAv and relative HYPER-MCAv reactivity data during NX1 and NX2 CO2 reactivity protocols.  For the NX1 trail the mean R2 values for HYPO-MCA CVR was 0.98  0.0 for PETCO2 CVR, 0.98  0.0 for PaCO2 CVR, and 0.99  0.0 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-MCA CVR was 0.96  0.0 for PETCO2 CVR, 0.95  0.0 for PaCO2 CVR, and 0.96  0.0 for Pred-PaCO2 CVR.  During the NX1 protocol (Panel A), there was no difference in PaCO2, PETCO2, and Pred-PaCO2 MCAv reactivity in the hypocapnic range (P=0.120).  In the hypercapnic range, PaCO2 MCAv reactivity was greater by 0.7  0.1 cm/s/mmHg compared to PETCO2 MCAv reactivity, and by 0.3  0.1 cm/s/mmHg compared to Pred-PaCO2 MCAv reactivity (P<0.001 and P=0.002, respectively).  In addition, the Pred-PaCO2 MCAv reactivity was greater by 0.4  0.0 cm/s/mmHg compared to the PETCO2 MCAv reactivity (P<0.001).  NX1 Relative ICA Reactivity (%/mmHg)-6-4-20246810NX2 Relative ICA Reactivity (%/mmHg)-6-4-20246810††* *A. B. Hypocapnia HypocapniaHypercapnia Hypercapnia  55 For the NX2 trail the mean R2 values for HYPO-MCAv CVR was 0.99  0.0 for PETCO2 CVR, 0.98  0.0 for PaCO2 CVR, and 0.99  0.0 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-MCA CVR was 0.98  0.0 for PETCO2 CVR, 0.97  0.0 for PaCO2 CVR, and 0.99  0.0 for Pred-PaCO2 CVR.  During the NX2 protocol (Panel B), there was no difference in PaCO2, PETCO2, and Pred-PaCO2 MCAv reactivity in the hypocapnic range (P=0.283).  In the hypercapnic range, PaCO2 MCAv reactivity was greater by 1.2  0.4 cm/s/mmHg compared to PETCO2 MCAv reactivity, and by 0.8  0.3 cm/s/mmHg compared to the Pred-PaCO2 MCAv reactivity (P<0.001 and P=0.035, respectively), there was no difference between Pred-PaCO2 MCAv reactivity and PETCO2 MCAv reactivity (P=0.237).  HYPO-MCAv PaCO2 CVR and HYPER MCAv PaCO2 CVR was compared between PFO+ and PFO- participants, no differences during both NX1 and NX2 CO2 reactivity protocols for HYPO- (P=0.250 and P=0.846) and HYPER- (P=0.417 and P=0.741) MCAv PaCO2 CVR were found between these two groups.  Figure 3.10: MCA hypocapnia and hypercapnia reactivity data during NX1 (Panel A) and NX2 (Panel B) protocols. (black bar), MCA reactivity indexed against PETCO2; (white bar), MCA reactivity indexed against PaCO2; (grey bar), MCA reactivity indexed against Pred-PaCO2; calculated using our previously derived algorithm. †P<0.05, PaCO2 hypercapnic MCA reactivity vs. PETCO2 and Pred-PaCO2 hypercapnic ICA reactivity. *P<0.05, Pred-PaCO2 hypercapnic MCA reactivity vs PETCO2 MCA reactivity.  3.4.2 Hypoxia reactivity data 3.4.2.1 Hypercapnic ventilatory reactivity data Figure 3.11 displays the comparison of HCVR data when plotted against PETCO2, PaCO2, and Pred-PaCO2 (calculated using our previously derived correction algorithms, equation 5).  For the A. B. NX1 Relative MCA Reactivity (%/mmHg)-4-20246NX2 Relative MCA Reactivity (%/mmHg)-4-20246HypocapniaHypercapniaHypocapniaHypercapnia*††  56 HX1 trial the mean R2 values for HCVR was 0.99  0.01 for PETCO2 HCVR, 0.98  0.01 for PaCO2 HCVR, and 0.98  0.01 for Pred-PaCO2 HCVR.  During the HX1 reactivity protocol, PaCO2 HCVR was greater than PETCO2 HCVR by 0.4  0.2 l/min/mmHg, but not different from Pred-PaCO2 reactivity (P=0.007 and P=0.596, respectively), and there was no difference between Pred-PaCO2 HCVR and PETCO2 HCVR (P=0.062).  For the HX2 trial the mean R2 values for HCVR was 0.96  0.01 for PETCO2 HCVR, 0.95  0.02 for PaCO2 HCVR, and 0.96  0.01 for Pred-PaCO2 HCVR.  During the HX2 reactivity protocol there were was no difference between PaCO2, PETCO2 and Pred-PaCO2 HCVR (P=0.262). Hypercapnic ventilatory response to PaCO2 was also compared between PFO+ and PFO- participants (see Figure 3.12, pg 57), we found no differences in HCVR between PFO+ and PFO- participants during NX1 (P=0.505) and NX2 (P=0.257) CO2 reactivity protocols.   Figure 3.11: Hypercapnic ventilatory reactivity data during HX1 (Panel A) and HX2 (Panel B) protocols.  (black bar), HCVR indexed against PETCO2; (white bar), HCVR indexed against PaCO2; (grey bar), HCVR indexed against Pred-PaCO2; calculated using our previously derived algorithm. *P<0.05, PaCO2 HCVR vs. PETCO2 HCVR.   HX1 Ventilatory Reactivity (l/min/mmHg)0123456*HX2 Ventilatory Reactivity (l/min/mmHg)0123456A. B. PETCO2PaCO2Pred-PaCO2PETCO2PaCO2Pred-PaCO2  57  Figure 3.12: Hypercapnic ventilatory reactivity data during HX1 (Panel A) and HX2 (Panel B) protocols between PFO+ (n=8) and PFO- (n=6) participants.  (black bar), PFO+ HCVR indexed against PaCO2; (white bar), PFO- HCVR indexed against PaCO2.  3.4.2.2 Internal carotid artery reactivity Figure 3.13 displays relative HYPO-ICA and relative HYPER-ICA data during HX1 and HX2 CO2 reactivity protocols.  Due to incomplete data sets, four participants were removed from Q̇ICA analysis (n=10).  For the HX1 trial the mean R2 values for HYPO-ICA CVR was 0.93  0.02 for PETCO2 CVR, 0.92  0.02 for PaCO2 CVR, and 0.93  0.02 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-ICA CVR was 0.96  0.02 for PETCO2 CVR, 0.97  0.02 for PaCO2 CVR, and 0.98  0.03 for Pred-PaCO2 CVR.  During the HX1 (Panel A) protocol, there were no differences between PETCO2, PaCO2 and Pred-PaCO2 relative ICA reactivity in the hypocapnic range (P=0.563).  In the hypercapnic range, PaCO2 and Pred-PaCO2 relative ICA reactivity were greater by 0.9  0.3 %/mmHg, and 0.6  0.1 %/mmHg compared to PETCO2 relative ICA reactivity (P=0.001 and P=0.030, respectively), and no difference was found between PaCO2 HCVR and Pred-PaCO2 HCVR (P=0.330).   For the HX2 trial the mean R2 values for HYPO-ICA CVR was 0.93  0.02 for PETCO2 CVR, 0.93  0.03 for PaCO2 CVR, and 0.93  0.03 for Pred-PaCO2 CVR. The mean R2 values for HYPER-ICA CVR was 0.93  0.02 for PETCO2 CVR, 0.94  0.03 for PaCO2 CVR, and 0.94  0.03 for Pred-PaCO2 CVR.  During the HX2 protocol (Panel B), PETCO2 relative ICA reactivity was lower than PaCO2 relative ICA reactivity by 0.2  0.1 %/mmHg, and Pred-PaCO2 relative ICA reactivity by 0.2  0.0 %/mmHg (P=0.005 and P=0.004, respectively) in the hypocapnic range. In the hypercapnic range, PaCO2 relative ICA reactivity was greater by 0.8  0.4 %/mmHg HX1 Ventilatory Reactivity (l/min/mmHg)0123456HX2 Ventilatory Reactivity (l/min/mmHg)0123456A. B. PFO + PFO- PFO + PFO-  58 compared to PETCO2 relative ICA reactivity (P=0.027), and there was no difference between Pred-PaCO2 relative ICA reactivity and PaCO2 (P=0.605) and PETCO2 (P=0.173) relative ICA reactivity. HYPO-ICA PaCO2 CVR and HYPER ICA PaCO2 CVR was compared between PFO+ and PFO- participants, no differences during both HX1 and HX2 CO2 reactivity protocols for HYPO- (P=0.096 and P=0.083) and HYPER- (P=0.502 and P=0.694) ICA PaCO2 CVR were found between these two groups.  Figure 3.13: ICA hypocapnia and hypercapnia reactivity data during HX1 (Panel A) and HX2 (Panel B) protocols. (black bar), ICA reactivity indexed against PETCO2; (white bar), ICA reactivity indexed against PaCO2; (grey bar), ICA reactivity indexed against Pred-PaCO2; calculated using our previously derived algorithm. *P<0.05, PETCO2 hypocapnic ICA reactivity vs hypocapnic PaCO2 and Pred-PaCO2 ICA reactivity. †P<0.05, vs PETCO2 ICA reactivity.  3.4.2.3 Middle cerebral artery reactivity Figure 3.14 displays HYPO-MCAv and HYPER-MCAv data during HX1 and HX2 CO2 reactivity protocols.  For the HX1 trial the mean R2 values for HYPO-MCAv CVR was 0.98  0.01 for PETCO2 CVR, 0.98  0.02 for PaCO2 CVR, and 0.98  0.02 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-MCAv CVR was 0.98  0.02 for PETCO2 CVR, 0.97  0.02 for PaCO2 CVR, and 0.98  0.02 for Pred-PaCO2 CVR.  During the HX1 protocol (Panel A), PETCO2 relative MCAv reactivity was lower by -0.3  0.1 %/mmHg compared to PaCO2 relative MCAv reactivity in the hypocapnic range (P=0.015).  In the hypercapnic range, PaCO2 and Pred-PaCO2 relative MCAv reactivity were greater by 0.4  0.1 %/mmHg (P=0.001) and 0.3  0.1 %/mmHg (P=0.011) compared to PETCO2 relative MCAv reactivity.  HX1 Relative ICA Reactivity (%/mmHg)-6-4-20246810HX2 Relative ICA Reactivity (%/mmHg)-6-4-20246810A. B. Hypocapnia HypocapniaHypercapniaHypercapnia*†††  59 For the HX2 trial the mean R2 values for HYPO-MCAv CVR was 0.98  0.02 for PETCO2 CVR, 0.96  0.01 for PaCO2 CVR, and 0.98  0.02 for Pred-PaCO2 CVR.  The mean R2 values for HYPER-MCAv CVR was 0.96  0.02 for PETCO2 CVR, 0.98  0.02 for PaCO2 CVR, and 0.97  0.03 for Pred-PaCO2 CVR.  During the HX2 protocol (Panel B), PETCO2 relative MCAv reactivity was lower by 0.2  0.1 %/mmHg compared to PaCO2 relative MCAv reactivity (P<0.001), and by 0.2  0.0 %/mmHg compared to Pred-PaCO2 relative MCAv reactivity (P<0.001) in the hypocapnic range. In the hypercapnic range, no differences were detected between PaCO2, PETCO2, and Pred-PaCO2 relative MCAv reactivity (P=0.075).  HYPO-MCAv PaCO2 CVR and HYPER MCAv PaCO2 CVR was compared between PFO+ and PFO- participants, no differences during both HX1 and HX2 CO2 reactivity protocols for HYPO- (P=0.441 and P=0.141) and HYPER- (P=0.197 and P=0.094) MCAv PaCO2 CVR were found between these two groups.  Figure 3.14: MCAv hypocapnia and hypercapnia reactivity data during HX1 (Panel A) and HX2 (Panel B) protocols.(black bar), MCAv reactivity indexed against PETCO2; (white bar), MCAv reactivity indexed against PaCO2; (grey bar), MCAv reactivity indexed against Pred-PaCO2; calculated using our previously derived algorithm. *P<0.05, PETCO2 hypocapnic relative MCAv reactivity vs hypocapnic PaCO2 and Pred-PaCO2 MCAv reactivity. †P<0.05, vs PETCO2 MCAv reactivity.   Research Question Four: performance of our PaCO2 correction algorithm  3.5.1 Comparison between new and old prediction algorithms for PaCO2 Table 3.12 displays the linear regression correction algorithms for PaCO2 derived from previous work (118), and additional correction algorithms derived from the current study data based on data HX1 Relative MCA Reactivity (%/mmHg)-4-20246HX2 Relative MCA Reactivity (%/mmHg)-4-20246* *† † HypocapniaHypocapniaHypercapnia HypercapniaA. B.   60 from NX1 and HX1 CO2 reactivity protocols.  Among the variables included in the stepwise multiple regression model (see section Statistics, pg 29), the following were significant contributors to the prediction equation for PaCO2: PETCO2, Baseline Pa-PETCO2 gradient, and VT for both normoxia and hypoxia CO2 reactivity protocols (P<0.001).   3.5.2 Normoxia and hypoxia data Figure 3.15 and Figure 3.17 display the relationship between PaCO2 and PETCO2 raw data, and PETCO2 from the current study data that has been adjusted using an algorithm proposed by Tymko et al. (118) and Peebles et al. (87) for both normoxia and hypoxia trials.  In addition to this, the performance of our new linear regression algorithms derived from NX1 and HX1 trials were applied to our NX2 and HX2 data, see in Figure 3.16 and Figure 3.18.  3.5.3 Performance PaCO2 correction algorithm in normoxia and hypoxia Table 3.13 displays a summary of the performance of our PaCO2 correction algorithms for both normoxia and hypoxia CO2 reactivity trials.  Between the NX1 and NX2 trials, there were no differences between the Pa-PETCO2 and PET-PaO2 gradients (see section NX1 vs. NX2 protocols, pg 33).  When we applied our PaCO2 prediction algorithm during NX2 trial, there was no difference in our predicted PaCO2 and measured PaCO2 the +4 mmHg CO2 stage (P=0.894), however there was during the +8 mmHg CO2 stage (P<0.001).  Between the HX1 and HX2 trials, there were no differences between the Pa-PETCO2 and PET-PaO2 gradients (see section HX1 vs. HX2 CO2 protocols, pg 39).  When we applied our PaCO2 prediction algorithm during NX2 trial, there was no difference between our predicted PaCO2 and measured PaCO2 for both the +4 mmHg (P=0.403) and +8 mmHg (P=0.860) CO2 stage.  Comparing the end-tidal-to-arterial gradients between normoxia and hypoxia trials, no differences were found in the Pa-PETCO2 gradient between normoxia and hypoxia (P=0.738), while the PET-PaO2 gradient was significantly larger in hypoxia trials compared to normoxia (P<0.001) during the CO2 reactivity protocols (see section Comparison of normoxia and hypoxia CO2 protocols, pg 44).    61 Table 3.12: A comparison of linear regression correction algorithms for PaCO2 derived from Tymko et al., and the NX1 CO2 reactivity protocol.   Tymko et al. (2015) Linear Regressions  (1) PaCO2= 0.363 + (0.958 *PETCO2) R2 = 0.95; P<0.001  (3) PaCO2 = 0.964 + (0.960 * PETCO2) + (0.331 * Baseline Pa-PETCO2) R2 = 0.95; P<0.001  Linear Regressions (Current Study) Normoxia Hypoxia  (e1) PaCO2= 2.189 + (0.941 *PETCO2) R2 = 0.95; P<0.001  (e2) PaCO2 = 2.059 + (0.941 * PETCO2) + (0.174 * Baseline Pa-PETCO2) R2 = 0.95; P<0.001    (e3) PaCO2 = 3.216 + (0.950 * PETCO2) - (0.998 * VT) + (0.376 * Baseline Pa-PETCO2) R2 = 0.99; P<0.001   (e4) PaCO2 = 3.196 + (0.948 * PETCO2) - (0.768 * VT) R2 = 0.98; P<0.001   (e1) PaCO2 = 2.531 + (0.931 * PETCO2) R2= 0.97; P<0.001  (e2) PaCO2 = 2.293 + (0.929 * PETCO2) - + (0.338 * Baseline Pa-PETCO2) R2 = 0.98; P<0.001    (e3) PaCO2 = 2.271 + (0.960 * PETCO2) - (0.635 * VT) + (0.405 * Baseline Pa-PETCO2) R2 = 0.98; P<0.001   (e4) PaCO2 = 2.551 + (0.955 * PETCO2) - (0.493 * VT) R2 = 0.97; P<0.001     Definition of Abbreviations:  PaCO2, arterial partial pressure of carbon dioxide; PETCO2, end-tidal partial pressure of carbon dioxide; Baseline Pa-PETCO2, baseline end-tidal-to-arterial carbon dioxide gradient; VT, tidal volume  62  Figure 3.15: Assessment of PaCO2 and PETCO2 relationship of CO2 during NX1 and NX2 CO2 reactivity protocols.   Data points were obtained during the last minute of each step change in PETCO2.  Dotted lines represent 95% confidence intervals.  Panel A, represents pooled linear regression for PaCO2 and PETCO2.  Panel B, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm proposed by Tymko et al. in 2015 (eTymkoPaCO2 = 0.363 + (0.958 *PETCO2).  Panel C, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted with an algorithm proposed by Peebles et al. in 2007 (ePeeblesPaCO2 = 2.367 + 0.884* PETCO2). b[1], slope; b[0], y-intercept.  Red line indicates a line of identity (i.e. x = y)  NX1NX225 30 35 40 45 50 55PETCO2 (mmHg)25303540455055R2 = 0.97b[1] = 1.04b[0] = -1.225 30 35 40 45 50 55eTymkoPaCO2 (mmHg)25303540455055R2 = 0.97b[1] = 0.99b[0] = -0.825 30 35 40 45 50 55ePeeblesPaCO2 (mmHg)25303540455055R2 = 0.97b[1] = 0.91b[0] = 1.3PaCO2 (mmHg)25 30 35 40 45 50 55PETCO2 (mmHg)25303540455055R2 = 0.96b[1] = 1.04b[0] = -1.1A. C.B.PaCO2 (mmHg)25 30 35 40 45 50 55eTymkoPaCO2 (mmHg)25303540455055R2 = 0.96b[1] = 0.99b[0] = -0.65PaCO2 (mmHg)25 30 35 40 45 50 55ePeeblesPaCO2 (mmHg)25303540455055R2 = 0.96b[1] = 0.91b[0] = 1.4  63  Figure 3.16: Assessment of PaCO2 and PETCO2 relationship of CO2 during NX2 CO2 reactivity protocols, using new algorithms derived from NX1 CO2 reactivity protocol.  Data points were obtained during the last minute of each step change in PETCO2.  Dotted lines represent 95% confidence intervals.  Panel A, represents pooled linear regression for PaCO2 and PETCO2.  Panel B, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm derived from NX1 CO2 reactivity protocol data (e3PaCO2 = 3.216 + (0.950 * PETCO2) - (0.998 * VT) + (0.376 * Baseline Pa-PETCO2)). Panel C, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm derived from NX1 CO2 reactivity protocol data (e4PaCO2 = 3.196 + (0.948 * PETCO2) - (0.768 * VT)). Panel D, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm (e1PaCO2 = 2.189 + (0.941 * PETCO2)). b[1], slope; b[0], y-intercept.  Red line indicates a line of identity (i.e. x = y)   PaCO2 (mmHg)25 30 35 40 45 50 55e1PaCO2 (mmHg)25303540455055PaCO2 (mmHg)25 30 35 40 45 50 55e4PaCO2 (mmHg)2530354045505525 30 35 40 45 50 55e3PaCO2 (mmHg)2530354045505525 30 35 40 45 50 55PETCO2 (mmHg)25303540455055R2 = 0.96b[1] = 1.04b[0] = -1.06R2 = 0.97b[1] = 1.03b[0] = -0.75R2 = 0.96b[1] = 1.01b[0] = -0.07A.C.B.R2 = 0.96b[1] = 0.98b[0] = 1.19D.  64   Figure 3.17: Assessment of PaCO2 and PETCO2 relationship of CO2 during HX1 and HX2 CO2 reactivity protocols.   Data points were obtained during the last minute of each step change in PETCO2.  Dotted lines represent 95% confidence intervals.  Panel A, represents pooled linear regression for PaCO2 and PETCO2.  Panel B, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm proposed by Tymko et al. in 2015 (eTymkoPaCO2 = 0.363 + (0.958 *PETCO2).  Panel C, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted with an algorithm proposed by Peebles et al. in 2007 (ePeeblesPaCO2 = 2.367 + 0.884* PETCO2). b[1], slope; b[0], y-intercept.  Red line indicates a line of identity (i.e. x = y).   PaCO2 (mmHg)25 30 35 40 45 50 55ePeeblesPaCO2 (mmHg)25303540455055PaCO2 (mmHg)25 30 35 40 45 50 55eTymkoPaCO2 (mmHg)25303540455055PaCO2 (mmHg)25 30 35 40 45 50 55PETCO2 (mmHg)2530354045505525 30 35 40 45 50 55ePeeblesPaCO2 (mmHg)2530354045505525 30 35 40 45 50 55eTymkoPaCO2 (mmHg)25303540455055HX1HX225 30 35 40 45 50 55PETCO2 (mmHg)25303540455055R2 = 0.97b[1] = 1.04b[0] = -1.3R2 = 0.97b[1] = 0.99b[0] = -0.91R2 = 0.97b[1] = 0.91b[0] = 1.2R2 = 0.96b[1] = 1.02b[0] = -0.75A. C.B.R2 = 0.96b[1] = 0.98b[0] = -0.35R2 = 0.96b[1] = 0.90b[0] = 1.7  65  Figure 3.18: Assessment of PaCO2 and PETCO2 relationship of CO2 during HX2 CO2 reactivity protocols, using new algorithms derived from HX1 CO2 reactivity protocol.  Data points were obtained during the last minute of each step change in PETCO2.  Dotted lines represent 95% confidence intervals.  Panel A, represents pooled linear regression for PaCO2 and PETCO2.  Panel B, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm derived from HX1 CO2 reactivity protocol data (e3PaCO2 = 2.271 + (0.960 * PETCO2) - (0.635 * VT) + (0.405 * Baseline Pa-PETCO2)).  Panel C, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm derived from HX1 CO2 reactivity protocol data (e4PaCO2 = 2.551 + (0.955 * PETCO2) - (0.493 * VT)).  Panel D, represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm derived from HX1 CO2 reactivity protocol data (e1PaCO2 = 2.531 + (0.931 * PETCO2)). b[1], slope; b[0], y-intercept.  Red line indicates a line of identity (i.e. x = y).    PaCO2 (mmHg)25 30 35 40 45 50 55e1PaCO2 (mmHg)25303540455055PaCO2 (mmHg)25 30 35 40 45 50 55e4PaCO2 (mmHg)2530354045505525 30 35 40 45 50 55e3PaCO2 (mmHg)2530354045505525 30 35 40 45 50 55PETCO2 (mmHg)25303540455055R2 = 0.97b[1] = 1.02b[0] = -0.75R2 = 0.98b[1] = 0.96b[0] = 1.60R2 = 0.97b[1] = 0.95b[0] = 1.73A.C.B.R2 = 0.97b[1] = 0.95b[0] = 1.83D.  66 Table 3.13: Summary of the performance of our PaCO2 correction algorithm during normoxia and hypoxia trials.   AB-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg NX1 Pa-PETCO2 gradient -0.3 ± 0.2 0.5 ± 0.2 -0.2 ± 0.3 0.3 ± 0.2 -0.7 ± 0.2† -1.1 ± 0.2†  PET-PaO2 gradient 3.5 ± 0.9† 4.1 ± 0.9† 3.5 ± 0.8† 4.7 ± 0.9† 2.9 ± 0.6† 2.1 ± 0.4†         NX2 Pa-PETCO2 gradient -0.3 ± 0.3 0.1 ± 0.4 0.1 ± 0.3 0.0 ± 0.3 -0.8 ± 0.2† -1.9 ± 0.3†  PET-PaO2 gradient 5.7 ± 0.7† 3.1 ± 1.0† 2.9 ± 1.0† 4.8 ± 0.8† 3.6 ± 0.7† 1.0 ± 0.8  Pred-PaCO2 – actual PaCO2 -0.4 ± 0.2 -0.1 ± 0.4 -0.4 ± 0.3 -0.7 ± 0.2 0.0 ± 0.2 -1.1 ± 0.2*           HX-DEF -8 mmHg -4 mmHg 0 mmHg +4 mmHg +8 mmHg HX1 Pa-PETCO2 gradient -0.2 ± 0.3 0.4 ± 0.3 0.2 ± 0.2 -0.2 ± 0.2 -0.7 ± 0.3† -0.9 ± 0.3†  PET-PaO2 gradient 6.7 ± 0.5† 5.6 ± 0.5† 6.4 ± 0.3† 6.7 ± 0.7 5.9 ± 0.4† 6.1 ± 0.6†         HX2 Pa-PETCO2 gradient -0.4 ± 0.3 0.5 ± 0.2 -0.1 ± 0.3 -0.1 ± 0.2 -0.5 ± 0.3† -0.7 ± 0.3†  PET-PaO2 gradient 5.5 ± 0.6† 6.0 ± 0.7† 6.5 ± 0.7† 6.4 ± 0.6 6.3 ± 0.6† 5.5 ± 0.6†  Pred-PaCO2 – actual PaCO2 -0.3 ± 0.2 -0.2 ± 0.2 -0.3 ± 0.3 -0.4 ± 0.1 -0.2 ± 0.2 0.0 ± 0.3 Definition of Abbreviations: AB-DEF, air breathing baseline controlled with our DEF system (normoxia baseline 2); HX-DEF, isocapnic hypoxia breathing controlled with our DEF system (hypoxia baseline 2); PaCO2, arterial partial pressure of carbon dioxide; PETCO2, end-tidal partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial carbon dioxide gradient; PET-PaO2, end-tidal-to-arterial oxygen gradient; Pred-PaCO2, predicted PaCO2, calculated using our correction algorithm from Tymko et al. 2015: PaCO2= 0.363 + (0.958 *PETCO2). †P<0.05, presence of an end-tidal-to-arterial gradient. *P<0.05, Pred-PaCO2 vs actual PaCO2 at +8 mmHg during NX2 protocol.   67 Chapter 4: Discussion  Summary of main findings The purpose of the current study was to (I) measure any differences in the end-tidal-to-arterial gradient between normoxia and hypoxia CO2 reactivity protocols, (II) determine whether having a PFO had an effect on both CO2 and O2 end-tidal-to-arterial gradients, (III) quantify differences (if any) between HCVR and CVR to CO2 when indexed against PETCO2 and PaCO2 in normoxia and hypoxia, and (IV) determine whether our previously derived PaCO2 correction algorithms accurately predict PaCO2 during DEF.  The primary findings from this study were (I) there were no differences in the Pa-PETCO2 gradient between normoxia and hypoxia, however, the PET-PaO2 gradient was larger in hypoxia compared to normoxia during a CO2 reactivity test, (II) presence of a PFO increased the PET-PaO2 gradient during the normoxia CO2 reactivity protocols, but this effect was not observed during the hypoxia CO2 reactivity protocols, (III) HCVR and CVR (MCAv and Q̇ICA) to CO2 profiles were significantly greater when plotted against PaCO2 compared to PETCO2 during both normoxia and hypoxia CO2 reactivity protocols, and (IV) our previously derived PaCO2 correction algorithm was successful at accounting for the PET-PaCO2 gradient during the normoxia CO2 reactivity protocol at +4 mmHg, and hypoxia CO2 reactivity protocol at both +4 and +8 mmHg PETCO2 steps.   Research Question One: any differences in the end-tidal-to-arterial gradients between normoxia and hypoxia? Despite the advantages of using a DEF system, particularly our system as it is portable, cost effective due to low gas requirements, and capable of controlling end-tidal gases within narrow limits, it, similar to other systems is affected by methodological constraints.  Arguably the most recognized methodological constraint is that similar to all DEF systems, our DEF system assumes that end-tidal gases are a surrogate measure for arterial blood gases, in which we know that there normally is a gradient between end-tidal and arterial blood gases.  4.2.1 The end-tidal-to-arterial CO2 gradient  There are a number of experimental and clinical conditions where PETCO2 does not accurately reflect PaCO2 such as changes in body position (14), age (80), exercise (59), low breathing   68 frequencies (56), and with CO2 administration (i.e. hypercapnia) (3, 93, 118).  Similar to previously reported data (20, 93, 127), our study found that there was a positive Pa-PETCO2 gradient during eupneic air breathing, not controlled using DEF (Baseline 1, AB).  This was in contrast to previously published work (118), in which the PaCO2 prediction algorithms were previously derived from (see section End-tidal vs arterial CO2?, pg 16).  In the previously published work, a negative Pa-PETCO2 gradient was observed in the participants during eupneic air breathing.  The negative Pa-PETCO2 gradient was attributed to the presence of a large apparatus deadspace within the breathing circuit (~250ml of deadspace), which adds to physiological deadspace, and the resulting trapped CO2 within the apparatus deadspace was inhaled with each breath.  However, exactly the same breathing apparatus from previous work was used during the current study suggesting that apparatus deadspace was not the reason for the measured resting negative Pa-PETCO2 gradients (see 2.5.2, pg 24 for details on our breathing apparatus).  Perhaps a better explanation was that a large negative Pa-PETCO2 gradient was present in three of nine subjects, likely shifting the mean bias enough to become statistically significant (118).   A novel finding was that there was a significant decrease in the Pa-PETCO2 gradient between the unforced air breathing baseline (baseline 1, AB), and the forced air breathing baseline (baseline 2, AB/HX-DEF) in both normoxia and hypoxia CO2 reactivity protocols, meaning that DEF in itself had an effect on the Pa-PETCO2 gradient.  This is attributed to the increases in V̇E during DEF air breathing, which led to a greater PICO2 that occupies physiological deadspace, possibly elevating end-tidal values upon expiration.  Although participants are being controlled at the same PETCO2 and PETO2 during the DEF baseline, almost all participants increased their V̇E slightly due to pseudo-stimulation probably from the noise made by the DEF solenoid valves when triggered.  By this logic, one would expect that the Pa-PETCO2 gradient would be larger during the hypoxic CO2 reactivity protocol during DEF isocapnic hypoxia air breathing compared to normoxia, however, this was not observed, despite an increase in PICO2.  Consistent with previous findings (118), no Pa-PETCO2 gradient was detected during normoxia CO2 reactivity protocols in the hypocapnic range, and this also held true during the hypoxia CO2 reactivity protocols.  During the mild hypercapnic steps (+4 and +8 mmHg PETCO2), a negative Pa-PETCO2 gradient (PETCO2 overestimating PaCO2) was present in both   69 normoxia and hypoxia (refer to Bland and Altman plots, see Figure 3.2 and Figure 3.5), and in opposition to one of our hypotheses, there was no difference in the Pa-PETCO2 gradients between normoxia and hypoxia CO2 reactivity protocols.   In Tymko et al. (118), a widening (i.e. more negative) Pa-PETCO2 gradient was observed at HA compared to LA.  Initially, it was thought that the effect was simply due to the heightened ventilatory response associated with hypobaric hypoxia (atm ~413 mmHg), which would require an increased FICO2 in order to maintain a given PETCO2 during DEF.  This effect paired with the potential increase in alveolar deadspace at HA due to HPV could be responsible for the difference in the Pa-PETCO2 gradient between LA and HA (67, 118).  In the current study, an acute isocapnic hypoxia bout (10-minutes) was administered prior to conducting the CO2 reactivity test in order to elicit a HPV response, similar to what may be observed at HA (116, 118), to increase alveolar deadspace.  However, even though V̇E mildly increased during hypoxic hypercapnia compared to normoxic hypercapnia, there was no difference in the Pa-PETCO2 gradient between the two protocols.  We speculate that the reason behind the negative finding was that we failed to deliver a powerful enough respiratory stimulus in order to achieve a difference in the Pa-PETCO2 gradient between normoxia and hypoxia CO2 protocols.  Although V̇E increased between the normoxia and hypoxia trials, PICO2 remained the same.  Looking back at previous data (118), there was a difference in the Pa-PETCO2 gradient between LA and HA when the largest stimulus was delivered during hypercapnia (+10 mmHg PETCO2), and the V̇E response was nearly 2-fold greater at HA compared to LA.  The V̇E response at HA necessitated a FICO2 delivery of 9.1  0.1% compared to 6.7  0.1% at LA.  In order to see a difference in the Pa-PETCO2 gradient between normoxia and hypoxia trials, we might have needed to deliver a more severe hypoxic stimulus.  However, the knowledge that the Pa-PETCO2 gradient remains unchanged with a ~15-20 l/min difference in V̇E between the normoxia and hypoxia CO2 reactivity protocols is extremely valuable for future applications of our DEF system.  4.2.2 The end-tidal-to-arterial O2 gradient Surprisingly, there is little literature available with respect to the PET-PaO2 gradient during changes in PETCO2 conducted by DEF.  Although experiments investigating the AaDO2 exist, it is difficult to assess the AaDO2 during DEF due to breath-by-breath fluctuations in inspired   70 fractions of O2 and CO2, necessitating the study of the PET-PaO2 gradient in its place.  Literature regarding the AaDO2 is inconsistent suggesting that the AaDO2 can widen or narrow following hypoxia (67, 91, 115).  During the normoxia trials the PET-PaO2 gradient did not change between eupneic unforced air breathing (AB; baseline 1) compared to room air breathing controlled with the DEF system (AB-DEF; baseline 2).  Throughout the normoxic CO2 reactivity protocol, the PET-PaO2 gradient remained unchanged from the DEF baseline with the exception of +8 mmHg PETCO2 stage where the PET-PaO2 gradient was significantly reduced compared to the AB-DEF baseline.  This novel observation was not expected and it may be due to a combination of (I) increased V̇/Q matching from hypercapnic induced pulmonary vasoconstriction, and (II) decreased administered PIO2 due to the increased V̇E during hypercapnia.  The latter theory is the most probable, as during hyperventilation less PIO2 is required to maintain a given PETO2 level, minimizing the difference between PETO2 and PaO2.  With this reasoning one would expect to see a decrease in the PET-PaO2 gradient in hypocapnia as well, and although this was not significant, the PET-PaO2 gradient was smaller during hyperventilation-induced hypocapnia, giving further support to this theory.  In contrast, this effect of the PET-PaO2 gradient decreasing during hypercapnia was not observed during our hypoxia CO2 reactivity trials.  In addition, the PET-PaO2 gradient was larger during hypoxia compared to the normoxia CO2 reactivity protocols.  This supports the notion that our acute isocapnic hypoxia stimulus increased alveolar deadspace within the lung.   4.2.3 Targeting arterial blood gases Currently, there are two sophisticated approaches to target arterial blood gases using computer-controlled systems.  One is the traditional DEF system, like our research group, and improving arterial blood gas targeting using linear regression algorithms (118), or two, by employing the RespirAct, a sequential gas delivery system (56, 106).  The sole purpose behind developing the RespirAct was to account for the discrepancy between end-tidal and arterial gases (106).  One of the fundamental features of the RespirAct is that the end-tidal-to-arterial gas gradient is minimized by filling physiological deadspace with previously expired gas (considered gas exchange neutral) rather than “fresh” inspirate (56, 106) which aims to control anatomical and alveolar deadspace gas content.  However, it works most effectively during paced ventilation, limiting its utility during exercise testing and during experiments investigating the control of   71 breathing (44).  The RespirAct was validated in a study that only recruited five participants, four of whom were over the age of 40 yrs, one of these participants had asthma, and one had a history of smoking, raising concern as to the validity of this study.  The RespirAct validation study involved manipulations in PETCO2 and PETO2 while coaching participants to breathe at different rates.  Compared to our DEF system (118) the Respiract is arguably less effective at targeting and controlling end-tidal gases, based on the results of its validation study (56), and the results from our DEF system validation study (118).   Research Question Two: were there any differences in the end-tidal-to-arterial gradients between PFO+ and PFO- participants? Within the circulatory system several potential intracardiac shunt pathways exist.  These pathways provide a low resistance pathway for blood to bypass the gas exchange sites of the lung, resulting in a reduction in pulmonary gas exchange efficiency.  The most common right-to-left shunt pathway is the PFO.  The formen ovale is a small flap-like opening in the atrial septum, which normally provides a direct pathway for blood between the right and left atria, important in the fetal circulation, but usually closes shortly after birth.  This potentially results in a widening of the alveolar-to-arterial CO2 gradient, and more likely, a widening in the AaDO2.  Changes in the alveolar-to-arterial gradient will likely also affect the end-tidal-to-arterial gradient.  Given that the DEF technique functions based on the assumption that end-tidal are a surrogate for arterial blood gases, it is prudent to account for any potential differences between end-tidal and arterial blood gases and determine whether DEF is a suitable option for controlling end-tidal gases in this population.  4.3.1 End-tidal-to-arterial carbon dioxide gradient Although possible, it is unlikely that a PFO would have a significant effect on the Pa-PETCO2 gradient, unless it was considerably large.  In the current study, participants that presented with a PFO were only patent upon Valsalva maneuver and were small (i.e. shunt scores of “1” and “2”).  In normoxia and hypoxia CO2 reactivity protocols, the Pa-PETCO2 gradient was the same between PFO+ and PFO- participants.      72 4.3.2 End-tidal-to-arterial oxygen gradient Due to the large resting gradient of O2 between atmospheric air and venous blood returning to the lung, it was more likely that the presence of a PFO may alter the PET-PaO2 gradient.  We found that the PET-PaO2 gradient was significantly larger in PFO+ participants compared to PFO- participants during the normoxia CO2 reactivity protocols.  This was unexpected due to the small size of the PFO’s that were screened, but these participants’ foramen ovale may have become more patent during increased thoracic pressures seen with increased V̇E during active hyperventilation in order to achieve hypocapnia, and during hypercapnia.  With this reasoning one would expect the same result in the hypoxia trials, however, this was not the case as the PET-PaO2 gradient was the same between PFO+ and PFO- participants during the hypoxia trials.  This was likely due to the narrowing of the gradient between the amount of O2 administered to maintain a PETO2 of 50 mmHg, and the venous blood returning to the lung (37, 118).  4.3.3 Effect of PFO on hypercapnic ventilatory reactivity Recent evidence suggests that there may be differences in the HCVR between PFO+ and PFO- populations (37).  This could be the result of differences in the Pa-PETCO2 gradient and/or the PET-PaO2 gradient.  If a difference in the PET-PaO2 gradient between PFO+ and PFO- participants was present during the hypoxia trials (PETO2 = 50 mmHg) where PaO2 lies near the steep portion of the oxygen-dissociation curve, it could affect HX-HCVR due to the additive effects of hypercapnia and hypoxia on V̇E (45, 47, 68).  We found in our study that there was no difference in the HCVR and HX-HCVR between PFO+ and PFO- participants in normoxia and hypoxia.  This was expected since we measured no difference in the PET-PaO2 gradient between PFO+ and PFO- participants during hypoxia CO2 reactivity protocols, and only a small gradient was found during normoxia CO2 reactivity protocols.     Research Question Three: are calculated cerebrovascular reactivity and ventilatory reactivity to CO2 attenuated by using PETCO2 instead of PaCO2? Cerebrovascular reactivity to CO2 is almost always quantified by plotting cerebral blood flow against either PETCO2 or PaCO2.  As described in section 1.3.1 and 1.3.2, quantifying CVR   73 and HCVR to PETCO2 during DEF may potentially lead to data misinterpretation, as PETCO2 does not uniformly reflect PaCO2.  As presented in Illustration 1.5 (pg 16), the HCVR can be potentially misinterpreted if indexed against PETCO2 as opposed to PaCO2, as PETCO2 overestimates PaCO2 in the hypercapnic range (118).  This same effect can be observed with CVR to CO2 (see Illustration 1.6, pg 17) during hypercapnia.  However, during hypocapnia, PETCO2 seems to be a reliable estimate of PaCO2, avoiding this potential confound (87, 118).   4.4.1 Hypercapnic ventilatory response in normoxia and hypoxia The normoxic HCVR was indexed against PETCO2, PaCO2, and Pred-PaCO2, which was calculated based on a previously derived correction algorithm (see equation 3).  As expected, HCVR was greater when plotted against PaCO2 compared to PETCO2 for both NX1 and NX2 CO2 reactivity protocols.  However, our results were inconsistent when plotted against Pred-PaCO2, indicating that our correction algorithm was not reliable when predicting PaCO2 HCVR.  These results were observed by Peebles et al. (87) when they conducted a HCVR test using two levels of administered CO2 delivered with a Douglas bag, however these results have been yet to be validated using DEF, and during hypoxia (87).  Similar results were found between normoxia and hypoxia CO2 reactivity protocols, in which HX-HCVR was greater when plotted against PaCO2 compared to PETCO2, however, these results were inconsistent since this effect was not seen in the HX2 trial.  A possible explanation of this is that the subject variability in HCVR reactivity is greater during the hypoxia CO2 reactivity protocol, leading to inconsistent results.   4.4.2 Middle cerebral artery velocity reactivity in normoxia and hypoxia Middle cerebral artery blood velocity was indexed against PaCO2, PETCO2, and Pred-PaCO2.  During both normoxia and hypoxia CO2 reactivity protocols, hypocapnic PETCO2 CVR to CO2 was lower compared to hypocapnic PaCO2 CVR to CO2.  This is in contrast to findings by Peebles et al. (87), where they found no differences in CVR to CO2 in the hypocapnic range.  A possible explanation for this is the different methodology used between the two studies, as we used a custom DEF system to finely control hypocapnia, while they solely relied upon active hyperventilation to achieve their desired levels of hypocapnia.  However,   74 Pred-PaCO2 and PaCO2 HYPO-CVR were the same in hypocapnia, rendering our prediction algorithm useful during hypocapnia when predicting MCAv PaCO2 CVR.  Similar to Peebles et al. (87), PaCO2 CVR was higher compared to PETCO2 CVR during the normoxia CO2 reactivity protocol.  This was expected during hypercapnia as a significant Pa-PETCO2 gradient was present, whereas there was not a significant Pa-PETCO2 gradient in the hypocapnic range.  The same effect was observed during the HX1 hypoxia CO2 reactivity protocol, but surprisingly, not for the HX2 hypoxia CO2 reactivity protocol, this could be attributed to the slightly smaller, although not significantly smaller, Pa-PETCO2 gradient observed during the HX2 compared to the HX1 CO2 reactivity protocol.   4.4.3 Internal carotid artery blood flow reactivity in normoxia and hypoxia To our knowledge this is the first study comparing relative Q̇ICA responses when indexing against PaCO2, PETCO2, and Pred-PaCO2.  During the normoxia and hypoxia CO2 reactivity protocol, there was no difference between hypocapnic PETCO2 CVR and PaCO2 CVR to CO2, with the exception of a lower PETCO2 CVR in one of the hypoxia CO2 reactivity protocols.  Despite this one difference, these results were expected as there was no Pa-PETCO2 gradient present during hypocapnia.  HYPER-Q̇ICA PaCO2 CVR was higher compared to PETCO2 CVR, this was consistent and expected throughout both normoxia and hypoxia trials as a significant Pa-PETCO2 gradient was present. HYPER-Q̇ICA Pred-PaCO2 CVR was greater than PETCO2 CVR in normoxia and hypoxia trials. However, Pred-PaCO2 CVR was not the same as PaCO2 CVR in all cases, meaning that our prediction equation is more accurate that PETCO2 CVR but slightly underestimates PaCO2 CVR.    Research Question Four: does our previously derived correction algorithm accurately predict PaCO2? One of the primary purposes of this study was to validate our previously derived correction algorithm (equation 3) that involved invasive measurements (i.e. arterial blood sampling).  We conducted two normoxia and two hypoxia CO2 reactivity protocols, and during the second protocol we applied our PaCO2 prediction algorithm in an attempt to take into account the Pa-  75 PETCO2 gradient.  In addition to this, we applied one of our non-invasive correction algorithms (equation 1) to our raw data to determine its performance.  4.5.1 Performance of previously derived PaCO2 invasive correction algorithms We quantified the performance of our algorithm by comparing the predicted PaCO2 value (our target) and the actual PaCO2 value measured.  The only Pa-PETCO2 gradient present during DEF was in the hypercapnic range, which we attempted to account for.  During the normoxia CO2 reactivity protocols, an Pa-PETCO2 gradient was present during both hypercapnic steps (+4 and +8 mmHg PETCO2 from baseline).  Our prediction algorithm successfully accounted for the Pa-PETCO2 gradient as there was no difference between Pred-PaCO2 and actual PaCO2 measured at +4 mmHg PETCO2.  However, we were unsuccessful at predicting the gradient at the most severe level of hypercapnia (+8 mmHg PETCO2), where there was a significant difference between Pred-PaCO2 and actual PaCO2.  Although the Pa-PETCO2 gradient was not predicted during the +8 mmHg PETCO2 step in normoxia, for reasons unknown, a larger, although not statistically significant, Pa-PETCO2 gradient presented itself during NX2 compared to the NX1 CO2 reactivity protocol at +8 mmHg PETCO2.  In light of this, the difference between the Pred-PaCO2 and actual PaCO2 was less than the Pa-PETCO2 gradient, meaning that our PaCO2 correction algorithm was successful to some extent.   During the hypoxia CO2 reactivity protocols, we were successful at predicting PaCO2 using our correction algorithm during both +4 mmHg and +8 mmHg PETCO2 stages, as there was no difference between Pred-PaCO2 and actual PaCO2.  Collectively, between the results for the normoxia and hypoxia CO2 reactivity protocols our invasive PaCO2 correction algorithm demonstrated its utility during DEF.  4.5.2 Performance of previously derived PaCO2 non-invasive correction algorithms Non-invasive correction algorithms have been proposed for predicting PaCO2 by Tymko et al. (118) (see equation 1) during a similar CO2 reactivity test using the same DEF system, and Peebles et al. (87) during hypocapnia (through active hyperventilation) and hypercapnia (increased fixed FICO2 inspirate).  As highlighted in Figure 3.15 and Figure 3.17, it was found that the correction algorithms proposed by Peebles et al. (87) could not accurately predict PaCO2 during the DEF protocols in normoxia and hypoxia, and this was likely due to the   76 different experimental conditions.  However, the non-invasive equation proposed by Tymko et al. (118) significantly improved both slope (b[1]) and y-intercept (b[0]) during both the normoxia and hypoxia CO2 reactivity protocols when plotting PETCO2 vs PaCO2, suggesting that this algorithm is useful when predicting PaCO2 based on PETCO2 during a CO2 reactivity test during DEF, and specifically advantageous as it does not require invasive measures.   4.5.3 Performance of newly derived PaCO2 correction algorithms Although the invasive and non-invasive algorithms derived by Tymko et al. (118) were successful, in an attempt to further improve the prediction of PaCO2, we ran multiple stepwise regression analyses to determine which variables would predict PaCO2 in both normoxia and hypoxia CO2 reactivity protocols.  These algorithms were derived based upon NX1 and HX1 CO2 reactivity protocol data (see Table 3.12, pg 61) and to determine their performance in Pred-PaCO2 they were applied to the NX2 and HX2 CO2 reactivity protocol data.  During the NX2 protocol the e4PaCO2 (e4PaCO2 = 3.196 + (0.948 * PETCO2) - (0.768 * VT)) algorithm made the greatest improvement to both b[1] and b[0]. During the hypoxia trial, all of the algorithms based on NX1 data were not effective when applied to the NX2 data, as there was no improvement in both b[1] and b[0] (see Figure 3.18).     Methodological considerations. One of the primary purposes of this research was to consider a fundamental caveat with the DEF technique; the assumption that end-tidal gases are a direct reflection of arterial blood gases.  The following sections are the methodological considerations for the current study, excluding the methodological considerations that are involved with DEF (see Methodological considerations with dynamic end-tidal forcing, pg 5).    4.6.1 Utility of our PaCO2 invasive correction algorithm One of the prediction algorithms we used (see equation 5) showed promise in predicting PaCO2 based on PETCO2 and the individuals’ baseline Pa-PETCO2 gradient.  Although this equation proved to be valuable by accounting for the Pa-PETCO2 gradient, it may not be as impactful since it involves either (I) the insertion of an arterial blood gas catheter, or (II) taking a reliable baseline arterial blood measurement (i.e. arterial puncture) while simultaneously collecting   77 respiratory measurements to calculate the Pa-PETCO2 gradient.  The latter is difficult to achieve due to the likelihood of participant hyperventilation from discomfort.  In the event where arterial blood measurements are not plausible, our non-invasive PaCO2 correction algorithm also showed utility in the current study (see equation 1).   4.6.2 The use of transcranial Doppler ultrasound to measure cerebral blood flow Another methodological consideration is the use of TCD. Transcranial Doppler ultrasound is a useful tool to obtain continuous measures of blood velocity in intracranial vessels in human participants.  However, velocity is only an index of flow if the diameter of the conduit remains constant, and only downstream arterioles are dilating (129).  Although some studies suggest that this assumption is valid in a narrow range (99, 120), recent studies suggest that the MCA may dilate across a narrow range of CO2 (26) or at more extreme values of O2 and CO2 (131), suggesting that velocity measures may underestimate flow during more extreme CO2 challenges (4).  Although TCD was employed in the current study, we also accounted for any changes in arterial diameter by conducting continuous hand-held measurements of volumetric flow within the ICA.   4.6.3 Reproducibility of arterial blood gas measurements Perhaps the most concerning criticism of our methodology was the reliability of the arterial blood gas measurements.  Figure 4.1 below, displays Bland and Altman plots showing the agreement between two arterial blood gas measurements taken on the exact same sample for PaCO2 and PaO2.  Taking into consideration that inter-variability between blood gas analyzers the approximate variation between the two measurements with our blood analyzer was 2 mmHg and 4 mmHg for PaCO2 and PaO2, respectively.  The magnitude of the mean Pa-PETCO2 gradient is well within the variability of the blood gas machine, however, the mean bias of each measurement is near zero. The root mean squared coefficient of variation was 1.45% and 1.66% for PaCO2 and PaO2, respectively   78  Figure 4.1: Bland and Altman plot for agreement between two consecutive arterial blood gas measurements (on the same sample) for CO2 (A) and O2 (B) during eupneic air breathing (n=30). Dotted lines represent the 95% confidence intervals and the continuous lines represent the mean bias.   4.6.4 Nasopharyngeal temperature as a surrogate for core body temperature Traditionally, core body temperature is usually measured using esophageal or rectal body temperature.  Given the nature of the experimental protocols where the CO2 stages were only four minutes in length, it is likely that any changes in core body temperature would not be reflected in rectal temperature due to time delay.  Initially, the methodology involved measuring esophageal temperature, however it was discovered that esophageal temperature was not an accurate measure of core body temperature as during increases in V̇E associated with the DEF CO2 reactivity protocol esophageal temperature would decrease by approximately 5-6 C, therefore it was found that nasopharyngeal temperature was a more appropriate measure.  Nasopharyngeal temperature has been considered a reliable measure of core body temperature (95), and likely a good measure of brain temperature as the ICA is located nearby (71).   Perspective and Significance. We hypothesized that (1) the Pa-PETCO2 would be larger in the background of hypoxia as opposed to normoxia during a CO2 reactivity test, (2) the PET-PaCO2 would remain unchanged, while the PET-PaO2 gradient would be greater in those with a PFO, (3) the HCVR and CVR would be lower when plotted against PETCO2 compared to PaCO2, and (4) a PaCO2 prediction Mean PaCO2 36 38 40 42 44 46PaCO2 measure 1 - PaCO2 meaure 2-6-4-20246Mean PaO2 80 84 88 92 96 100PaO2 measure 1 - PaO2 measure 2-6-4-20246A. B.  79 algorithm will appropriately correct for the Pa-PETCO2.  In agreement with our hypotheses, we demonstrated that the PET-PaCO2 remained unchanged, while the PET-PaO2 gradient was greater in those with a PFO during normoxic but not hypoxic CO2 reactivity protocol, (II) the HCVR and CVR was greater when indexed against PaCO2 compared to PETCO2, and (III) our previously derived invasive and non-invasive PaCO2 prediction algorithms appropriately predicted PaCO2.  Our hypotheses were not supported in that we found that the Pa-PETCO2 gradient was not different between normoxia and hypoxia CO2 reactivity protocols despite differences in V̇E between the two protocols, and that the PET-PaO2 gradient was greater in the hypoxia protocol compared to the normoxia protocol, which was unexpected.   The correction algorithms derived are specific to the CO2 and O2 protocols employed, and possibly only our DEF system. However, these previously derived algorithms have now been validated in an independent sample (current study).  We recommend that the end-tidal-to-arterial gas gradients be considered when performing common ventilatory and vascular reactivity protocols while using DEF, as these gradients have proven that they could potentially result in data misinterpretation (i.e. under/over estimating reactivity measures).  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End-tidal-to-arterial CO2 and O2 gas gradients at low- and high-altitude during dynamic end-tidal forcing.  Title: End-tidal-to-arterial CO2 and O2 gas gradients at low- and high-altitude during dynamic end-tidal forcing.  Authors: Michael M. Tymko1                           Philip N. Ainslie1 David B. MacLeod2 Chris K. Willie1 Glen E. Foster1  Affiliations: 1Centre for Heart, Lung, and Vascular Health, School of Health and Exercise Science, University of British Columbia, Kelowna, Canada.  2Department of Anesthesiology, Duke University Medical Center, Durham, NC, USA.  Correspondence: Glen E. Foster, PhD. School of Health and Exercise Science Faculty of Health and Social Development University of British Columbia. 3333 University Way, Kelowna, BC, V1V 1V7  Telephone: 250-807-8224 Fax: 250-807-9665 Email: glen.foster@ubc.ca  Running head:  End-tidal-to-arterial gas gradients during end-tidal forcing  101  A.1 Abstract We sought to characterize and quantify the performance of a portable dynamic end-tidal forcing (DEF) system in controlling the partial pressure of arterial CO2 (PaCO2) and O2 (PaO2) at low- (LA; 344m) and high-altitude (HA; 5050m) during an isooxic CO2 test, and an isocapnic O2 test, commonly used to measure ventilatory and vascular reactivity in humans (n=9).  The isooxic CO2 tests involved step changes in the partial pressure of end-tidal carbon dioxide (PETCO2) of -10, -5, 0, +5 and +10 mmHg from baseline.  The isocapnic O2 test consisted of a 10-min hypoxic step (PETO2=47mmHg) from baseline at LA, and a 5-min euoxic step (PETO2=100mmHg) from baseline at HA.  At both altitudes, PETO2 and PETCO2 were controlled within narrow limits (<1mmHg from target) during each protocol.  During the isooxic CO2 test at LA, PETCO2 consistently overestimated PaCO2 (P<0.01) at both baseline (2.10.5mmHg) and hypercapnia (+5mmHg: 2.10.7mmHg; +10mmHg: 1.90.5mmHg). This Pa-PETCO2 gradient was approximately two-fold greater at HA (P<0.05).  At baseline at both altitudes, PETO2 overestimated PaO2 by a similar extent (LA: 6.92.1mmHg; HA: 4.50.9mmHg; both P<0.001). This overestimation persisted during isocapnic hypoxia at LA (6.90.6mmHg), and during isocapnic euoxia at HA (3.81.2mmHg). Step-wise multiple regression analysis, on the basis of the collected data, revealed that it may be possible to predict an individual’s arterial blood gases during DEF.  Future research is needed to validate these prediction algorithms, and determining the implications of end-tidal-to-arterial gradients in the assessment of ventilatory and/or vascular reactivity.   Key words:  End-tidal forcing, high-altitude, gas exchange  102  A.2 Introduction The partial pressure of end-tidal gas, measured at the relatively flat portion of the expiratory phase, is often used as a surrogate for alveolar gas and considered a reasonable index for the partial pressure of arterial blood.  Dynamic end-tidal forcing (DEF) provides a method of altering arterial blood gases by means of controlling end-tidal gases – an approach that has been used for investigating the control of breathing (10, 19) and cerebral (30, 47), pulmonary (46), and cardiac (5) vascular function.  The DEF approach uses predictive feed-forward algorithms, feedback information, and error reduction algorithms to control the end-tidal partial pressure of carbon dioxide (PETCO2) and the end-tidal partial pressure of oxygen (PETO2) by adjusting the necessary fraction of inspired CO2 (FICO2) and O2 (FIO2) on a breath-by-breath basis.   Several different DEF systems have been developed over the past few decades, each with their own strengths and weaknesses (21, 36, 39, 42).  Most recently, a compact portable DEF system by Koehle et al. (21) has been developed.  This system is portable and involves filling medical grade CO2, N2 and “air” into a reservoir bag by the time-dependent control of three gas solenoid valves (one for each mixing component).  Similar to this DEF system (21), we recently developed a DEF system capable of controlling end-tidal gases while being portable.  This system uses a mixture of three gases: O2, CO2, and N2 to deliver the desired inspiratory gas volume into a 6L capacity reservoir bag.  This DEF system has effectively controlled end-tidal gases independent of breathing frequency during isocapnic and poikilocapnic hypoxia (14, 32), and during a hyperthermic intervention to correct marked hypocapnia  (1).   A common pitfall with DEF systems, first recognized by Swanson & Bellville (42), is the assumption that end-tidal gas partial pressures accurately represent arterial blood gas pressures in all conditions (arterial blood gases being the stimulus for changes in vascular tone and chemoreflex activity).  However, even in otherwise healthy individuals, there is a gradient between end-tidal and arterial blood gases likely due to a combination of deadspace mixing (alveolar and anatomical), ventilation-perfusion (V̇/Q̇) mismatching, diffusion  103  limitation, intrapulmonary and intracardiac shunting (11, 24, 41). The end-tidal-to-arterial PCO2 gradient (Pa-PETCO2) is usually positive, but can be altered with body position (4) and age (27).  Moreover, in some cases this gradient can be negative (i.e. PETCO2> PaCO2) with exercise (18), and low breathing frequencies (17).  Caution has been advised by Robbins et al. (35) when interpreting PETCO2 during hypercapnic conditions with DEF, as administered CO2 occupies physiological deadspace during inspiration and mixes with alveolar gas during expiration, inflating PETCO2 to a greater extent than PaCO2 (17, 35).  The same effect can be observed with increases in apparatus deadspace (12).  There is a vast amount of literature reporting CO2 gradients clinically, but there is less for experimental chemosensitivity testing (6, 47, 50) and predictive correction equations for the end-tidal-to-arterial gradient have been previously derived in order to accurately predict PaCO2 during exercise and with fixed fraction inspirate but not DEF (18, 29).  In contrast, there are few reports on the end-tidal-to-arterial PO2 gradient (PET-PaO2; calculated as PETO2-PaO2) in the context of chemoreflex or vascular (e.g., cardiac, pulmonary, cerebral, etc.) testing during DEF.  It is well established that the alveolar-to-arterial difference is greater for O2 compared to CO2.  This means that end-tidal gas may not reflect PaO2 accurately, in contrast to PaCO2 (40).  Chemosensitivity and vascular function tests often manipulate O2 to low levels (hypoxia), and could be potentially harmful in individuals with abnormally large PET-PaO2 gradients.   Due to the compact nature of our DEF system, it was used on a high-altitude (HA) research expedition to 5050m (barometric pressure ~413 mmHg).  However, it is unknown how well end-tidal gases reflect arterial blood gases at extreme HA during DEF, which is extremely important due to recent innovations in DEF systems making them more portable for HA physiology research. The strong humoral response of hypoxia elicits several physiological adaptations including heterogeneous hypoxic pulmonary vasoconstriction (HPV) (46), resulting in an increase in V̇/Q̇ ratio in some areas of the lung, which has been previously shown to increase physiological deadspace (22).  In addition, because of HA-associated increases in chemosensitivity, a higher FICO2 and lower FIO2 would be required to maintain PETCO2 and PETO2 during DEF (9, 37).  The implications of a heightened ventilatory  104  sensitivity means that higher CO2 and O2 concentrations will occupy physiological deadspace at HA, potentially altering the end-tidal-to-arterial gas gradients.   The purpose of the present study was to quantify the end-tidal-to-arterial gas gradient for both CO2 and O2 at LA and HA using a portable DEF system during (1) an isooxic CO2 protocol, and (2) an isocapnic O2 protocol. To our knowledge, end-tidal gas control systems have not been used above 4300m, and end-tidal-to-arterial gas gradients have not been directly measured during DEF at or above 5050m (28).  We hypothesized that PETCO2 would overestimate PaCO2 to a greater extent during a CO2 protocol at HA, while PETO2 would overestimate PaO2 at a lesser extent at HA due to a reduction in the arterial-venous gas gradient.  105  A.3 Materials and methods Ethical approval.  All experimental procedures and protocols were approved by the clinical research ethics board at the University of British Columbia and the Nepal Health Medical Research Council, and conformed to the Declaration of Helsinki.  All participants provided written informed consent prior to participation in this study.  This study was part of a larger research expedition conducted in April-June in 2012 (14, 23).  As such, participants took part in a number of studies conducted in Kelowna, BC and during three weeks at the Ev-K2 CNR Pyramid Laboratory located near Mt. Everest basecamp at 5050m.    Participants.  To assess pulmonary and cardiovascular health, altitude naïve participants (n=12) were initially screened at LA (elevation = 344m, Kelowna, BC) for normal pulmonary function (FEV1/FVC >0.75), normal resting blood pressure (systolic blood pressure <140 and diastolic blood pressure <90 mmHg), and were required to fill out a medical history questionnaire.  Three of these subjects were unable to participate in the HA protocols due to time constraints and one case of appendicitis; these subjects were excluded from our mean data analysis. Participants (n=9; all male) included in mean data analysis were non-smokers, had no previous history of cardiovascular, cerebrovascular or respiratory diseases, were not taking any medications during testing, and had a mean  SEM age of 32.0  1.9 years, height of 180.0  1.1 cm, weight of 81.9  3.7 kg, and body mass index of 25.3  1.6 kg/m2.  All participants took acetazolamide (125 mg, 3xday) during the trek to HA to minimize the risk of acute mountain sickness but medication was discontinued 24-hours before ascending from Pheriche (4300m) to the Pyramid lab, which was at least 5 days prior to experimentation. The 8-10 day ascent profile to 5050m has been described in detail elsewhere (14, 23, 48).   Respiratory Measurements.  All respiratory parameters were acquired at 200 Hz using an analog-to-digital converter (Powerlab/16SP ML 880; ADInstruments, Colorado Springs, CO, USA) interfaced with a personal computer.  Commercially available software was used to analyze ventilatory and cardiovascular variables (LabChart V7.1, ADInstruments, Colorado Springs, CO, USA).  Throughout all procedures, subjects breathed through a mouthpiece  106  (with nose clip), a bacteriological filter and a two-way non-rebreathing valve (7900 series, Hans Rudolph, Shawnee, KS, USA).  The resistance to airflow for this breathing apparatus is 0.80 and 0.73 cmH2O/l/s at flow rates of 1.5 and 3.0 l/s, respectively.  Respired gas pressures were sampled at the mouth, dried with nafion tubing, and analyzed for PETO2 and PETCO2 (ML206; ADinstruments, Colorado Springs, CO, USA).  Gas analyzers were calibrated before each experiment with gases of known concentration using the same sample line used in the experiment.  Measured PO2 and PCO2 were time-corrected for gas analyzer sample delay and the values corresponding to the end of expiration (i.e. when respiratory flow crossed zero in the positive to negative direction) were identified as the PETO2 and PETCO2.  Respiratory flow was measured near the mouth using a pneumotachograph (HR 800L, HansRudolph, Shawnee, KS, USA) and a differential pressure amplifier (ML141, ADinstruments, Colorado Springs, CO, USA), which was calibrated with a 3 l syringe at a variety of expected flow rates.  Total apparatus deadspace was measured at 250 ml.   End-Tidal Forcing. The PETO2 and PETCO2 were controlled by a portable DEF system.  This system uses independent gas solenoid valves for O2, CO2, and N2 and controls the volume of each gas being delivered to the inspiratory reservoir through a mixing and humidification chamber.  PETO2, PETCO2, tidal volume (VT), breathing frequency (FB), and minute ventilation (V̇E) was determined for each breath online using specifically designed software (Labview 13.0, National Instruments, Austin, TX, USA).  Using feedback information regarding PETO2, PETCO2, inspired VT, and expired VT the DEF system adjusts the inspirate to bring end-tidal gas to the desired target values.  Feed-forward control of the inspirate is based on estimates of baseline metabolic O2 consumption and CO2 production and employs the alveolar gas equation to determine the required FICO2 and FIO2.  While feedback control is accomplished using a proportional and integral error reduction control system.  This system has been used previously to control end-tidal gases during physiological stressors (1, 14, 32).  End-tidal steady-state was determined once values were within one mmHg of the desired target point for at least three consecutive breaths.  107   Arterial Blood Sampling.  After local anaesthesia (2% lidocaine), a 20-gauge catheter (Radial artery catheter, Arrow International, Reading, PA, USA) was placed transcutaneously into the radial artery using ultrasound guidance and a modified Seldinger technique.  The catheter was connected to a commercially available arterial blood sampling kit (VAMP Adult, Edwards Lifescience, Irvine, CA, USA), allowing for repeated sampling and flushing with 0.9% saline.  Before sampling, the deadspace volume was withdrawn and an arterial sample (3mL) was collected in pre-heparanized syringes (safePICO syringes, Radiometer, Copenhagen, Denmark).  Air bubbles were immediately evacuated from the syringe and blood gas analysis was performed within 30-seconds of sampling with a gas analyser (ABL90 FLEX, Radiometer, Copenhagen, Denmark).  The blood gas analyzer was calibrated every 8 hours using manufacturer’s standard internal quality checks and external ampoule-based quality checks were routinely performed to confirm internal calibrations.  Reported variables that were calibrated and analyzed included: PaO2, PaCO2, and arterial oxyhaemoglobin (SaO2).  Following cannulation, all subjects rested quietly in a supine position breathing room air for at least 30-minutes to ensure normal baseline measurements prior to beginning the experimental protocol.   Experimental Design This study was conducted in two parts: LA and HA investigations.  Prior to each experiment, all participants abstained from exercise and alcohol for 24 hrs, and caffeine for 12 hrs.  All testing was conducted while participants lay in supine position. Baseline measurements are made during the last two minutes of a five minute period of eupnea breathing room-air.  Low-Altitude Protocols (344m).  LA studies involved both CO2-LA and O2-LA protocols.  A minimum of 15-minutes separated each protocol, and protocols were not randomized (CO2 protocol completed first) to avoid any confounds involving carry-over effects of sympathetic nervous activity associated with exposure to acute hypoxia (49).    108  Protocol CO2-LA.  This protocol was selected because it can be used to assess the ventilatory and cerebral vascular response to CO2 (47).  PETO2 was controlled immediately after baseline at 100 mmHg for the duration of the protocol.  Following 5-minutes of baseline, an arterial blood gas sample was collected and then PETCO2 was altered and controlled in a stepwise fashion at -10, -5, 0, +5, +10, and +15 mmHg from individual baseline values.  Targeted PETCO2 in the hypocapnic range was achieved through active hyperventilation and controlled by the DEF system.  Each step change in PETCO2 lasted for approximately 3-minutes after steady-state was reached and an arterial blood sample was carefully collected during the final 30-seconds of each stage.  Protocol O2-LA.  This protocol was selected because it is often used to determine both the ventilatory and vascular responses to isocapnic hypoxia (46, 47).  In this protocol, PETCO2 was controlled at +1 mmHg above baseline values for the entire duration of the protocol except for baseline.  Following 5-minutes of baseline, an arterial blood sample was collected and then PETO2 was decreased to 47 mmHg.  This value (PETO2 = 47 mmHg) was selected based on previous research at the same location showing that subjects on average have a baseline PETO2 of 47 mmHg at 5050m and this facilitated a comparison to the baseline state in the HA condition (13).  After PETO2 reached its target value (time = 0-minutes), arterial blood samples were collected at 7- and 10-minutes.   High-Altitude Protocols.  HA studies involved two DEF protocols to CO2 and to relief of hypoxia (protocols CO2-HA and O2-HA respectively).  A minimum of 15-minutes separated each protocol with the CO2 protocol conducted first.  Protocol CO2-HA.  Similar to protocol CO2-LA, CO2-HA was used to investigate cerebral vascular and chemoreflex sensitivity to step changes in CO2.  PETO2 was controlled at baseline values for the entire duration of the protocol (46.8 ± 1.0 mmHg). Following a 5-minute baseline, an arterial blood sample was collected and then PETCO2 was controlled in a  109  stepwise fashion at -10, -5, 0, +5, and +10 mmHg from baseline values.  Targeted PETCO2 in the hypocapnic range was achieved through active hyperventilation and controlled by the DEF system.  Due to heightened chemosensitivity at HA, a step change of +15 mmHg PETCO2 was too uncomfortable for most participants to endure, and was therefore excluded from this protocol. Each step change in PETCO2 lasted for 3-minutes after steady-state was reached and an arterial blood sample was carefully collected during the final 30-seconds of each stage.  Protocol O2-HA. O2-HA was used to address the changes in end-tidal to arterial gradient with the relief of environmental hypoxia.  In this protocol, PETCO2 was controlled at +1 mmHg above baseline values for the entire duration of the protocol.  Following a 3-minute baseline, an arterial blood sample was collected, and then PETO2 was controlled at 100 mmHg (relative hyperoxia).  This value was selected to restore PETO2 to LA values.  After end-tidal gases were stable (time = 0-minutes), arterial blood samples were collected at 4- and 5-minutes.  The O2 protocol was shorter at HA compared to LA in order to conserve limited gas supply at the field-based laboratory.  Statistical Analysis  To ensure an adequate power (>0.80) to detect a change in Pa-PETCO2 gradient from rest to hypercapnia a sample size calculation was conducted based on the PETCO2 and PaCO2 values reported by Peebles et al. (2007).  To increase our power of detection for the comparison of LA and HA parameters and to account for the possibility of subject illness and dropout at HA the sample size was increased to n=12.  All data were analysed using SigmaStat V11.5 (Systat, Chicago, IL, USA).  For each protocol baseline measurements were averaged over two minutes immediately prior to the start of the test.    During the CO2 protocols (CO2-LA and CO2-HA), all measurements were averaged over a one minute period at the end of each CO2 stage, just prior to arterial blood sampling.  During the O2 protocols (O2-LA and O2-HA), all measurements were averaged one minute prior to minute seven and ten at LA, and at minute four and five at HA, in association with when the arterial blood samples were collected.  These two measurements, taken near the end of the  110  O2 protocol for both LA and HA, were subsequently averaged together.  Two-way (Altitude: LA vs HA X PETCO2 step: -10, -5, 0, +5, +10) repeated measures analysis of variance (RM ANOVA) was used to compare differences (P<0.05) in respiratory, cardiovascular and arterial blood measurements between LA and HA for CO2 tests.  One-way (PETCO2 step: -10, -5, 0, +5, +10, +15) RM ANOVA was used to compare differences (P<0.05) in respiratory, cardiovascular and arterial blood measurements within LA data during the CO2 test, as the additional +15 mmHg PETCO2 step at LA hindered comparisons to HA.  When significant F-ratios were detected, post-hoc comparisons within LA and HA were made using Tukey’s HSD.  Paired T-tests were used to determine 1) differences between LA and HA for all outcome measures during the CO2 tests, 2) differences between end-tidal gases and arterial blood gases for both CO2 and O2 tests within LA and HA, and 3) differences between baseline and O2 condition within LA (isocapnic hypoxia) and HA (isocapnic euoxia).  Bland-Altman plots were used to assess the agreement between end-tidal and arterial blood gases.  The 95% limits of agreement were calculated by using 1.96 x S.D. of the bias (8).  Multiple forward stepwise regression analysis was used to determine which variables could accurately predict arterial blood gases.  For each condition, regression equations were determined for both PaCO2 and PaO2 as the dependent variable.  The following independent variables were included in the regression analysis: V̇E, VT, FB, FIO2, FICO2, PETO2, PETCO2 and the baseline PET-PaO2 and Pa-PETCO2 gradient.  P<0.05 was used to determine the tolerance for including independent variables in the regression model.  All data are expressed as mean values  SEM.   111  A.4 Results CO2-LA Test.  Table 1 displays ventilatory data for the isooxic CO2 test at both LA and HA.  As expected, V̇E, VT, and FB were elevated compared to baseline throughout hypocapnia (-10 and -5 mmHg PETCO2 step) and hypercapnia (+5, +10, and +15 mmHg PETCO2 step) ranges.  FICO2 was elevated, while FIO2 was decreased from baseline in both hypo- and hyper-capnic ranges (P<0.05).  The average difference between measured PETCO2 and target PETCO2 (target PETCO2 = -10, -5, 0, +5, +10, and +15 mmHg from baseline) was 0.6  0.3 mmHg, and the difference between measured PETO2 and target PETO2 (target PETO2 = 100 mmHg) was -0.2 ± 0.1 mmHg throughout the protocol.  Table 2 displays end-tidal and arterial blood gas values at baseline and during the isooxic CO2 test at both LA and HA.  At LA, significant (P<0.01) Pa-PETCO2 gradients were present at baseline, +5 mmHg and +10 mmHg PETCO2 steps.  Multiple stepwise regression analysis determined that PETCO2 (P<0.001), and the Pa-PETCO2 gradient at baseline (P=0.08) were the best predictors of PaCO2 during DEF at LA.  Linear regression analysis identified the following equations for accurately predicting PaCO2:  (1) PaCO2 = 0.363 + (0.958 * PETCO2) R2 = 0.95; p<0.001  (2) PaCO2 = 0.964 + (0.960 * PETCO2) + (0.331 * Baseline Pa-PETCO2)  R2 = 0.95; p<0.001  We found that there was a PET-PaO2 gradient at LA baseline (P<0.001), and +15 mmHg (P=0.02) PETCO2 step (see Table 2).  The PET-PaO2 gradient was reduced during DEF (i.e. during the PETCO2 steps) compared to the uncontrolled baseline (Table 2; P<0.001).  Based upon these findings it was deemed unnecessary to correct for the PET-PaO2 gradient during the CO2 test at LA.    112   CO2-HA Test.  As illustrated in table 1, V̇E and FB were significantly elevated compared to baseline throughout both hypocapnic (-10 and -5 mmHg PETCO2 steps) and hypercapnic (+5 and +10 mmHg PETCO2 steps) ranges.  VT remained unchanged during the hypocapnic range but was significantly increased during hypercapnia.  FICO2 was elevated from baseline at each PETCO2 step with the exception of -10 mmHg, while FIO2 was decreased from baseline in both hypo- and hyper-capnic ranges (P<0.05).  The average difference between measured PETCO2 and target PETCO2 (PETCO2 = -10, -5, 0, +5, and +10 mmHg from baseline) was 0.40.2 mmHg, and the difference between measured PETO2 and target PETO2 (target PETO2 = 46.8 mmHg) was 0.1±0.3 mmHg throughout the protocol.  Displayed in table 2, we found that there was a Pa-PETCO2 gradient (P<0.01) not only at baseline, but also during each stage of hypocapnia and hypercapnia.  In addition, throughout the CO2 protocol the Pa-PETCO2 gradient was significantly greater (i.e. more negative) during the hypercapnic steps compared to the hypocapnic steps (P<0.05).  Multiple stepwise regression analysis determined that PETCO2 (P<0.001), and the baseline Pa-PETCO2 gradient (P<0.001) were the best predictors of PaCO2 when using DEF at HA.  Using linear regression analysis including these variables the following equations were derived which accurately predict PaCO2:  (3) PaCO2 = 0.143 + (0.861 * PETCO2)     R2 = 0.92; p<0.001  (4) PaCO2 = 2.899 + (0.861 * PETCO2) + (0.675 * Baseline Pa-PETCO2)  R2 = 0.97; p<0.001  We also found that there was a PET-PaO2 gradient present during baseline (P<0.001) and all stages of hypocapnia (P<0.01) and hypercapnia (P<0.001), as seen in table 2.  This gradient remained unchanged from baseline throughout the CO2 protocol meaning that the PET-PaO2  113  gradient can be corrected for during the CO2 protocol using the individual’s baseline PET-PaO2 gradient (P<0.001).  Comparison of CO2-LA and CO2-HA Tests.  Differences between HA and LA for respiratory and arterial blood gases are reported in tables 1 and 2.  Breathing frequency was significantly elevated at HA compared to LA during baseline, hypocapnia, and hypercapnia.  V̇E and VT were unchanged at baseline and during active hyperventilation between LA and HA, but were higher at HA during hypercapnia.  The FICO2 was significantly greater at HA compared to LA at -10 mmHg, +5 mmHg, and +10 mmHg (P<0.05).  The FIO2 was unchanged between LA and HA in the hypocapnic range, but was lower during HA in the hypercapnic range (P<0.05).  The Pa-PETCO2 gradient (P<0.05) was significantly greater (i.e. more negative) at HA compared to LA at baseline, +5, and +10 PETCO2 steps (see table 2, and figures 1 and 2).  As mentioned, there was no Pa-PETCO2 gradient found during hypocapnia at LA, nor did we achieve a +15 mmHg PETCO2 step at HA; thus, these comparisons were not made.    The PET-PaO2 gradient (P<0.05) was significantly greater (i.e. more positive) at HA compared to LA during DEF at -10, -5, +5, and +10 mmHg PETCO2 steps (see table 2).  O2-LA Test.  Table 3 displays ventilatory and arterial blood data during the isocapnic O2 test at both LA and HA.  V̇E, VT, and FB significantly increased from baseline during isocapnic hypoxia.  We found that there was a PET-PaO2 gradient at baseline (6.92.1 mmHg, P<0.001) and during isocapnic hypoxia (6.90.6 mmHg, P<0.001).  The average difference between measured PETO2 and target PETO2 (target PETO2 = 47 mmHg) was 0.10.2 mmHg, and the difference between measured PETCO2 and target PETCO2 (target PETCO2 = +1 mmHg from baseline) was -0.40.3 mmHg throughout the protocol.  The FIO2 was lower during isocapnic hypoxia compared to baseline, while the FICO2 was higher during isocapnic hypoxia  114  compared to baseline (P<0.05).  There was no difference of the PET-PaO2 gradient between baseline and isocapnic hypoxia (see table 3).  Multiple stepwise regression analysis identified PETO2 (P<0.001), and the baseline PETO2-PaO2 gradient (P<0.001) as suitable predictors of PaO2 during DEF at LA.  Linear regression analysis identified the following equations for accurately predicting PaO2:  (5) PaO2 = -6.024 + (0.986 * PETO2)   R2 = 0.94; p < 0.001  (6) PaO2 = -2.520 + (0.995 * PETO2) - (0.592 * Baseline PET-PaO2)    R2 = 0.98; p < 0.001  Whilst there was a Pa-PETCO2 gradient during the O2 protocol at baseline (-2.0  0.4 mmHg, P<0.001), this gradient was abolished during isocapnic hypoxia (table 3).   O2-HA Test.  In table 3, V̇E and VT did not change from baseline after administration of isocapnic euoxia, but FB was elevated.  The difference between measured PETO2 and target PETO2 (target PETO2 = 100 mmHg) was -0.1  0.2 mmHg, and the difference between measured PETCO2 and target PETCO2 (target PETCO2 = 100 mmHg) was -0.5  0.3 mmHg from our target (target PETCO2 = +1 mmHg from baseline) throughout the protocol.  Also shown in table 3, we found that there was a PET-PaO2 gradient at baseline, and during isocapnic euoxia.  The FIO2 and FICO2 were higher during isocapnic euoxia compared to baseline (P<0.05).  There was no difference in the PET-PaO2 gradient between these conditions.  Multiple stepwise regression analysis identified PETO2 (P<0.001), and the baseline PET-PaO2 gradient (P<0.001) as the best predictors of PaO2 when using DEF at HA.  Linear regression analysis derived the following equations which could accurately predict PaO2:   115  (7) PaO2 = -4.826 + (1.010 * PETO2)   R2 = 0.99; p < 0.001  (8) PaO2 = -1.880 + (1.012 * PETO2) - (0.695 * Baseline PET-PaO2)   R2 = 1.0; p < 0.001  In addition, we found that there was a Pa-PETCO2 gradient at baseline and isocapnic euoxia, displayed in table 3.  This gradient remained unchanged from baseline throughout the O2 protocol meaning that PaCO2 can be accounted for based on the individual’s PET-PaCO2 gradient during the O2-HA protocol (P<0.001).    Comparison of O2-LA and O2-HA Tests.  There were no significant differences when comparing the PET-PaO2 gradient between baseline at LA to isocapnic euoxia at HA (P=0.15), and between isocapnic hypoxia at LA to baseline at HA (P=0.08), where PETO2 values were similar to one another.  It was deemed unreasonable for the O2 tests conducted at LA and HA to be directly compared to one another outside of baseline since the two tests were fundamentally different from one another and as a result we did not conduct any other statistical analysis between LA and HA.   116  A.5 Discussion  The primary purpose of this study was to quantify the end-tidal-to-arterial gradients for both O2 and CO2 at LA and, for the first time, HA during an isooxic CO2 and an isocapnic O2 protocol.  The main findings from this study are that (1) The Pa-PETCO2 gradient is greater at HA (i.e. more negative) compared to LA, especially during hypercapnia; (2) The PET-PaO2 gradient remained the same between LA and HA at baseline, and was unchanged during isocapnic hypoxia (at LA) and relative hyperoxia (at HA); and (3) our recently developed computer-controlled portable DEF system effectively targets PETO2 and PETCO2 at both LA and HA.  Based upon the results from this study, correction equations were derived using multiple stepwise regression analysis. This approach was utilized in order to improve predictions of arterial blood gases at SL and HA during DEF.   A DEF system for controlling end-tidal gases at LA and HA. Our DEF system has been previously used to control end-tidal gases in other recent studies (1, 14, 32), but its performance has not yet been formally quantified.  This DEF system is inexpensive and portable making it possible to perform studies in remote HA environments as well as at LA.  To our knowledge, this is the first study to use a computer controlled DEF system above 5000m (28).  On average this DEF system was able to control end-tidal gases within 1-mmHg from our target end-tidal values.  In addition, there were no differences in the ability of our DEF system to target PETO2 and PETCO2 between LA and HA environments (see Figures 1 and 3).   Relationship between PETCO2 and PaCO2 at LA and HA. There are a number of reported conditions where PETCO2 does not accurately reflect PaCO2, such as: exercise, aging, body position, and in patients with abnormal lung conditions such as pulmonary oedema and pneumonia (4, 18, 25, 27, 31).  In contrast to some previously reported data (7, 35), our study found that there was a negative Pa-PETCO2 gradient at baseline similar to that reported by Peebles et al. (29).  Apparatus deadspace could potentially explain  117  our negative Pa-PETCO2 gradient at baseline, but perhaps a better explanation is that we found large negative Pa-PETCO2 gradients in three of our subjects at LA shifting our mean bias enough to become statistically significant.  In addition, we found a negative Pa-PETCO2 during hypercapnia (+5, and +10 mmHg) at LA.  No gradient presented itself in the +15 mmHg PETCO2 step at LA, however the Pa-PETCO2 gradient was close to significance (P=0.071).  In contrast, at HA there was a Pa-PETCO2 gradient at baseline and during both hypocapnia and hypercapnia, and these gradients were of greater magnitude (i.e. more negative) when compared to the gradients found at LA.  The widening of the Pa-PETCO2 gradient between LA and HA could potentially be the result of a diffusion limitation from subclinical mild HA pulmonary oedema (44).  However, the participants in our study had no signs or symptoms of acute mountain sickness at the time of experimentation (experimentation took place between days 5-10 at HA).  In addition, there was no change in the PET-PaO2 gradient between LA and HA, which further supports the belief that diffusion limitation remains unchanged.  The most likely explanation for the differences in the Pa-PETCO2 gradient between LA and HA is increased alveolar deadspace (26, 44).  The Pa-PETCO2 gradient was not only greater at HA compared to LA, but it also widened progressively (i.e. became more negative) during hypercapnia at HA, this trend can be observed in figure 2.  One explanation for this phenomenon is the heightened ventilatory response at HA necessitates an increased FICO2 to maintain a given PETCO2 during DEF, increasing the fraction of CO2 trapped in physiological and apparatus deadspace and thereby inflating the measured PETCO2.  In addition to chronic hypoxia leading to an increase in alveolar deadspace, there is also evidence demonstrating that administered PCO2 can cause small airway dilation and pulmonary vasoconstriction augmenting the HPV response, potentially further increasing physiological deadspace during hypercapnia which could explain the Pa-PETCO2 gradient differences between hypocapnia and hypercapnia (2, 3, 20, 38).  A controversial mechanism suggested by Gurtner & Traystman (16) for negative gas gradients observed during steady-state hypercapnia is referred to as the ‘charged membrane hypothesis’. Here, the suggested mechanism is that the generation of an electrical negative field near the capillary endothelial surface within the lung results in  118  dissociation of weak acids and elevates local hydrogen ion concentration.  The increase in local hydrogen ion concentration shifts the equilibrium of the bicarbonate buffering relationship toward an increased PCO2 (16).  However, this hypothesis is based on 1) anesthetized dogs during an extreme level of hypercapnia that is not tolerated by healthy humans, and 2) measured alveolar-to-arterial gradients, in contrast to end-tidal-to-arterial gradients.  Therefore, we speculate that our observations are more likely due to an effect of physiological deadspace on end-tidal gas sampling.   During our isocapnic O2 test, participants endured a hypoxic step change at LA and a euoxic step change at HA, while PETCO2 was controlled at +1 mmHg above baseline values.  We discovered that there was a Pa-PETCO2 gradient at LA during our uncontrolled baseline period, but this gradient was abolished during isocapnic hypoxia.  In contrast, at HA, there was a Pa-PETCO2 gradient of the same magnitude at baseline and during isocapnic euoxia.  At baseline, PETCO2 overestimated PaCO2 at HA to a greater degree compared to LA.   Our data indicate that during an acute isocapnic hypoxia test at LA the Pa-PETCO2 gradient is negligible, but is maintained during hypoxia (eg. high-altitude, normobaric hypoxia).  Differences in PETO2 and PaO2 at LA and HA. Surprisingly, there is little literature available with respect to the PET-PaO2 gradient during DEF at LA or HA.  Although experiments investigating the alveolar-to-arterial oxygen difference (AaDO2) between LA and HA exist, it is difficult to assess the AaDO2 during DEF necessitating the study of the PET-PaO2 gradient in its place.  However, literature regarding the AaDO2 is inconsistent suggesting that the AaDO2 can widen or narrow following ascent to HA at baseline (22, 33, 45).  During DEF, inspired gas concentrations are constantly changing making it difficult to assess alveolar PO2.  The PET-PaO2 gradient differs from the AaDO2 because it includes some mixing of alveolar gas with physiological deadspace gas.   During the isocapnic hypoxia test at LA, both baseline measures and measurements taken near the end of the protocol showed that PETO2 consistently overestimated PaO2, and these gradients measured at LA were unchanged from each other.  Intuitively, the PET-PaO2  119  gradient would increase with isocapnic hypoxia due to increased alveolar deadspace from hypoxic pulmonary vasoconstriction, however, the hypoxic stimulus we administered (10-minutes) would have resulted in a modest change in pulmonary artery pressure compared to that at high altitude (15).  The isocapnic O2 protocol used at HA was fundamentally different than the one used at LA.  The HA isocapnic O2 test involved administering higher amounts of O2 from baseline to values that were similar to those seen during baseline at LA (euoxia).  For this reason we did not compare the PET-PaO2 gradient between the LA and HA outside of our baseline measures, as we technically delivered different stimuli.  The unchanged PET-PaO2 gradient seen at baseline between LA and HA in healthy subjects was somewhat unexpected as there are likely HA-related changes in alveolar deadspace volume.  However, this is likely explained by the reduction in atmospheric PO2 at HA, making the inspiratory PO2 to PaO2 difference lower, which will minimize the difference between end-tidal and arterial PO2.  The concomitant changes of alveolar deadspace and the reduction of atmospheric PO2 associated with HA may oppose each other in such a manner that the net result is an unchanged PET-PaO2 gradient observed during baseline between LA and HA.  Changes in alveolar deadspace not only occur during hypoxia (acute and chronic), but it may also occur during hyperoxia (i.e. increased FiO2) (50).  During hyperoxia, there is a redistribution of pulmonary blood from poorly perfused alveoli to highly perfused alveoli due to pulmonary vasodilation potentially further increasing physiological deadspace during hypercapnia (50).  The end result is an increase in alveolar deadspace volume which has the potential of widening the PET-PaO2 gradient.  However, we did not observe this effect at HA, as the PET-PaO2 gradient was unchanged between baseline and isocapnic euoxia (PETO2 = 100 mmHg).  It is possible that our euoxia stimulus was not strong enough or applied for too short a time period to elicit a significant response.   Can PaCO2  be predicted from PETCO2 during end-tidal forcing? Previous equations have been proposed for predicting PaCO2 by Jones et al. (18) during  120  exercise, and Peebles et al. (29) during hypocapnia (through active hyperventilation) and hypercapnia (increased fixed FICO2 inspirate). However, as highlighted in figure 5, we found that the correction algorithms proposed by Jones et al. (18) and Peebles et al. (29) could not accurately predict PaCO2 during our DEF protocols (likely due to the different experimental conditions); therefore, multiple stepwise regression analysis was used to determine which variables might predict PaCO2 at LA and HA.  Based on these variables we conducted linear regression analysis with PaCO2 as our dependent variable to better predict PaCO2 at LA and HA.  Our results suggest that a correction algorithm that includes PETCO2 as a variable will more accurately predict PaCO2.    Methodological Considerations. Acetazolamide (a carbonic anhydrase inhibitor), decreases the risk of high-altitude sickness by increasing central chemoreflex drive and improving arterial oxygenation (43).  Acetazolamide administration has been proven useful for safe acclimatization to extreme altitude, and effectively reduces the risk of pulmonary edema (thus diffusion limitation), and alveolar deadspace due to improved V̇/Q̇ matching (44, 46).  Therefore, if acetazolamide had long-lasting effects (i.e. 5-days beyond its discontinuation), it could be argued that our results have underestimated the true difference in end-tidal-to-arterial gradient between LA and HA during DEF. This possibility would seem unlikely, however, given the rapid clearance of acetazolamide (<36 hrs) (34). Despite the advantages of our DEF system being portable, cost effective (due to low gas requirements), and capable of controlling end-tidal gases within narrow limits, it, similar to other systems is affected by methodological constraints.  First, similar to all DEF systems, hypocapnic steps can only be achieved by asking participants to actively hyperventilate, driving their PETCO2 below desired targets where the DEF is then capable of controlling and fine tuning PETCO2 by adding CO2 into the breathing circuit.  Thus, with hypocapnia, end-tidal gas control is dependent upon the ability of the participant to maintain a constant rate of ventilation. Second, similar to the DEF system developed by Koehle et al. (21), our DEF system functions by mixing inspirate into a reservoir bag on a breath-by-breath basis.  In an  121  ideal world the reservoir bag would be emptied with each breath so that only the inspirate for the next breath is available to the subject at any given time.  However, in order to deal with variation in VT with each breath a small amount of extra air remains in the reservoir bag to prevent its complete emptying.  This safety volume is normally 100 ml more than the current VT but can be adjusted by the DEF operator to maintain the reservoir volume as small as possible.  In addition, the algorithm for the system estimates the volume of each gas remaining in the reservoir and takes these volumes into account before injecting the next volume of gases to better target the correct fractions of inspired O2 and CO2.  Nonetheless, inaccuracies in this prediction or inflation in the estimated reservoir volume can lead to a sluggish response or a delay in the systems response to a perturbation.  Third, our DEF system uses 100% CO2, O2, and N2 compressed gas to fill our inspirate reservoir.  Although avoiding pre-mixed gases is usually less expensive, it does present some potential safety issues as it is possible for the participant to be administered 100% N2 during a severe hypoxic step.  Therefore it is crucial to employ a well-trained DEF device operator to recognize these potential issues.  If a problem arises in which the participant’s end-tidal values are not being controlled properly, 100% O2 can be delivered to the participant for safety reasons.  Finally, similar to all DEF system, our DEF system assumes the end-tidal gases are a surrogate for arterial blood gases.    Perspectives and Significance. We hypothesized that PETCO2 would overestimate PaCO2 to a greater extent during a CO2 protocol at HA, while PETO2 would overestimate PaO2 to a lesser extent at HA due to a reduction in the arterial-venous gas gradient.  In agreement with our hypothesis, we demonstrated that during DEF PaCO2 is systematically overestimated by PETCO2 at baseline, hypocapnia and hypercapnia at HA, but only at baseline and hypercapnia at LA.  This overestimation of PaCO2 is exacerbated at HA compared to LA.  In contrast to our hypothesis, we found that PETO2 overestimates PaO2 by the same magnitude at LA and HA, and these gradients remained unchanged throughout the O2 protocols.  The correction algorithms derived are specific to our experimental conditions including our study sample, altitude, the  122  CO2 and O2 protocols employed, and possibly only our DEF system. However, these algorithms need to be validated in an independent sample during similar protocols.  We recommend that the end-tidal-to-arterial gas gradients be considered when performing common ventilatory and vascular reactivity protocols to CO2 and O2 as they could potentially effect the measured response.  With future validation, it may be possible to use the regression equations reported here to estimate arterial blood gases in the event that invasive measurements are not feasible.  123  Acknowledgements:  This study was carried out within the framework of the Ev-K2-CNR Project in collaboration with Nepal Academy of Science and Technology as foreseen in the Memorandum of Understanding between Nepal and Italy, and thanks to a contribution from the Italian National Research Council.  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Regional brain blood flow in man during acute changes in arterial blood gases. J Physiol 590: Pt 14: 3261-3275, 2012. 48. Willie CK, Smith KJ, Day TA, Ray LA, Lewis NC, Bakker A, Macleod DB and Ainslie PN. Regional cerebral blood flow in humans at high altitude: gradual ascent and 2 wk at 5,050 m. J Appl Physiol 116: 7: 905-10, 2014. 49. Xie A, Skatrud JB, Puleo DS and Morgan BJ. Exposure to hypoxia produces long-lasting sympathetic activation in humans. J Appl Physiol 91: 4: 1555-62, 2001. 50. Yamauchi H, Ito S, Sasano H, Azami T, Fisher J and Sobue K. 1. Bain AR, Smith KJ, Lewis NC, Foster GE, Wildfong KW, Willie CK, Hartley GL, Cheung SS and Ainslie PN. Regional changes in brain blood flow during severe passive hyperthermia: effects of PaCO2 and extracranial blood flow. J Appl Physiol 115: 5: 653-9, 2013.   130  Table A.1: Ventilatory data during an isooxic CO2 test at LA and HA.   Baseline -10 mmHg  -5 mmHg 0 mmHg +5 mmHg +10 mmHg         +15 mmHg V̇E (l min-1) LA 14.3 ± 0.7 39.0 ± 5.0* 34.2 ± 4.8* 19.7 ± 2.2 21.5 ± 1.8* 44.5 ± 5.4* 57.6 ± 5.7*  HA 19.9 ± 1.0† 46.8 ± 3.5* 40.5 ± 4.4* 28.6 ± 2.6† 53.3 ± 7.2*† 84.2 ± 5.9*†   PETCO2: <0.001 ; Altitude: <0.001 ; Interaction: <0.001 VT (l) LA 1.0 ± 0.1 1.7 ± 0.3* 1.5 ± 0.3* 1.1 ± 0.2 1.4 ± 0.1* 2.2 ± 0.1* 2.5 ± 0.2*  HA 1.2 ± 0.1† 1.6 ± 0.2 1.4 ± 0.2 1.2 ± 0.1 2.1 ± 0.1*† 2.6 ± 0.1*†   PETCO2: <0.001 ; Altitude: 0.210 ; Interaction: <0.001 FB  (breaths min-1) LA 15.1 ± 0.8 24.5 ± 2.3* 24.2 ± 3.0* 19.7 ± 2.7 15.2 ± 1.1 20.3 ± 2.7 24.1 ± 3.1*  HA 17.2 ± 1.0 31.5 ± 2.4*† 30.1 ± 2.8* 24.6 ± 2.4* 24.9 ± 2.5*† 32.5 ± 2.7*†   PETCO2: <0.001 ; Altitude: 0.004 ; Interaction: 0.013 FICO2 (%) LA 0.2 ± 0.0 1.9 ± 0.3* 3.4 ± 0.2* 2.9 ± 0.4* 5.0 ± 0.3* 6.7 ± 0.1* 7.4 ± 0.1*  HA 0.2 ± 0.1 0.2 ± 0.3† 3.2 ± 0.4* 4.0 ± 0.5* 7.3 ± 0.3*† 9.1 ± 0.1*†  PETCO2: <0.001 ; Altitude: 0.028 ; Interaction: <0.001 FIO2 (%) LA 20.8 ± 0.1 17.1 ± 0.2* 17.4 ± 0.2* 19.7 ± 0.6 18.2 ± 0.3* 16.6 ± 0.1* 16.3 ± 0.1*  HA 21.0 ± 0.2 16.1 ± 0.2*† 16.6 ± 0.4* 18.4 ± 0.7* 15.8 ± 0.5*† 14.7 ± 0.2*†  PETCO2: <0.001 ; Altitude: 0.005 ; Interaction: 0.012 V̇E, minute ventilation; VT, tidal volume; FB, breathing frequency; FICO2, fraction of inspired carbon dioxide; FIO2, fraction of inspired oxygen. Reported p-values represent differences between LA and HA.  *P<0.05, vs baseline. †P<0.05 low-altitude vs. high-altitude. Values are mean  SEM.  The P-values for the main effects of PETCO2 step and altitude, and the P-value of the interaction effect are displayed underneath each measured variable.  131  Table A.2 End-tidal and arterial blood gases during an isooxic CO2 test at LA and HA.   Baseline -10 mmHg -5 mmHg 0 mmHg +5 mmHg +10 mmHg +15 mmHg PETCO2  (mmHg) LA 41.5 ± 0.4 31.9 ± 0.4* 37.0 ± 0.4* 42.4 ± 0.4 46.9 ± 0.6* 52.3 ± 0.4* 57.2 ± 0.4*  HA 28.7 ± 0.5† 19.0 ± 0.5*† 24.1 ± 0.5*† 29.1 ± 0.5† 34.2 ± 0.5*† 39.3 ± 0.5*†   PETCO2: <0.001; Altitude: <0.001; Interaction: 0.469 PaCO2 (mmHg) LA 39.4 ± 0.5 30.9 ± 0.6* 36.2 ± 0.5* 40.5 ± 0.8 44.9 ± 0.5* 50.4 ± 0.5* 55.9 ± 1.0*  HA 24.6 ± 0.9† 16.2 ± 0.7*† 21.2 ± 0.9*† 25.5 ± 0.7† 29.1 ± 0.7*† 34.1 ± 0.8*†   PETCO2: <0.001; Altitude: <0.001; Interaction: 0.310 Pa-PETCO2 (mmHg)  LA -2.1 ± 0.5‡ -1.1 ± 0.6 -0.7 ± 0.4 -1.8 ± 0.7‡ -2.1 ± 0.7‡ -1.9 ± 0.5‡ -1.3 ± 0.9  HA -4.1 ± 0.7‡† -2.7 ± 0.4‡ -2.9 ± 0.7‡ -3.6 ± 0.7‡† -5.1 ± 0.7‡† -5.2 ± 0.7‡†  PETCO2: <0.001; Altitude: <0.001; Interaction: 0.349 PETO2 (mmHg)  LA 93.2 ± 1.1 100.0 ± 0.2* 100.1 ± 0.1* 98.7 ± 0.9* 99.9 ± 1.0* 100.0 ± 0.2* 100.0 ± 0.2*  HA 46.8 ± 1.0† 46.6 ± 0.5† 46.7 ± 0.5† 46.6 ± 0.4† 46.5 ± 0.5† 46.8 ±0.5†   PETCO2: <0.001; Altitude: <0.001; Interaction: <0.001 PaO2 (mmHg)  LA 88.1 ± 2.2 100.2 ± 1.6* 100.1 ± 0.9* 95.1 ± 2.1* 98.9 ± 1.6* 101.9 ± 0.6* 102.6 ± 0.8*  HA 41.7 ± 1.2† 42.8 ± 1.0† 41.5 ± 0.9† 41.2 ± 1.0† 41.3 ± 0.6† 41.8 ± 0.6†   PETCO2: <0.001; Altitude: <0.001; Interaction: <0.001 PET-PaO2(mmHg) LA 5.1 ± 1.4‡ -0.2 ± 1.5 0.0 ± 0.9 3.5 ± 1.5‡ 1.0 ± 0.8 -1.9 ± 0.6 -2.7 ± 0.9‡  HA 5.1 ± 1.0‡ 3.8 ± 1.0‡ 5.1 ± 1.0‡ 5.4 ± 1.0‡ 5.2 ± 0.8‡ 5.0 ± 0.9‡  PETCO2: <0.001; Altitude: 0.016; Interaction: 0.003 PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial gas gradient of carbon dioxide; PETO2, end-tidal partial pressure of oxygen; PaO2, arterial partial pressure oxygen; PET-PaO2, end-tidal-to-arterial gas gradient of oxygen.  Reported p-values represent differences between LA and HA (statistical significance set at P<0.05). *P<0.05, vs baseline. †P<0.05 low-altitude vs high-altitude.  ‡P<0.05, end-tidal vs arterial gases (i.e significant gradient).  P<0.05, Pa-PETCO2 hypercapnia vs hypocapnia (-10 and -5 mmHg) at HA. Values are mean  SEM. The P-values for the main effects of PETCO2 step and altitude, and the P-value of the interaction effect are displayed in the last three columns. 132  Table A.3 Ventilatory and arterial blood gas data during an isocapnic O2 test at LA and HA.  Altitude Baseline Isocapnic Hypoxia Altitude Baseline Isocapnic Euoxia PETCO2 mmHg  LA 40.6 ± 0.5 41.2 ± 0.4 HA 28.6 ± 0.6 29.1 ± 0.6 PaCO2 mmHg LA 38.6 ± 0.7 41.4 ± 0.7* HA 24.8 ± 0.8 26.0 ± 0.9 Pa-PETCO2 mmHg LA -2.0 ± 0.4‡ 0.1 ± 0.5 HA -3.7 ± 0.7‡ -3.1 ± 0.6‡ PETO2 mmHg LA 82.9 ± 1.8 46.9 ± 0.2* HA 45.7 ± 0.7 100.1 ± 0.4* PaO2 mmHg LA 76.0 ± 2.4 40.0 ± 0.5* HA 41.2 ± 0.5 96.3 ± 1.3* PET-PaO2 mmHg LA 6.9 ± 2.1‡ 6.9 ± 0.6‡ HA 4.5 ± 0.9‡ 3.8 ± 1.2‡ V̇E l min-1 LA 12.6 ± 0.7 37.3 ± 4.4* HA 19.6 ± 1.5 25.3 ± 2.9* VT l LA 0.9 ± 0.0 1.8 ± 0.1* HA 1.3 ± 0.1 1.2 ± 0.1 FB breaths min-1 LA 14.9 ± 0.8 20.3 ± 2.2* HA 16.3 ± 1.5 22.4 ± 2.7* FIO2 % LA 20.7 ± 0.1 9.2 ± 0.2* HA 20.9 ± 0.2 34.2 ± 0.7* FICO2 % LA 0.3 ± 0.0 4.5 ± 0.2* HA 0.2 ± 0.1 2.4 ± 0.6* PETCO2, end-tidal partial pressure of carbon dioxide; PaCO2, arterial partial pressure of carbon dioxide; Pa-PETCO2, end-tidal-to-arterial gas gradient of carbon dioxide; PETO2, end-tidal partial pressure of oxygen;  PaO2, arterial partial pressure oxygen; PET-PaO2, end-tidal-to-arterial gas gradient of oxygen; V̇E, minute ventilation; VT, tidal volume; FB, breathing frequency; FIO2, fraction of inspired oxygen; FICO2, fraction of inspired carbon dioxide. *P<0.05, vs baseline. ‡P<0.05, end-tidal vs arterial gases (i.e significant gradient). Values are mean  SEM.  133  Figure Legends. Figure 1. Representative tracing of isooxic CO2 test at LA and HA.  Open circles () represent PETO2, end-tidal partial pressure of oxygen; closed circles () represent PETCO2, end-tidal partial pressure of carbon dioxide.  Top panels A (LA) and B (HA), represents mean data (15-second time bins)  SEM from the isooxic CO2 test in 9 participants.  The numeric values below the PETCO2 data represents the mean end-tidal-to-arterial gas gradient for carbon dioxide.  Bottom panels C (LA) and D (HA), represents raw breath-by-breath data from one participant from the isooxic CO2 test.   Figure 2. Bland and Altman plot of differences between PaCO2 and PETCO2 during an isooxic CO2 test at LA and HA.  (), -10 mmHg; (), -5 mmHg; (▼), 0 mmHg; (∆), +5 mmHg; (■), +10 mmHg; (□), +15 mmHg.  Dotted lines represent the 95% confidence intervals and the continuous lines represents the mean bias.  Panel A., representing the end-tidal-to-arterial difference at LA, and Panel B, represents the end-tidal-to-arterial difference at HA during the CO2 test.   Figure 3. Representative tracing of the isocapnic O2 test at LA and HA.  Open circles () represent PETO2, end-tidal partial pressure of oxygen; closed circles () represent PETCO2, end-tidal partial pressure of carbon dioxide.  Top panels A (LA) and B (HA), represents mean data (15-second time bins)  SEM from the isocapnic O2 test in 9 participants.  The numeric values above PETO2 data represents the end-tidal-to-arterial gas gradient for oxygen.  Bottom panels C (LA) and D (HA), represents raw breath-by-breath data from one participant during the isocapnic O2 test.    Figure 4. Bland and Altman plot of differences between PaO2 and PETO2 partial pressures of O2 during an isocapnic O2 test at LA and HA.  Dotted lines represent the 95% confidence intervals and the continuous lines represent the mean bias.  Panel A., representing the end-tidal-to-arterial difference at LA, and Panel B., represents the end-tidal-to-arterial difference at HA.   134   Figure 5. Assessment of PaCO2 and PETCO2 relationship of CO2 during an isooxic CO2 test at LA and HA.  Data points were obtained during the last minute of each step change in PETCO2.  Dotted lines represent 95% confidence intervals.  Panel A represents pooled linear regression for PaCO2 and PETCO2.  Panel B represents pooled linear regression for PaCO2 and PETCO2 from the current study data that has been adjusted using an algorithm proposed by Jones et al. (1979) (ePaCO2 = 5.5 + 0.9* PETCO2– VT).  Panel C represents pooled linear regression for PaCO2and PETCO2from the current study data that has been adjusted with an algorithm proposed by Peebles et al. in 2007 (ePaCO2 = 2.367 + 0.884* PETCO2). b[1], slope; b[0], y-intercept.   135  Figure A.1: Representative tracing of isooxic CO2 test at LA and HA  PETCO2 and PETO2 (mmHg)020406080100120020406080100120Baseline -10 mmHg -5 mmHg 0 mmHg+5 mmHg +10 mmHg +15 mmHg Baseline -10 mmHg -5 mmHg 0 mmHg +5 mmHg +10 mmHg-2.1-1.1-0.7-1.8-2.1-1.9-1.3-4.1-2.7-2.9-3.6-5.1-5.22 min2 minTime (s)0 200 400 600 800 1000 1200PETCO2 and PETO2 (mmHg)020406080100120Time (s)0 200 400 600 800 1000020406080100120A.D.C.B. 136  Figure A.2: Bland and Altman plot of differences between PaCO2 and PETCO2 during an isooxic CO2 test at LA and HA  PaCO2 and PETCO2 mean (mmHg)25 30 35 40 45 50 55 60 65PaCO2 - PETCO2 (mmHg)-10-8-6-4-20246PaCO2 and PETCO2 mean (mmHg)10 15 20 25 30 35 40-10-8-6-4-20246A.B. 137  Figure A.3: Representative tracing of the isocapnic O2 test at LA and HA       Time (s)0 100 200 300 400 500 600 700PETCO2 and PETO2 (mmHg)020406080100120Time (s)0 100 200 300 400020406080100120A. B.PETCO2 and PETO2 (mmHg)0204060801001200204060801001206.96.9 4.53.8BaselineBaselineIsocapnic Hypoxia Isocapnic EuoxiaC. D. 138   Figure A.4: Bland and Altman plot of differences between PaO2 and PETO2 partial pressures of O2 during an isocapnic O2 test at LA and HA            PaO2 and PETO2 mean (mmHg)41 42 43 44 45PETO2 - PaO2 (mmHg)-2024681012PaO2 and PETO2 mean (mmHg)94 95 96 97 98 99 100 101-2024681012A. B.  139  Figure A.5: Assessment of PaCO2 and PETCO2 relationship of CO2 during an isooxic CO2 test at LA and HA       20 30 40 50 60PETCO2 (mmHg) 203040506070R2 = 0.92PaCO2 (mmHg)10 15 20 25 30 35 40PETCO2 (mmHg) 1020304050R2 = 0.9220 30 40 50 60ePeeblesPaCO2 (mmHg)20304050607020 30 40 50 60eJonesPaCO2 (mmHg)203040506070PaCO2 (mmHg)10 15 20 25 30 35 40eJonesPaCO2 (mmHg)1020304050R2 = 0.91R2 = 0.87PaCO2 (mmHg)10 15 20 25 30 35 40ePeeblesPaCO2 (mmHg)1020304050R2 = 0.92R2 = 0.92b[1] = 0.80b[1] = 0.97b[1] = 1.07b[1] = 0.88b[1] = 0.95b[1] = 0.86A.B. C.b[0] = 3.09 b[0] = 7.96b[0] = 5.10b[0] = 2.13 b[0] = 5.89b[0] = 4.26HASL 140  Appendix B: University of British Columbia ethics certificate of full board approval      The University of British Columbia Office of Research (Services) Ethics Clinical Research Ethics Board – Room 210, 828 West 10th Avenue, Vancouver, BC V5Z 1L8     ETHICS CERTIFICATE OF FULL BOARD APPROVAL     PRINCIPAL INVESTIGATOR: INSTITUTION / DEPARTMENT: UBC CREB NUMBER: Glen E. Foster  UBC/UBCO Health & Social Development/UBCO Health and Exercise Sciences  H14-01128 INSTITUTION(S) WHERE RESEARCH WILL BE CARRIED OUT:  Institution Site UBC Okanagan Other locations where the research will be conducted: N/A   CO-INVESTIGATOR(S): Michael Tymko Joshua C. Tremblay   SPONSORING AGENCIES: - British Columbia Knowledge Development Fund (BCKDF) - "Cardiopulmonary Laboratory for Experimental and Applied Physiology" - Canada Foundation for Innovation - "Cardiopulmonary Laboratory for Experimental and Applied Physiology" - Natural Sciences and Engineering Research Council of Canada (NSERC) - "Intermittent Hypoxia and Cardiopulmonary Adaptation " - UBCO Faculty of Health and Social Development - "Start-Up Funds"  PROJECT TITLE: Validation of end-tidal-to-arterial correction algorithms during a CO2 reactivity test in normoxia and hypoxia.  THE CURRENT UBC CREB APPROVAL FOR THIS STUDY EXPIRES:  June 10, 2015 The full UBC Clinical Research Ethics Board has reviewed the above described research project, including associated documentation noted below, and finds the research project acceptable on ethical grounds for research involving human subjects and hereby grants approval.  This approval applies to research ethics issues only. The approval does not obligate an institution or any of its departments to proceed with activation of the study. The Principal Investigator for the study is responsible for identifying and ensuring that resource impacts from this study on any institution are properly negotiated, and that other institutional policies are followed. The REB assumes that investigators and the coordinating office of all trials continuously review new information for findings that indicate a change should be made to the protocol, consent documents or conduct of the trial and that such changes will be brought to the attention of the REB in a timely manner.  REB FULL BOARD MEETING REVIEW DATE:   June 10, 2014     DOCUMENTS INCLUDED IN THIS APPROVAL: DATE DOCUMENTS APPROVED:  141   Document Name Version Date Protocol: Protocol Version 4 July 2, 2014 Consent Forms: Informed Consent Version 4 July 2, 2014 Advertisements: Advertisement Version 3 June 24, 2014 Questionnaire, Questionnaire Cover Letter, Tests: ETF Subject Data sheet Version 1 May 5, 2014 ETF Questionnaire Version 3 June 24, 2014    July 2, 2014 CERTIFICATION: In respect of clinical trials:  1. The membership of this Research Ethics Board complies with the membership requirements for Research Ethics Boards defined in Division 5 of the Food and Drug Regulations. 2. The Research Ethics Board carries out its functions in a manner consistent with Good Clinical Practices. 3. This Research Ethics Board has reviewed and approved the clinical trial protocol and informed consent form for the trial which is to be conducted by the qualified investigator named above at the specified clinical trial site. This approval and the views of this Research Ethics Board have been documented in writing.    The documentation included for the above-named project has been reviewed by the UBC CREB, and the research study, as presented in the documentation, was found to be acceptable on ethical grounds for research involving human subjects and was approved by the UBC CREB.    Approval of the Clinical Research Ethics Board by one of:   Dr. Peter Loewen, Chair Dr. Stephen Hoption Cann, Associate Chair       142  Appendix C: NX1, NX2, HX1, and HX2 CO2 reactivity protocol raw data Table C.1: Subject demographic raw data for each participant SUBJECT AGE WEIGHT (KG) HEIGHT (CM) BMI (KG/M2) PFO (REST) PFO (VAL) 1 23 63.6 178 20.1 - + 2 24 89 178 28.1 - - 3 35 97.7 188 27.6 - + 4 39 68 174 22.5 - + 5 - - - - - - 6 22 68 181 20.8 - + 7 - - - - - - 8 29 103 176 33.3 - - 9 - - - - - - 10 20 71 178 22.4 - - 11 26 57 157 23.2 - - 12 29 51 163 19.1 - + 13 21 61 160 23.9 - - 14 21 72.3 172 24.4 - + 15 21 73 178 23 - + 16 21 65.5 175 21.4 - - 17 27 77 180 23.8 - + 18 29 76 175 24.8 - - MEAN 25.8 72.8 174.2 23.8 - - SEM 1.4 3.6 2.1 0.9 - -  Definition of abbreviations: PFO = patent foramen ovale; VAL = Valsalva maneuver;     143   Table C.2: Pulmonary function raw data for each participant. SUBJECT FVC %PRE FEV1 %PRE FEV1/FVC (%) 1 4.8 92 3.5 78 73 2 5.9 109 4.8 104 81 3 5.3 94 4.3 88 81 4 5.5 116 4.1 105 74 5 - - - - - 6 5.2 92 4.1 88 78 7 - - - - - 8 7.3 139 5.9 135 81 9 - - - - - 10 4.9 97 3.8 86 78 11 4 115 3.2 111 80 12 3.9 108 2.5 85 65 13 4.1 112 3.4 111 83 14 5.7 120 4.5 108 79 15 5.7 104 5.0 111 89 16 5.2 98 4.4 100 85 17 5.8 106 4.7 101 81 18 6.1 119 4.7 110 80 MEAN 5.3 108.0 4.2 101.4 79.2 SEM 0.2 3.3 0.2 3.7 1.4  Definition of abbreviations: FVC = forced vital capacity; %PRE = percent predicted; FEV1 = forced expired volume in one second.     144   Table C.3: Pulmonary raw data for each participant SUBJECT TLC (L) %PRE VC (L) %PRE IC (L) 1 5.3 77 5.0 92 2.7 2 7.6 111 6.2 114 4.5 3 6.8 89 5.7 101 4.1 4 7.2 109 5.9 124 3.3 5 - - - - - 6 6.5 92 5.3 94 3.1 7 - - - - - 8 - - - - - 9 - - - - - 10 5.5 84 5.0 107 2.8 11 5.0 108 4.2 122 2.9 12 5.1 101 3.6 111 2.3 13 4.7 98 4.2 114 2.5 14 7.2 120 5.7 132 3.8 15 7.4 108 5.9 108 4.5 16 6.6 101 5.2 98 3.3 17 7.1 102 5.8 3.8 3.8 18 7.8 118 6.2 121 4.1 MEAN 6.4 101.2 5.3 102.9 3.4 SEM 0.2 3.2 0.2 7.9 0.1  Definition of abbreviations: TLC = total lung capacity; %PRE = percent predicted; VC = vital capacity; IC = inspiratory capacity.    145  Table C.4: Subject demographic raw data for each participant SUBJECT FRC (L) %PRE ERV RV %PRE 1 2.58 78 2.02 0.56 55 2 3.15 95 1.86 1.43 95 3 2.72 70 1.39 1.1 57 4 3.88 119 2.73 1.33 77 5 - - - - - 6 3.4 99 2.09 1.2 79 7 - - - - - 8 - - - - - 9 - - - - - 10 2.62 82 2.1 0.48 37 11 2.14 84 1.51 0.84 70 12 2.79 101 1.67 1.08 78 13 2.26 85 1.77 0.59 51 14 3.4 115 1.95 1.47 121 15 2.94 89 1.32 1.52 104 16 3.29 104 2.09 1.46 103 17 3.33 97 2 1.32 82 18 3.69 114 2.2 1.61 106 MEAN 3.0 95.1 1.9 1.1 79.6 SEM 0.1 3.7 0.0 0.0 6.2 Definition of abbreviations: FRC = functional residual capacity; %PRE = percent predicted; ERV = expiratory residual volume; RV = residual volume.     146   Table C.5: Subject demographic raw data for each participant SUBJECT DLCO %PRE DLCO/VA %PRE VA %PRE 1 36.3 98 6.7 126 5.3 78 2 42.4 115 6.1 115 6.9 101 3 32.1 89 4.7 99 6.7 90 4 30.9 95 4.6 94 6.6 101 5 - - - - -  6 38.5 102 6.1 114 6.3 89 7 - - - - -  8 34 96 4.6 87 7.3 110 9 - - - - -  10 25.8 68 4.6 84 5.5 81 11 23.5 96 5.1 102 4.5 94 12 22.1 88 4.5 91 4.9 97 13 23.5 92 5.1 102 4.5 91 14 27.6 75 4 71 6.9 106 15 37.2 100 5.5 103 6.6 97 16 35 96 5.6 103 6.2 93 17 38.8 107 5.4 104 7.1 103 18 30.5 86 4.4 83 6.9 104 MEAN 31.8 93.5 5.1 98.5 6.1 95.6 SEM 1.6 3.0 0.1 3.6 0.2 2.3 Definition of abbreviations: DLCO = diffusion capacity of the lung for carbon monoxide transfer; %PRE = percent predicted; DLCO/VA = DLCO corrected for alveolar ventilation; VA = alveolar ventilation.     147   Table C.6: Minute ventilation (l/min) raw data for each participant NX1 CO2 reactivity protocol.  Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system.  SUBJECT AB AB-DEF -8 -4 0 4 8 1 12.8 15.6 19.6 21.5 19.2 25.0 35.1 2 12.2 14.9 23.9 21.3 15.0 35.1 58.8 3 - - - - - - - 4 10.2 19.1 30.0 23.5 18.0 26.0 30.0 5 - - - - - - - 6 13.9 21.0 21.4 16.1 17.6 27.5 41.6 7 - - - - - - - 8 13.6 24.1 17.9 21.4 21.1 18.6 53.5 9 - - - - - - - 10 11.1 14.3 18.0 15.5 14.4 19.7 28.6 11 10.5 13.9 17.2 15.1 16.1 23.9 29.3 12 11.2 14.0 11.6 15.2 15.0 20.1 25.5 13 14.4 20.2 28.1 27.6 24.4 49.2 64.2 14 10.0 9.4 19.2 15.6 13.0 16.9 24.4 15 16.1 14.3 25.7 27.1 17.2 24.1 36.6 16 14.3 21.2 28.5 23.8 21.0 27.8 38.1 17 13.3 19.4 23.1 22.9 17.0 35.7 48.2 18 9.4 14.7 26.0 21.6 17.3 19.9 30.2 MEAN 12.4 16.9 22.1 20.6 17.6 26.4 38.9 SEM 0.5 1.0 1.3 1.1 0.7 2.2 3.2  148  Table C.7: Minute ventilation (l/min) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 16.2 21.6 31.4 20.1 19.4 31.6 42.9 2 14.5 17.6 24.4 27.5 19.4 30.4 59.6 3 - - - - - - - 4 12.3 15.3 23.8 24.8 22.0 27.2 34.1 5 - - - - - - - 6 14.4 15.7 30.1 24.2 22.6 26.9 42.2 7 - - - - - - - 8 14.0 19.7 26.9 21.4 16.4 28.5 48.9 9 - - - - - - - 10 12.7 16.5 17.7 18.7 17.5 23.2 26.9 11 9.8 12.2 15.0 13.5 14.0 22.0 32.5 12 12.8 13.1 11.4 15.1 9.8 18.8 27.6 13 13.9 21.1 25.7 28.9 18.5 40.0 67.6 14 11.4 14.2 17.4 18.0 16.8 22.4 28.4 15 16.5 17.8 28.7 27.6 20.0 29.2 43.5 16 12.3 14.0 24.4 25.8 24.9 26.3 38.9 17 10.4 16.4 19.5 19.0 22.0 37.7 47.6 18 11.9 15.1 25.2 21.8 16.4 18.5 33.6 MEAN 13.1 16.5 23.0 21.9 18.5 27.3 41.0 SEM 0.5 0.7 1.5 1.2 0.9 1.6 3.1 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system.    149   Table C.8: Minute ventilation (l/min) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system.   SUBJECT AB AB-DEF -8 -4 0 4 8 1 14.9 34.4 16.0 18.3 35.4 49.2 62.3 2 11.7 30.2 26.5 28.0 21.9 45.6 81.4 3 10.5 26.9 23.5 21.4 32.3 50.5 70.5 4 12.3 24.3 23.7 27.5 28.5 32.9 38.8 5 - - - - - - - 6 15.0 31.6 24.5 27.8 32.8 51.6 64.2 7 - - - - - - - 8 13.2 30.0 24.2 20.8 23.8 42.7 56.6 9 - - - - - - - 10 16.7 21.7 17.5 15.4 20.7 27.8 34.9 11 10.6 22.4 19.7 15.0 21.5 30.0 43.1 12 9.8 18.6 12.5 14.4 18.7 23.7 33.2 13 - - - - - - - 14 11.4 21.8 21.3 21.1 24.9 29.4 40.1 15 13.8 41.1 27.1 25.7 26.8 45.3 68.5 16 14.0 27.9 30.2 28.5 30.5 37.1 50.1 17 13.5 23.2 22.3 24.7 28.2 50.2 62.0 18 11.5 28.5 20.6 22.3 27.4 50.8 83.5 MEAN 12.8 27.3 22.1 22.2 26.7 40.5 56.4 SEM 0.5 1.5 1.2 1.2 1.2 2.5 4.2  150   Table C.9: Minute ventilation (l/min) raw data for each participant during the HX2 CO2 reactivity protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system.  SUBJECT AB AB-DEF -8 -4 0 4 8 1 14.9 43.4 18.8 23.5 41.6 55.8 66.9 2 11.7 27.1 23.2 18.0 22.2 60.1 100.0 3 10.5 35.3 23.1 20.5 26.7 49.0 63.6 4 12.3 25.1 24.1 19.9 28.0 34.4 50.3 5 - - - - - - - 6 15.0 30.5 23.4 22.6 32.8 49.5 58.3 7 - - - - - - - 8 13.2 28.7 19.6 27.0 24.5 35.0 57.5 9 - - - - - - - 10 16.7 25.9 15.3 18.2 23.3 30.9 37.4 11 10.6 28.6 17.7 17.5 24.0 34.0 41.5 12 9.8 22.8 12.6 14.7 23.9 26.1 34.4 13 - - - - - - - 14 11.4 22.8 21.1 17.9 24.4 43.8 47.4 15 13.8 27.6 28.3 15.1 29.6 62.6 69.4 16 14.0 25.8 26.6 31.8 30.8 35.6 55.1 17 13.5 33.8 25.1 24.2 31.5 44.4 59.7 18 11.5 24.1 21.2 18.4 22.8 40.2 75.6 MEAN 12.8 28.7 21.4 20.7 27.6 43.0 58.4 SEM 0.5 1.4 1.1 1.2 1.3 2.8 4.3  151   Table C.10. Tidal volume (l) raw data for each participant. NX1 CO2 reactivity protocol.SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.9 1.1 0.7 0.6 1.2 1.6 2.3 2 1.0 1.1 1.2 1.0 1.0 1.7 1.9 3 - - - - - - - 4 0.9 1.5 1.3 1.0 1.1 1.9 2.3 5 - - - - - - - 6 0.9 1.3 1.9 2.0 0.9 1.5 1.8 7 - - - - - - - 8 0.8 1.4 1.0 1.2 1.2 1.1 2.8 9 - - - - - - - 10 0.8 1.0 0.9 1.0 0.9 1.4 2.1 11 0.8 0.9 1.4 1.4 1.0 1.4 1.8 12 0.5 0.6 1.1 1.5 0.7 1.1 1.5 13 0.7 1.0 1.9 1.7 1.5 1.2 1.7 14 1.0 1.6 2.9 2.6 2.3 2.5 3.0 15 1.0 1.0 0.9 1.0 1.0 1.4 1.7 16 0.7 1.0 1.5 1.3 1.1 1.2 1.7 17 0.9 1.8 2.6 2.5 1.9 2.1 2.4 18 1.3 1.9 3.2 2.6 2.0 2.5 3.3 MEAN 0.9 1.2 1.6 1.5 1.3 1.6 2.1 SEM 0.0 0.0 0.2 0.1 0.1 0.1 0.1 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system.    152   Table C.11. Tidal volume (l) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.9 1.4 0.9 0.7 1.0 1.6 2.1 2 0.9 1.1 0.9 0.8 1.1 1.4 2.1 3 - - - - - - - 4 0.8 1.1 1.0 1.0 1.0 1.6 2.3 5 - - - - - - - 6 0.7 1.1 2.5 2.6 1.3 1.6 1.9 7 - - - - - - - 8 0.8 1.0 1.5 1.2 1.0 1.4 2.5 9 - - - - - - - 10 1.1 1.2 2.3 2.6 1.1 1.6 2.1 11 0.7 0.9 1.4 1.4 0.9 1.6 1.8 12 0.5 0.7 1.0 1.6 0.7 1.6 1.6 13 0.7 1.1 1.9 1.9 0.9 1.2 1.9 14 0.9 1.9 2.5 2.7 2.9 3.0 3.2 15 1.0 1.1 1.2 0.9 0.9 1.3 1.8 16 0.7 0.8 1.8 1.8 1.1 1.3 1.8 17 1.0 1.4 2.5 2.4 2.1 2.0 2.4 18 0.9 1.6 2.6 2.4 1.9 1.8 3.1 MEAN 0.8 1.2 1.7 1.7 1.3 1.6 2.2 SEM 0.0 0.0 0.1 0.1 0.1 0.1 0.1  153   Table C.12. Tidal volume (l) raw data for each participant during the HX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.8 1.7 1.0 1.1 1.7 2.2 2.5 2 0.9 1.3 0.8 0.8 1.1 2.1 2.2 3 1.0 1.6 2.3 2.1 1.8 2.5 2.9 4 0.8 1.8 2.1 2.0 2.2 2.6 2.7 5 - - - - - - - 6 0.8 1.3 1.8 2.2 1.4 2.1 2.2 7 - - - - - - - 8 0.8 1.5 1.2 1.0 1.1 2.3 3.1 9 - - - - - - - 10 0.6 1.6 1.5 1.4 1.5 2.0 2.7 11 0.7 1.2 1.2 0.9 1.2 1.8 2.0 12 0.6 1.4 1.3 1.3 1.5 1.8 1.8 13 - - - - - - - 14 0.9 3.3 3.3 3.3 3.4 3.2 2.5 15 1.0 1.7 0.8 0.9 1.1 1.7 2.4 16 0.7 1.3 1.6 1.3 1.4 1.8 2.4 17 0.8 1.9 1.5 1.6 1.6 2.7 3.1 18 1.0 1.9 2.1 2.2 2.1 3.2 3.2 MEAN 0.8 1.7 1.6 1.6 1.6 2.3 2.5 SEM 0.0 0.1 0.1 0.1 0.1 0.1 0.1 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     154   Table C.13. Tidal Volume (l) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.8 1.9 1.7 1.2 1.8 2.3 2.6 2 0.9 1.6 0.7 0.8 1.1 1.8 2.4 3 1.0 2.2 2.3 2.1 1.9 2.7 3.0 4 0.8 1.8 1.8 1.7 2.4 3.1 3.2 5 - - - - - - - 6 0.8 1.5 2.1 2.1 1.7 2.0 1.9 7 - - - - - - - 8 0.8 1.5 1.0 1.4 1.3 1.7 3.0 9 - - - - - - - 10 0.6 1.8 1.1 1.3 1.7 2.6 2.9 11 0.7 1.6 1.2 1.1 1.4 1.9 2.2 12 0.6 1.2 1.3 1.1 1.3 1.6 1.7 13 - - - - - - - 14 0.9 2.8 3.0 2.9 2.8 2.7 2.6 15 1.0 1.2 1.1 1.0 1.4 2.2 2.5 16 0.7 1.2 1.4 1.5 1.3 1.7 2.3 17 0.8 2.3 1.6 1.7 2.0 2.4 3.0 18 1.0 3.0 2.7 2.2 2.5 3.5 3.2 MEAN 0.8 1.8 1.6 1.6 1.8 2.3 2.6 SEM 0.0 0.1 0.1 0.1 0.1 0.1 0.1 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     155   Table C.14. Breathing frequency (breaths/min) raw data for each participant NX1 CO2 reactivity protocol. Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system  SUBJECT AB AB-DEF -8 -4 0 4 8 1 13.1 13.3 26.8 35.3 15.9 15.1 14.8 2 12.0 13.0 19.4 21.2 14.9 19.8 30.5 3 - - - - - - - 4 10.7 12.4 21.9 22.1 16.0 13.3 12.8 5 - - - - - - - 6 15.4 15.5 11.1 8.1 18.2 17.9 22.6 7 - - - - - - - 8 16.4 16.4 17.5 16.7 16.4 15.8 18.5 9 - - - - - - - 10 13.2 14.3 19.2 15.4 15.1 14.0 13.5 11 12.8 15.4 12.2 10.4 15.9 16.5 15.9 12 19.2 20.8 10.0 9.6 19.7 17.8 17.2 13 19.9 20.1 14.1 15.5 16.3 40.0 37.4 14 9.7 5.8 6.5 5.9 5.6 6.6 8.1 15 16.1 13.3 26.9 25.9 16.0 16.8 21.5 16 19.7 20.2 18.4 17.8 18.9 22.4 21.7 17 14.0 10.7 8.7 9.0 9.0 17.0 20.2 18 7.3 7.5 8.0 8.2 8.3 7.9 9.1 MEAN 14.2 14.2 15.8 15.8 14.7 17.2 18.8 SEM 0.9 1.1 1.7 2.1 1.0 1.9 2.0  156   Table C.15. Breathing frequency (breaths/min) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 17.2 15.0 34.6 26.7 18.0 19.8 30.6 2 14.6 15.8 25.1 31.1 16.8 21.0 28.4 3 - - - - - - - 4 14.3 13.8 22.3 23.0 21.4 16.8 14.5 5 - - - - - - - 6 18.3 14.2 12.3 9.1 17.5 16.5 21.6 7 - - - - - - - 8 17.3 18.4 17.6 16.5 15.5 20.3 19.1 9 - - - - - - - 10 11.9 13.6 7.6 7.0 15.3 14.1 12.4 11 12.8 12.6 10.1 9.0 14.2 13.6 17.3 12 21.6 17.4 10.5 9.4 12.5 11.6 16.6 13 17.7 18.3 13.4 14.7 19.5 32.1 34.5 14 12.0 7.4 6.8 6.6 5.6 7.2 8.9 15 16.3 14.9 22.6 30.6 22.1 21.9 23.7 16 15.9 15.8 13.0 13.8 21.7 20.1 21.1 17 9.5 11.1 7.5 7.8 10.4 18.5 19.5 18 12.3 9.1 9.6 8.8 8.4 9.8 10.5 MEAN 15.1 14.1 15.2 15.3 15.6 17.4 19.9 SEM 0.8 0.8 2.1 2.2 1.2 1.5 1.9  157   Table C.16. Breathing frequency (breaths/min) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 17.6 19.5 15.5 16.6 20.5 22.3 24.9 2 12.3 22.1 30.3 33.4 19.8 21.6 36.7 3 10.6 16.3 10.1 9.8 17.2 20.0 24.0 4 14.3 13.3 11.1 13.6 12.8 12.4 14.2 5 - - - - - - - 6 17.0 23.2 13.8 12.5 23.1 24.6 28.4 7 - - - - - - - 8 16.2 19.4 18.9 19.0 19.9 18.2 17.7 9 - - - - - - - 10 24.4 13.1 11.5 11.0 13.5 13.3 12.8 11 14.0 18.5 15.5 16.2 16.6 16.3 20.8 12 15.2 13.4 9.2 10.8 12.3 12.6 18.4 13 - - - - - - - 14 12.0 6.5 6.4 6.4 7.3 9.1 15.9 15 13.3 24.1 31.7 27.3 23.1 25.4 28.1 16 17.7 20.6 18.3 22.2 21.6 20.4 20.6 17 15.6 12.3 14.8 15.2 16.9 18.1 19.6 18 10.5 14.9 9.4 9.9 12.6 15.7 25.7 MEAN 15.1 17.0 15.5 16.0 16.9 17.8 22.0 SEM 0.9 1.2 1.9 1.9 1.2 1.2 1.6  158   Table C.17. Breathing frequency (breaths/min) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 17.6 21.9 10.7 19.3 22.5 24.1 25.7 2 12.3 18.0 29.4 22.5 18.6 33.1 41.6 3 10.6 15.5 9.8 9.3 13.7 17.7 20.8 4 14.3 13.4 13.2 11.1 11.6 11.0 15.3 5 - - - - - - - 6 17.0 19.9 10.8 10.7 19.1 24.6 30.1 7 - - - - - - - 8 16.2 18.9 17.9 19.3 17.7 19.4 18.6 9 - - - - - - - 10 24.4 14.4 13.2 13.9 13.5 11.8 12.8 11 14.0 17.2 13.9 15.7 16.4 17.7 18.8 12 15.2 17.9 9.9 12.7 17.7 15.5 19.7 13 - - - - - - - 14 12.0 8.0 7.0 6.2 8.5 15.6 18.2 15 13.3 21.4 24.3 15.0 20.7 28.4 27.6 16 17.7 21.1 19.1 21.2 23.5 20.3 23.5 17 15.6 14.7 15.6 14.3 15.7 18.5 19.8 18 10.5 7.7 7.6 8.1 9.0 11.2 22.9 MEAN 15.1 16.4 14.4 14.2 16.3 19.2 22.5 SEM 0.9 1.1 1.6 1.2 1.1 1.6 1.8 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    159   Table C.18. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each participantNX1 CO2 reactivity protocol.SUBJECT AB AB-DEF -8 -4 0 4 8 1 43.4 44.3 37.7 41.2 45.0 48.9 53.0 2 43.4 45.1 36.6 41.3 45.0 49.1 53.1 3 - - - - - - - 4 41.6 42.8 34.3 38.3 42.5 46.5 50.8 5 - - - - - - - 6 37.8 39.8 30.9 36.1 40.4 44.1 47.5 7 - - - - - - - 8 40.4 42.0 33.7 38.2 42.4 46.0 50.1 9 - - - - - - - 10 43.8 44.1 35.8 40.2 44.4 48.3 52.1 11 36.7 38.9 30.9 34.9 39.0 43.1 47.1 12 38.1 39.5 29.9 35.5 39.4 43.2 47.2 13 37.7 38.9 31.0 34.9 38.3 43.0 47.1 14 38.9 38.7 31.1 35.3 40.4 43.9 47.9 15 37.3 37.9 30.2 34.1 37.9 42.2 46.1 16 40.5 42.6 35.3 38.7 42.5 46.4 50.8 17 37.9 41.6 32.2 36.7 40.7 44.6 48.6 18 35.0 37.2 28.7 32.7 37.4 41.0 44.7 MEAN 39.5 41.0 32.7 37.0 41.1 45.0 49.0 SEM 0.7 0.6 0.7 0.6 0.6 0.6 0.6 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    160   Table C.19. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 40.9 45.5 36.7 40.8 44.3 49.2 53.3 2 43.0 46.2 37.2 41.6 46.2 50.0 54.4 3 - - - - - - - 4 40.6 43.0 34.6 38.7 43.0 47.2 51.8 5 - - - - - - - 6 38.6 39.0 31.6 34.9 40.4 43.7 47.6 7 - - - - - - - 8 41.4 42.0 33.7 38.2 41.7 46.2 51.2 9 - - - - - - - 10 41.6 45.2 37.0 40.8 45.5 49.8 53.7 11 38.3 40.3 31.8 36.0 40.3 44.6 48.6 12 37.7 40.1 30.5 35.1 39.5 44.1 47.8 13 38.3 40.2 31.3 35.9 39.7 43.9 48.6 14 38.3 39.4 30.7 35.3 39.5 44.1 47.6 15 36.5 38.7 29.9 34.1 38.1 42.7 46.7 16 41.9 42.8 33.9 38.5 42.8 46.8 50.8 17 40.2 41.3 32.4 36.8 41.8 45.5 49.6 18 35.8 37.4 28.4 33.5 37.7 40.9 46.0 MEAN 39.5 41.5 32.8 37.2 41.5 45.6 49.8 SEM 0.5 0.7 0.7 0.6 0.6 0.7 0.7  161   Table C.20. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 41.4 43.0 35.1 39.0 42.9 47.0 51.0 2 42.3 43.5 35.2 39.3 43.8 47.3 51.8 3 37.3 38.4 29.6 34.1 38.1 42.1 46.2 4 41.0 42.5 34.5 38.6 42.7 46.8 50.8 5 - - - - - - - 6 36.2 37.3 29.0 32.7 37.0 40.9 44.9 7 - - - - - - - 8 39.1 41.3 31.6 36.7 40.9 44.8 48.7 9 - - - - - - - 10 42.5 42.7 34.9 39.3 43.1 46.9 51.0 11 36.7 37.6 29.5 33.6 37.4 41.7 45.6 12 38.7 39.2 30.2 35.3 39.2 43.2 47.0 13 - - - - - - - 14 38.3 39.7 31.2 35.7 39.9 43.7 47.5 15 37.7 39.2 31.0 35.3 39.1 43.2 47.3 16 41.2 41.0 33.7 37.8 42.2 45.5 49.8 17 38.0 39.0 30.5 35.0 39.1 43.2 47.1 18 37.0 37.7 29.5 33.8 37.7 41.7 45.8 MEAN 39.1 40.2 31.8 36.2 40.2 44.1 48.2 SEM 0.5 0.5 0.6 0.5 0.6 0.5 0.5 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    162   Table C.21. Partial pressure end-tidal carbon dioxide (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 41.4 43.2 33.2 38.9 43.0 47.1 51.0 2 42.3 43.7 35.2 39.2 43.7 48.3 51.2 3 37.3 38.5 29.5 35.4 38.7 42.7 47.0 4 41.0 43.0 34.7 38.3 43.1 46.8 51.2 5 - - - - - - - 6 36.2 37.2 28.0 32.9 37.2 41.0 45.0 7 - - - - - - - 8 39.1 41.2 31.9 36.0 41.5 45.7 49.3 9 - - - - - - - 10 42.5 44.8 35.6 40.4 43.3 47.9 52.2 11 36.7 38.5 29.9 34.2 38.4 42.8 46.8 12 38.7 40.7 30.4 36.1 39.8 44.0 48.2 13 - - - - - - - 14 38.3 40.8 32.1 37.1 40.6 44.6 48.9 15 37.7 39.2 30.6 34.8 39.1 43.7 47.6 16 41.2 42.1 33.4 37.7 41.6 44.8 50.1 17 38.0 39.3 30.3 35.1 39.3 43.1 47.7 18 37.0 38.6 29.4 34.3 38.9 42.7 46.8 MEAN 39.1 40.8 31.7 36.5 40.6 44.7 48.8 SEM 0.5 0.5 0.6 0.5 0.5 0.5 0.5  163   Table C.22. Partial pressure end-tidal oxygen (mmHg) raw data for each participant. NX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 89.1 90.2 88.6 89.1 89.8 90.8 89.1 2 92.8 87.7 89.8 87.1 90.1 89.5 89.4 3 - - - - - - - 4 93.1 92.4 93.7 93.6 94.1 93.9 93.9 5 - - - - - - - 6 95.7 91.8 91.8 88.2 90.3 92.0 92.3 7 - - - - - - - 8 92.6 93.3 89.6 91.3 94.8 93.1 92.9 9 - - - - - - - 10 88.6 87.5 89.0 88.9 88.2 88.5 89.0 11 102.5 103.6 102.9 103.0 103.1 103.1 102.5 12 93.2 91.7 94.9 93.4 90.9 93.8 93.8 13 100.6 99.3 96.6 97.9 99.9 98.0 97.2 14 96.9 100.3 95.0 94.9 93.4 95.4 95.8 15 99.2 94.4 99.2 94.8 96.0 96.3 96.1 16 98.6 96.0 93.9 90.7 93.5 93.9 93.8 17 98.2 92.2 93.0 92.4 93.8 93.1 93.9 18 96.1 100.1 99.8 100.3 90.3 99.7 100.3 MEAN 95.5 94.3 94.1 93.3 93.4 94.4 94.3 SEM 1.0 1.2 1.1 1.2 1.0 1.0 1.0 Definition of abbreviations: NX1 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    164   Table C.23. Partial pressure end-tidal oxygen (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 96.2 89.3 91.1 82.4 88.2 88.9 90 2 92.3 88.6 89.6 87.4 88.9 89.0 89.3 3 - - - - - - - 4 92.1 94.3 94.5 94.0 91.1 92.9 93.5 5 - - - - - - - 6 92.0 92.5 90.9 92.0 92.4 91.2 92.3 7 - - - - - - - 8 90.7 93.9 91.5 90.2 94.5 91.1 91.1 9 - - - - - - - 10 90.1 88.2 88.5 89.8 88.4 89.7 89.5 11 98.5 102.6 103.1 103.3 99.6 103.0 102.9 12 97.8 93.3 94.7 94.0 91.5 92.8 93.4 13 98.3 98.1 98.9 97.2 100.1 97.8 98.5 14 93.2 92.7 93.6 93.5 94.8 92.6 94.7 15 100.3 96.6 96.8 95.6 96.8 97.1 97.3 16 93.8 92.8 93.4 93.0 92.6 94.8 93.7 17 92.7 93.6 92.6 94.1 93.5 92.9 92.9 18 100.8 99.3 100.8 98.9 98.1 102.1 100.4 MEAN 94.9 94.0 94.3 93.2 93.6 94.0 94.2 SEM 0.9 1.0 1.0 1.2 1.0 1.1 1.0  165   Table C.24. Partial pressure end-tidal oxygen (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 92.2 50.4 48.4 50.3 50.2 50.2 50.0 2 96.8 50.6 49.8 50.5 50.1 51.4 50.9 3 95.7 50.9 52.7 51.4 50.8 51.1 51.1 4 93.4 51.0 50.9 51.1 50.8 50.7 51.1 5 - - - - - - - 6 97.4 50.6 49.9 50.3 50.8 51.0 51.0 7 - - - - - - - 8 94.4 50.3 51.0 52.3 51.3 51.0 51.3 9 - - - - - - - 10 90.9 50.4 50.1 49.5 50.2 50.8 50.3 11 99.8 51.2 51.0 50.4 51.2 51.0 51.5 12 93.7 51.2 49.1 51.5 50.8 50.2 50.5 13 - - - - - - - 14 93.2 50.8 50.3 50.6 50.7 50.9 50.7 15 92.7 51.2 50.6 51.1 51.0 51.1 51.4 16 94.5 51.5 51.7 50.7 51.0 51.3 51.3 17 94.6 50.3 51.8 51.5 51.5 50.9 51.0 18 91.7 50.0 50.1 49.9 50.0 50.4 50.2 MEAN 94.3 50.7 50.5 50.8 50.7 50.9 50.9 SEM 0.6 0.1 0.2 0.1 0.1 0.0 0.1  166   Table C.25. Partial pressure end-tidal oxygen (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 92.2 50.0 50.9 49.7 50.1 50.0 50.1 2 96.8 50.8 50.6 50.0 50.7 51.0 50.8 3 95.7 51.2 51.7 50.0 51.1 51.0 51.1 4 93.4 50.4 51.0 50.8 50.8 50.8 50.7 5 - - - - - - - 6 97.4 50.3 51.6 49.8 50.8 50.5 50.9 7 - - - - - - - 8 94.4 50.2 52.3 53.4 51.3 50.6 51.0 9 - - - - - - - 10 90.9 49.7 52.9 48.2 50.1 50.4 50.4 11 99.8 50.6 51.1 50.6 51.0 51.1 51.0 12 93.7 50.1 51.8 49.7 51.1 50.7 50.6 13 - - - - - - - 14 93.2 50.5 50.2 50.0 51.0 50.6 51.5 15 92.7 50.8 51.4 51.7 51.1 51.4 51.7 16 94.5 51.6 51.4 51.3 51.5 51.2 51.5 17 94.6 50.9 50.5 50.9 51.0 50.9 50.7 18 91.7 50.2 51.0 49.9 50.1 50.0 50.3 MEAN 94.3 50.5 51.3 50.4 50.8 50.7 50.9 SEM 0.6 0.1 0.1 0.3 0.1 0.1 0.1 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    167   Table C.26. Partial pressure of inspired carbon dioxide (mmHg) raw data for each participant NX1 CO2 reactivity protocol.  SUBJECT AB AB-DEF -8 -4 0 4 8 1 1.0 18.0 1.0 9.2 23.8 34.3 43.3 2 4.2 18.5 18.2 24.0 19.3 40.3 48.0 3 - - - - - - - 4 3.5 25.1 20.2 24.3 25.5 35.1 41.8 5 - - - - - - - 6 5.3 23.9 12.4 20.3 24.9 33.6 40.8 7 - - - - - - - 8 6.4 29.2 10.2 18.6 25.6 28.6 43.9 9 - - - - - - - 10 1.2 15.3 12.1 13.0 16.0 32.4 41.3 11 4.3 20.3 12.3 19.1 23.2 34.9 40.3 12 8.5 20.6 2.4 21.5 24.1 33.4 39.2 13 1.7 20.9 19.3 22.5 22.8 35.1 39.5 14 2.4 3.1 11.3 17.1 20.8 28.6 36.3 15 0.6 1.5 1.5 12.4 9.6 24.9 32.8 16 7.6 27.4 20.7 24.6 27.1 37.6 44.3 17 5.3 26.2 15.1 23.6 24.6 36.0 41.8 18 0.6 16.2 14.4 17.0 21.3 27.1 36.2 MEAN 3.8 19.0 12.2 19.1 22.0 33.0 40.7 SEM 0.6 2.1 1.7 1.2 1.1 1.0 0.9 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    168   Table C.27. Partial pressure of inspired carbon dioxide (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 1.0 26.7 15.1 10.4 23.3 38.3 44.4 2 5.8 29.0 21.5 29.3 27.9 39.6 47.2 3 - - - - - - - 4 5.3 22.5 19.0 27.2 30.2 38.0 43.6 5 - - - - - - - 6 5.8 15.3 18.5 20.8 26.4 33.0 41.1 7 - - - - - - - 8 7.0 22.1 18.0 21.0 20.1 36.6 42.8 9 - - - - - - - 10 1.0 21.6 16.6 22.6 25.1 35.5 42.5 11 4.3 20.8 11.1 18.2 23.2 35.2 41.5 12 9.5 20.5 7.7 19.9 15.8 33.6 40.5 13 1.1 23.5 16.6 24.7 20.4 34.6 40.7 14 4.0 18.4 7.5 18.8 22.7 33.5 36.3 15 0.7 13.7 8.9 12.0 12.8 28.2 35.3 16 6.4 13.9 18.1 26.4 31.2 36.4 43.9 17 3.2 23.9 14.2 21.1 29.4 37.7 42.3 18 0.7 13.6 11.5 17.6 18.9 24.6 37.9 MEAN 4.0 20.4 14.6 20.7 23.4 34.6 41.4 SEM 0.7 1.2 1.1 1.3 1.4 1.0 0.8 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    169   Table C.28. Partial pressure of inspired carbon dioxide (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 1.3 30.6 3.6 16.5 32.3 38.5 43.4 2 5.4 33.9 19.1 28.3 28.8 39.9 45.2 3 3.5 24.6 10.2 21.9 30.2 36.8 41.4 4 5.8 31.5 20.8 28.2 33.5 38.9 42.9 5 - - - - - - - 6 5.9 28.6 12.1 21.2 28.7 34.7 39.6 7 - - - - - - - 8 4.9 28.7 9.5 18.0 28.0 37.6 42.5 9 - - - - - - - 10 1.8 26.2 14.2 15.9 25.9 34.5 41.0 11 4.4 26.7 13.8 16.6 27.7 35.2 40.4 12 6.7 27.0 6.8 20.6 27.6 35.6 40.5 13 - - - - - - - 14 4.0 29.2 17.1 24.7 30.7 35.8 39.8 15 0.7 26.7 6.2 13.0 22.5 33.2 39.3 16 6.8 27.1 21.3 26.2 31.9 37.5 42.4 17 6.0 24.3 8.6 19.5 29.0 36.8 41.1 18 1.0 24.8 10.9 18.6 26.1 34.7 38.5 MEAN 4.2 27.8 12.4 20.6 28.8 36.4 41.3 SEM 0.5 0.7 1.4 1.2 0.7 0.4 0.4  170   Table C.29. Partial pressure of inspired carbon dioxide (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 1.3 32.3 2.6 20.4 33.2 39.2 43.3 2 5.4 29.5 18.6 15.9 30.8 42.3 41.5 3 3.5 29.3 14.2 22.6 28.1 36.4 40.9 4 5.8 32.9 22.1 26.1 34.6 39.4 44.6 5 - - - - - - - 6 5.9 28.3 8.6 18.3 28.5 34.8 39.3 7 - - - - - - - 8 4.9 29.6 7.6 20.8 30.6 36.1 43.2 9 - - - - - - - 10 1.8 30.2 5.8 20.5 29.7 37.4 42.9 11 4.4 29.9 12.0 20.9 29.4 36.1 40.4 12 6.7 30.4 2.4 21.3 30.5 35.9 41.4 13 - - - - - - - 14 4.0 29.3 16.4 22.2 30.2 37.9 41.7 15 0.7 22.9 9.6 8.1 25.0 34.2 40.0 16 6.8 30.8 16.1 28.8 32.3 36.5 42.8 17 6.0 29.4 13.8 22.6 29.9 37.2 41.7 18 1.0 25.9 10.9 16.4 24.6 34.2 39.6 MEAN 4.2 29.3 11.5 20.3 29.8 37.0 41.7 SEM 0.5 0.6 1.5 1.2 0.7 0.5 0.4 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     171   Table C.30. Core body temperature (C) raw data for each. NX1 CO2 reactivity protocol. Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 36.7 36.7 36.7 36.7 36.7 36.7 36.7 2 36.5 36.5 36.5 36.5 36.5 36.4 36.3 3 - - - - - - - 4 36.7 36.6 36.4 36.5 36.4 36.4 36.2 5 - - - - - - - 6 36.6 36.6 36.6 36.6 36.6 36.6 36.6 7 - - - - - - - 8 36.2 36.2 36.2 36.2 36.2 36.2 36.1 9 - - - - - - - 10 36.5 36.5 36.4 36.5 36.5 36.5 36.4 11 36.7 36.7 36.7 36.7 36.8 36.7 36.6 12 35.4 36.3 36.3 36.2 36.5 36.5 36.4 13 36.8 36.8 36.7 36.8 36.8 36.8 36.7 14 36.3 36.3 36.3 36.3 36.3 36.3 36.3 15 36.5 36.6 36.2 36.2 36.6 36.5 36.5 16 36.2 36.2 36.2 36.2 36.1 36.1 36.1 17 36.5 36.6 36.5 36.5 36.6 36.6 36.5 18 36.1 36.2 36.2 36.2 36.2 36.3 36.3 MEAN 36.4 36.5 36.4 36.4 36.5 36.5 36.4 SEM 0.0 0.0 0.0 0.0 0.0 0.0 0.0  172   Table C.31. Core body temperature (C) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 36.8 36.8 36.8 36.8 36.8 36.8 36.8 2 36.4 36.4 36.4 36.4 36.4 36.4 36.3 3 - - - - - - - 4 36.4 36.5 36.4 36.4 36.4 36.4 36.4 5 - - - - - - - 6 36.8 36.7 36.7 36.7 36.7 36.6 36.6 7 - - - - - - - 8 36.4 36.4 36.3 36.3 36.3 36.2 35.3 9 - - - - - - - 10 36.4 36.4 36.3 36.3 36.3 36.3 36.2 11 36.6 36.6 36.5 36.6 36.7 36.6 36.6 12 35.8 36.4 36.3 36.2 36.6 36.5 36.3 13 36.8 36.8 36.8 36.8 36.8 36.7 36.7 14 36.4 36.4 36.5 36.5 36.5 36.5 36.5 15 36.5 36.6 36.3 36.4 36.7 36.5 36.1 16 36.2 36.3 36.2 36.2 36.2 36.2 36.1 17 36.4 36.6 36.5 36.5 36.6 36.5 36.5 18 36.1 36.3 36.2 36.1 36.2 36.3 36.2 MEAN 36.4 36.5 36.4 36.4 36.5 36.5 36.3 SEM 0.0 0.0 0.0 0.0 0.0 0.0 0.0 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     173   Table C.32. Core body temperature (C) raw data for each participant during the HX1 CO2 reactivity protocol Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 36.8 36.8 36.8 36.8 36.8 36.8 36.8 2 36.5 36.4 36.3 36.4 36.4 36.4 36.2 3 36.4 36.2 36.2 36.4 36.3 36.2 35.9 4 36.5 36.5 36.3 36.4 36.4 36.4 36.3 5 - - - - - - - 6 36.7 36.5 36.5 36.6 36.6 36.6 36.5 7 - - - - - - - 8 35.4 35.3 35.3 35.2 35.2 35.2 35.3 9 - - - - - - - 10 36.3 36.3 36.3 36.3 36.4 36.4 36.4 11 36.7 36.4 36.4 36.5 36.5 36.4 36.3 12 36.3 36.4 36.4 36.3 36.4 36.4 36.3 13 - - - - - - - 14 36.4 36.5 36.4 36.5 36.5 36.4 36.4 15 36.5 36.1 36.1 36.2 36.4 35.9 35.4 16 36.3 36.0 36.0 36.0 36.0 36.0 35.9 17 36.6 36.3 36.4 36.4 36.5 36.4 36.3 18 36.4 36.3 36.3 36.3 36.4 36.4 36.2 MEAN 36.4 36.3 36.3 36.3 36.3 36.3 36.2 SEM 0.0 0.0 0.0 0.0 0.0 0.0 0.1  174   Table C.33. Core body temperature (C) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 36.8 36.8 36.8 36.8 36.8 36.8 36.8 2 36.5 36.2 36.3 36.3 36.3 36.3 36.2 3 36.4 36.4 36.3 36.4 36.4 36.2 36.2 4 36.5 36.3 36.2 36.3 36.3 36.3 36.2 5 - - - - - - - 6 36.7 36.6 36.6 36.6 36.6 36.6 36.5 7 - - - - - - - 8 35.4 35.6 35.6 35.5 35.5 35.4 35.4 9 - - - - - - - 10 36.3 36.3 36.3 36.4 36.4 36.4 36.4 11 36.7 36.6 36.5 36.6 36.6 36.6 36.5 12 36.3 36.3 36.3 36.4 36.5 36.5 36.4 13 - - - - - - - 14 36.4 36.3 36.4 36.5 36.4 36.4 36.2 15 36.5 35.8 35.9 36.3 36.3 35.5 35.4 16 36.3 36.2 36.1 36.1 36.1 36.0 35.9 17 36.6 36.5 36.5 36.5 36.5 36.5 36.4 18 36.4 36.4 36.3 36.4 36.5 36.4 36.3 MEAN 36.4 36.3 36.3 36.4 36.4 36.3 36.2 SEM 0.0 0.0 0.0 0.0 0.0 0.1 0.1 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     175   Table C.34. Oxygen saturation of hemoglobin (%) raw data for each participant. NX1 CO2 reactivity protocolSUBJECT AB AB-DEF -8 -4 0 4 8 1 98.1 98.0 98.1 97.0 97.5 97.9 97.7 2 98.3 96.6 97.2 97.3 97.2 97.3 96.9 3 - - - - - - - 4 97.3 98.3 98.3 98.0 97.8 98.2 97.8 5 - - - - - - - 6 96.4 97.0 97.8 96.2 95.4 95.9 96.4 7 - - - - - - - 8 96.8 98.1 98.0 97.7 97.7 97.1 98.0 9 - - - - - - - 10 97.6 97.1 98.2 97.4 97.3 97.1 96.9 11 99.3 98.8 98.9 99.3 98.4 98.4 98.4 12 98.4 98.4 99.4 99.4 98.7 98.4 98.4 13 98.0 98.6 99.2 98.4 98.4 98.6 98.9 14 97.1 97.1 97.3 96.7 96.2 96.5 97.2 15 96.1 94.5 96.6 96.9 96.7 96.3 96.4 16 97.1 96.7 97.6 97.6 97.2 97.1 96.3 17 98.3 98.2 98.4 98.3 98.3 98.2 98.3 18 98.8 98.9 99.1 99.3 99.1 98.8 98.4 MEAN 97.7 97.6 98.1 97.8 97.6 97.6 97.6 SEM 0.2 0.3 0.2 0.2 0.2 0.2 0.2 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     176   Table C.35. Oxygen saturation of hemoglobin (%) raw data for each participant during the NX2 CO2 reactivity protocol Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 98.4 97.6 98.2 97.7 97.7 97.1 96.9 2 97.8 97.9 98.3 98.0 97.8 97.6 97.7 3 - - - - - - - 4 97.9 98.3 98.4 98.3 98.3 98.3 98.3 5 - - - - - - - 6 95.3 95.7 98.2 97.3 96.3 96.3 96.6 7 - - - - - - - 8 96.4 96.6 97.6 96.9 96.7 96.9 96.6 9 - - - - - - - 10 98.2 97.9 98.4 98.4 97.7 97.6 97.2 11 98.4 98.7 99.3 99.3 99.2 98.6 98.4 12 99.4 99.1 99.4 99.4 98.4 98.4 98.4 13 98.4 98.4 99.1 99.0 98.4 98.4 98.9 14 97.1 98.3 98.3 97.9 97.2 97.1 97.4 15 96.4 96.2 97.8 97.6 96.9 96.1 97.4 16 95.2 95.4 97.3 96.9 96.6 96.5 96.5 17 98.5 98.3 99.0 98.4 98.3 98.3 98.3 18 99.1 98.8 99.0 99.3 99.4 99.0 98.4 MEAN 97.6 97.7 98.4 98.2 97.8 97.6 97.6 SEM 0.3 0.3 0.1 0.2 0.2 0.2 0.2  177   Table C.36. Oxygen saturation of hemoglobin (%) raw data for each participant during the HX1 CO2 reactivity protocol SUBJECT AB AB-DEF -8 -4 0 4 8 1 98.3 84.3 87.8 83.9 83.2 82.2 82.0 2 98.3 84.6 85.4 84.1 84.0 84.4 83.6 3 98.4 84.5 88.9 86.0 82.1 80.2 80.6 4 98.7 78.3 86.5 84.0 80.1 78.5 80.0 5 - - - - - - - 6 96.5 81.5 86.8 85.0 81.3 80.5 79.0 7 - - - - - - - 8 98.1 85.1 85.9 84.8 84.2 83.8 84.8 9 - - - - - - - 10 97.6 83.1 86.1 84.1 81.3 81.6 82.1 11 98.4 81.0 84.8 80.0 80.9 78.6 77.0 12 98.9 83.3 88.7 85.0 82.2 82.2 79.9 13 - - - - - - - 14 97.1 83.8 87.1 86.1 84.5 82.6 81.1 15 96.3 82.3 85.7 82.9 82.5 81.9 81.0 16 96.7 83.2 85.8 85.3 83.1 81.7 81.4 17 98.3 86.3 88.7 86.8 86.0 85.2 84.2 18 98.4 87.0 90.2 87.6 86.4 85.6 83.5 MEAN 97.9 83.4 87.0 84.7 83.0 82.1 81.4 SEM 0.2 0.5 0.4 0.4 0.4 0.5 0.5 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     178   Table C.37. Oxygen saturation of hemoglobin (%) raw data for each participant during the HX2 CO2 reactivity protocol.  SUBJECT AB AB-DEF -8 -4 0 4 8 1 98.3 85.4 88.5 84.8 84.4 83.3 81.7 2 98.3 85.7 86.3 85.5 85.0 83.2 81.7 3 98.4 83.7 87.7 82.6 82.7 82.3 83.2 4 98.7 80.6 86.0 83.2 78.9 78.1 77.3 5 - - - - - - - 6 96.5 82.2 87.3 83.9 80.7 80.3 79.2 7 - - - - - - - 8 98.1 86.2 88.6 83.9 85.7 84.4 83.1 9 - - - - - - - 10 97.6 84.0 88.0 82.0 82.0 81.8 82.3 11 98.4 80.2 84.2 80.7 80.6 78.7 77.1 12 98.9 82.4 86.6 82.6 81.0 80.3 78.5 13 - - - - - - - 14 97.1 84.4 88.0 84.4 82.8 82.7 79.3 15 96.3 83.3 85.5 81.6 81.1 80.5 86.2 16 96.7 80.8 84.7 84.5 83.0 82.0 80.2 17 98.3 85.3 89.3 87.4 85.2 84.3 83.2 18 98.4 86.9 89.4 88.0 86.3 85.4 83.8 MEAN 97.9 83.7 87.1 83.9 82.8 82.0 81.2 SEM 0.2 0.5 0.4 0.5 0.5 0.5 0.6 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     179   Table C.38. Heart rate (bpm) raw data for each participant. NX1 CO2 reactivity protocol Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 55.2 57.3 62.4 61.7 67.1 73.8 76.6 2 69.5 71.7 74.3 70.0 69.4 77.1 83.9 3 - - - - - - - 4 52.4 54.1 62.1 57.9 54.2 59.9 60.3 5 - - - - - - - 6 59.5 61.3 63.3 59.4 62.1 65.8 74.3 7 - - - - - - - 8 47.5 49.3 46.8 54.4 48.6 48.0 57.8 9 - - - - - - - 10 70.6 73.0 73.2 65.5 66.1 75.6 81.5 11 73.7 77.2 79.7 76.6 79.0 86.2 89.3 12 52.7 52.7 59.1 60.9 51.2 59.9 70.4 13 78.9 81.6 74.1 76.2 81.8 89.9 94.1 14 59.5 60.7 70.0 68.6 69.1 70.4 73.6 15 65.3 66.6 69.2 68.4 66.2 71.6 75.1 16 50.0 53.9 60.6 53.5 54.3 55.1 60.0 17 55.3 61.9 67.5 64.9 59.0 67.5 74.2 18 67.8 69.6 72.7 72.3 68.3 72.5 71.3 MEAN 61.3 63.6 66.8 65.0 64.0 69.5 74.4 SEM 2.4 2.5 2.1 1.9 2.5 2.9 2.7  180   Table C.39. Heart rate (bpm) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 54.7 60.2 60.3 54.6 60.7 64.7 70.9 2 68.6 65.9 64.8 63.9 65.6 68.9 77.0 3 - - - - - - - 4 50.9 51.3 58.3 53.6 49.8 54.8 57.7 5 - - - - - - - 6 64.3 64.6 64.6 62.9 66.3 65.2 71.4 7 - - - - - - - 8 53.8 52.7 54.1 55.4 45.3 51.1 62.8 9 - - - - - - - 10 74.3 79.5 76.7 76.5 78.8 83.4 82.5 11 67.2 71.3 76.5 73.2 73.6 81.0 87.9 12 52.7 58.6 62.2 59.8 51.7 63.4 73.9 13 76.1 83.1 77.4 79.3 72.5 87.6 98.4 14 66.2 66.0 69.5 70.2 71.3 70.8 72.6 15 69.1 74.9 71.8 69.0 66.9 69.4 75.2 16 49.3 50.9 53.7 52.9 55.6 56.2 58.9 17 53.7 55.9 59.4 59.1 63.6 69.6 75.1 18 64.1 66.4 69.5 67.2 66.8 66.8 69.9 MEAN 61.8 64.4 65.6 64.1 63.5 68.1 73.9 SEM 2.3 2.6 2.0 2.2 2.5 2.7 2.7 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     181   Table C.40. Heart rate (bpm) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 58.7 73.4 65.4 64.6 71.7 80.8 84.1 2 61.5 76.9 70.1 69.0 70.6 69.4 79.4 3 59.4 76.8 75.4 69.0 76.5 83.4 92.3 4 49.8 64.7 63.4 58.7 63.3 62.5 63.5 5 - - - - - - - 6 66.0 76.8 71.7 68.9 71.2 80.5 87.4 7 - - - - - - - 8 48.4 70.0 61.0 54.5 60.6 60.1 64.8 9 - - - - - - - 10 73.5 94.9 80.8 91.2 94.3 97.1 102.5 11 79.2 95.6 99.9 90.9 89.4 91.4 99.7 12 56.2 76.8 72.6 63.9 67.2 75.3 85.5 13 - - - - - - - 14 66.2 71.1 69.2 66.7 70.2 72.1 77.5 15 69.0 82.3 77.3 78.4 75.2 78.7 86.5 16 48.1 56.0 61.5 58.6 57.0 61.6 63.2 17 58.0 71.2 76.9 73.4 72.3 84.0 84.1 18 65.9 76.3 79.1 78.9 78.7 82.4 97.1 MEAN 61.4 75.9 73.2 70.5 72.7 77.1 83.4 SEM 2.3 2.6 2.5 2.8 2.6 2.8 3.2  182   Table C.41. Heart rate (bpm) raw data for each participant during the HX2 CO2 reactivity protocol.  Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 58.7 85.2 78.2 74.0 75.8 86.2 91.2 2 61.5 71.7 71.0 74.5 73.7 78.1 84.8 3 59.4 76.0 68.9 71.8 71.7 78.7 87.4 4 49.8 64.2 56.6 62.0 60.8 63.1 68.0 5 - - - - - - - 6 66.0 74.8 74.1 74.4 72.2 82.4 89.3 7 - - - - - - - 8 48.4 63.2 50.1 53.4 54.1 61.1 69.6 9 - - - - - - - 10 73.5 100.0 82.2 91.2 97.2 96.2 104.0 11 79.2 95.0 92.0 88.1 89.5 93.6 99.9 12 56.2 77.4 74.2 68.4 68.9 74.3 80.5 13 - - - - - - - 14 66.2 70.1 66.9 61.3 66.9 73.3 85.6 15 69.0 82.9 80.1 77.8 79.2 83.5 85.4 16 48.1 56.6 61.7 58.1 61.1 62.5 64.4 17 58.0 75.3 69.7 71.9 74.1 78.2 85.0 18 65.9 74.1 72.2 70.8 74.0 79.1 87.0 MEAN 61.4 76.2 71.3 71.3 72.8 77.9 84.4 SEM 2.3 3.0 2.7 2.7 2.8 2.7 2.8  183   Table C.42. Mean arterial pressure (mmHg) raw data for each participant. NX1 CO2 reactivity protocolDefinition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 84.6 88.1 87.5 87.7 91.4 96.2 102.9 2 80.0 80.4 78.4 80.5 84.7 90.4 93.8 3 - - - - - - - 4 80.8 80.8 82.1 86.1 86.0 91.0 93.4 5 - - - - - - - 6 81.9 89.6 85.4 81.1 84.3 90.2 95.8 7 - - - - - - - 8 98.0 94.1 92.5 94.5 95.9 105.6 106.8 9 - - - - - - - 10 82.3 80.7 80.7 82.7 81.6 83.0 88.2 11 88.4 89.4 89.9 89.7 89.3 89.4 92.5 12 82.4 82.4 75.9 80.4 78.8 86.0 89.1 13 85.6 88.4 86.3 90.7 89.1 104.0 105.9 14 73.9 74.1 83.2 78.9 80.2 77.8 85.9 15 90.3 88.3 94.3 93.0 94.9 98.7 103.5 16 68.3 82.0 84.4 83.6 85.5 87.4 85.6 17 79.8 75.4 75.2 77.0 82.2 83.0 85.0 18 97.5 98.8 96.4 98.1 99.1 101.9 105.0 MEAN 83.8 85.2 85.2 86.0 87.3 91.7 95.2 SEM 2.0 1.7 1.6 1.6 1.5 2.1 2.0  184   Table C.43. Mean arterial pressure (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 87.4 88.2 86.9 87.4 87.5 92.0 95.2 2 83.2 84.4 81.3 82.5 84.2 84.8 87.1 3 - - - - - - - 4 86.6 84.9 87.7 90.3 86.5 92.4 95.4 5 - - - - - - - 6 76.8 81.2 80.1 79.7 82.5 87.6 91.4 7 - - - - - - - 8 85.8 91.0 84.9 87.4 89.9 94.5 98.6 9 - - - - - - - 10 85.5 85.8 85.4 87.3 90.2 92.4 92.6 11 89.6 88.4 86.3 86.8 89.7 89.5 95.5 12 75.1 76.5 75.8 78.1 75.8 80.8 87.8 13 83.0 95.0 93.8 95.6 92.9 102.3 110.8 14 85.9 88.5 92.6 89.1 88.3 90.2 96.6 15 89.4 92.8 91.8 93.7 92.2 96.5 100.1 16 79.1 85.0 80.7 82.1 84.4 86.7 87.5 17 83.9 79.8 78.8 82.5 83.4 87.6 92.2 18 94.8 92.1 94.5 96.4 97.4 99.4 100.4 MEAN 84.7 86.7 85.8 87.1 87.5 91.2 95.1 SEM 1.3 1.3 1.5 1.4 1.3 1.4 1.6  185   Table C.44. Mean arterial pressure (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 91.5 96.1 93.7 94.2 98.7 102.1 106.4 2 84.9 85.9 81.3 82.9 84.8 89.9 100.1 3 94.4 103.7 100.6 98.9 100.0 105.6 112.3 4 82.2 85.8 82.8 82.6 85.6 90.0 93.0 5 - - - - - - - 6 83.0 91.3 80.0 88.4 91.4 94.1 104.9 7 - - - - - - - 8 92.0 101.9 95.5 100.8 103.6 103.7 107.9 9 - - - - - - - 10 83.7 87.8 84.5 90.8 91.6 91.5 94.3 11 89.3 94.8 89.3 89.4 89.3 92.6 97.4 12 75.1 78.9 74.3 73.8 78.2 81.9 86.6 13 - - - - - - - 14 85.9 90.0 88.9 90.4 95.9 102.3 109.8 15 97.5 100.6 93.3 98.9 102.8 106.0 113.2 16 84.2 89.1 83.2 86.5 90.6 88.6 90.4 17 75.4 79.8 80.1 87.5 86.2 89.4 95.7 18 90.9 104.7 100.5 101.2 102.9 104.2 113.5 MEAN 86.4 92.2 87.7 90.4 93.0 95.8 101.8 SEM 1.7 2.1 2.0 2.0 2.0 2.0 2.3  186   Table C.45. Mean arterial pressure (mmHg) raw data for each participant during the HX2 CO2 reactivity test. SUBJECT AB AB-DEF -8 -4 0 4 8 1 91.5 105.4 99.6 100.1 101.9 104.5 110.0 2 84.9 83.5 86.7 90.3 89.1 93.6 108.6 3 94.4 100.3 95.5 98.4 101.7 106.0 111.6 4 82.2 85.6 80.7 85.0 88.0 91.2 96.6 5 - - - - - - - 6 83.0 86.5 80.1 87.7 83.0 95.3 105.6 7 - - - - - - - 8 92.0 100.5 97.4 96.7 98.4 106.7 108.2 9 - - - - - - - 10 83.7 88.7 89.2 87.5 86.3 92.9 97.6 11 89.3 91.6 90.7 90.5 90.6 94.2 100.4 12 75.1 79.7 76.5 75.4 78.7 81.0 85.5 13 - - - - - - - 14 85.9 103.2 103.9 98.8 102.5 109.5 118.4 15 97.5 103.8 102.3 105.9 107.4 111.9 115.8 16 84.2 85.5 79.4 84.1 84.3 83.3 89.7 17 75.4 79.4 76.4 79.9 77.6 81.8 89.6 18 90.9 92.8 92.8 97.1 98.3 103.6 109.8 MEAN 86.4 91.9 89.4 91.2 92.0 96.8 103.4 SEM 1.7 2.3 2.4 2.2 2.4 2.6 2.6 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     187   Table C.46. Middle cerebral artery velocity (cm/s) raw data for each participant NX1 CO2 reactivity Protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 47.7 51.1 42.3 48.3 53.5 55.0 71.8 2 71.9 75.0 60.6 71.1 78.7 88.6 105.9 3 - - - - - - - 4 55.8 59.8 49.7 54.3 69.0 55.1 78.3 5 - - - - - - - 6 66.8 71.7 54.0 60.0 76.7 80.9 92.1 7 - - - - - - - 8 54.4 57.0 45.4 49.7 54.6 64.0 80.7 9 - - - - - - - 10 48.2 52.0 42.4 46.8 51.4 55.1 64.6 11 67.1 71.9 55.6 63.2 73.3 83.1 98.9 12 67.1 70.2 51.0 60.6 70.6 79.1 87.3 13 61.4 65.9 49.5 55.5 62.6 77.1 83.9 14 57.2 56.0 47.9 50.4 57.1 64.1 70.4 15 62.7 63.9 52.9 58.2 63.8 72.8 84.2 16 80.8 86.2 67.2 75.4 86.0 100.0 106.0 17 70.9 79.1 57.0 65.4 75.1 86.5 97.2 18 43.1 42.7 34.6 40.2 46.0 50.4 56.3 MEAN 61.1 64.5 50.7 57.1 65.6 72.3 84.1 SEM 2.7 3.1 2.1 2.5 3.0 3.9 3.8 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     188   Table C.47. Middle cerebral artery velocity (cm/s) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 54.0 66.6 46.5 54.5 59.6 67.1 81.5 2 67.4 77.0 58.6 68.0 76.6 84.6 100.9 3 - - - - - - - 4 62.1 64.5 53.2 58.0 64.6 71.9 83.0 5 - - - - - - - 6 61.2 66.9 53.1 56.4 74.6 80.4 92.6 7 - - - - - - - 8 58.9 58.3 47.1 52.3 58.6 66.2 83.4 9 - - - - - - - 10 50.2 53.3 44.1 47.3 56.1 59.3 65.7 11 81.2 86.9 66.8 77.0 86.9 99.4 110.2 12 66.4 73.2 55.2 63.5 72.8 82.5 89.2 13 62.3 66.5 48.7 56.6 65.9 82.4 92.9 14 56.6 56.0 48.5 52.7 57.3 65.0 72.0 15 61.1 65.9 48.4 53.9 60.7 68.7 80.7 16 83.0 85.1 63.1 72.4 85.5 98.9 107.7 17 70.1 75.4 57.9 67.5 80.1 92.0 101.0 18 39.7 41.1 33.0 37.8 42.9 46.0 55.5 MEAN 62.4 66.9 51.7 58.4 67.3 76.0 86.9 SEM 2.9 3.1 2.1 2.6 3.2 3.9 4.0 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     189   Table C.48. Middle cerebral artery velocity (cm/s) raw data for each participant during the HX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 54.6 62.1 52.9 58.9 71.3 73.5 87.2 2 66.8 77.5 63.4 72.2 77.3 89.7 112.4 3 57.1 66.8 53.8 58.2 65.9 80.0 94.7 4 57.4 69.3 52.9 62.4 70.6 80.9 87.7 5 - - - - - - - 6 62.5 80.3 58.2 61.7 77.2 89.7 108.1 7 - - - - - - - 8 50.3 57.4 44.3 54.5 60.3 87.3 101.6 9 - - - - - - - 10 50.0 54.2 46.1 54.8 58.3 63.0 66.6 11 70.6 71.9 49.0 60.8 73.8 95.6 106.7 12 72.3 80.2 61.0 70.8 80.5 90.0 99.4 13 - - - - - - - 14 56.6 56.3 45.8 48.8 55.0 66.3 72.6 15 64.0 68.6 57.1 62.9 68.5 79.0 90.0 16 88.3 82.5 64.2 76.2 86.8 100.2 116.0 17 70.6 76.8 59.1 70.3 80.6 91.8 109.7 18 43.1 48.4 37.4 42.5 47.9 58.3 70.0 MEAN 61.7 68.0 53.2 61.1 69.6 81.8 94.5 SEM 2.9 2.8 2.0 2.4 2.8 3.2 4.1 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     190   Table C.49. Middle cerebral artery velocity (cm/s) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 54.6 71.3 45.7 60.5 69.8 79.0 90.0 2 66.8 67.7 50.3 56.9 70.0 88.4 121.0 3 57.1 64.5 48.9 58.8 62.7 77.0 89.2 4 57.4 69.9 55.5 65.5 77.3 83.9 95.3 5 - - - - - - - 6 62.5 73.8 56.6 67.1 79.1 90.8 104.4 7 - - - - - - - 8 50.3 53.6 44.3 49.8 56.1 64.2 80.7 9 - - - - - - - 10 50.0 61.1 48.1 57.5 58.4 61.3 68.8 11 70.6 81.6 63.3 69.8 73.0 96.2 106.6 12 72.3 79.7 54.6 72.3 76.6 83.9 98.3 13 - - - - - - - 14 56.6 59.6 48.0 56.0 60.5 71.0 76.4 15 64.0 72.6 54.5 63.9 71.7 77.0 95.6 16 88.3 91.9 71.0 79.1 91.3 99.4 120.2 17 70.6 76.6 58.5 67.8 79.7 89.4 107.9 18 43.1 46.6 36.1 42.5 48.5 59.6 69.1 MEAN 61.7 69.3 52.5 62.0 69.6 80.1 94.5 SEM 2.9 3.0 2.2 2.4 2.9 3.2 4.3 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     191   Table C.50. Arterial pH raw data for each participant.NX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 7.41 7.40 7.46 7.42 7.41 7.38 7.35 2 7.39 7.38 7.44 7.41 7.39 7.37 7.34 3 - - - - - - - 4 7.43 7.43 7.50 7.46 7.42 7.41 7.38 5 - - - - - - - 6 7.41 7.41 7.51 7.46 7.40 7.38 7.36 7 - - - - - - - 8 7.43 7.43 7.48 7.46 7.42 7.39 7.38 9 - - - - - - - 10 7.43 7.41 7.47 7.45 7.41 7.39 7.37 11 7.41 7.39 7.47 7.43 7.39 7.36 7.33 12 7.43 7.42 7.51 7.46 7.41 7.39 7.36 13 7.42 7.41 7.48 7.45 7.41 7.38 7.35 14 7.43 7.45 7.51 7.47 7.43 7.41 7.38 15 7.41 7.42 7.48 7.45 7.41 7.40 7.37 16 7.43 7.44 7.48 7.45 7.43 7.40 7.37 17 7.44 7.43 7.50 7.47 7.42 7.40 7.37 18 7.46 7.46 7.53 7.50 7.45 7.42 7.40 MEAN 7.42 7.42 7.49 7.45 7.41 7.39 7.37 SEM 0.0 0.0 0.01 0.01 0.0 0.0 0.01 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     192   Table C.51. Arterial pH raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 7.42 7.40 7.45 7.42 7.40 7.37 7.35 2 7.41 7.39 7.45 7.42 7.39 7.37 7.34 3 - - - - - - - 4 7.44 7.43 7.47 7.46 7.42 7.39 7.38 5 - - - - - - - 6 7.40 7.41 7.50 7.46 7.40 7.38 7.36 7 - - - - - - - 8 7.42 7.42 7.49 7.46 7.42 7.40 7.39 9 - - - - - - - 10 7.43 7.41 7.48 7.44 7.41 7.38 7.35 11 7.41 7.39 7.48 7.41 7.38 7.35 7.32 12 7.42 7.41 7.51 7.47 7.42 7.39 7.37 13 7.41 7.40 7.47 7.43 7.41 7.37 7.35 14 7.43 7.44 7.52 7.47 7.44 7.40 7.39 15 7.42 7.42 7.50 7.46 7.42 7.40 7.38 16 7.41 7.41 7.49 7.46 7.42 7.40 7.38 17 7.43 7.42 7.50 7.46 7.42 7.40 7.37 18 7.46 7.45 7.52 7.49 7.45 7.42 7.39 MEAN 7.42 7.41 7.49 7.45 7.41 7.39 7.37 SEM 0.00 0.00 0.01 0.01 0.00 0.00 0.01 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     193   Table C.52. Arterial pH raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 7.42 7.42 7.47 7.44 7.41 7.38 7.36 2 7.41 7.41 7.46 7.44 7.42 7.39 7.36 3 7.42 7.42 7.52 7.47 7.43 7.40 7.38 4 7.42 7.43 7.50 7.47 7.44 7.41 7.38 5 - - - - - - - 6 7.41 7.42 7.50 7.47 7.43 7.40 7.37 7 - - - - - - - 8 7.44 7.45 7.51 7.48 7.44 7.42 7.39 9 - - - - - - - 10 7.42 7.42 7.48 7.44 7.42 7.39 7.37 11 7.41 7.41 7.47 7.44 7.41 7.38 7.34 12 7.43 7.43 7.50 7.46 7.42 7.40 7.37 13 - - - - - - - 14 7.43 7.43 7.51 7.47 7.44 7.41 7.37 15 7.42 7.43 7.48 7.45 7.43 7.40 7.37 16 7.42 7.43 7.52 7.46 7.44 7.41 7.39 17 7.44 7.44 7.51 7.47 7.44 7.40 7.37 18 7.45 7.44 7.50 7.47 7.44 7.41 7.38 MEAN 7.42 7.42 7.50 7.46 7.43 7.40 7.37 SEM 0.00 0.00 0.00 0.00 0.00 0.00 0.00  194   Table C.53. Arterial pH raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 7.42 7.41 7.49 7.44 7.41 7.38 7.35 2 7.41 7.40 7.46 7.43 7.41 7.38 7.35 3 7.42 7.43 7.43 7.46 7.43 7.40 7.37 4 7.42 7.43 7.50 7.47 7.43 7.40 7.37 5 - - - - - - - 6 7.41 7.43 7.52 7.47 7.43 7.40 7.37 7 - - - - - - - 8 7.44 7.45 7.50 7.50 7.44 7.41 7.39 9 - - - - - - - 10 7.42 7.41 7.47 7.44 7.42 7.39 7.36 11 7.41 7.40 7.48 7.44 7.40 7.37 7.34 12 7.43 7.41 7.49 7.45 7.41 7.38 7.35 13 - - - - - - - 14 7.43 7.43 7.50 7.46 7.43 7.39 7.36 15 7.42 7.42 7.47 7.43 7.40 7.38 7.35 16 7.42 7.44 7.50 7.47 7.43 7.41 7.39 17 7.44 7.44 7.51 7.47 7.44 7.41 7.37 18 7.45 7.45 7.52 7.48 7.44 7.41 7.37 MEAN 7.42 7.42 7.49 7.46 7.42 7.39 7.36 SEM 0.00 0.00 0.01 0.01 0.00 0.00 0.00 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     195   Table C.54. Partial pressure of arterial carbon dioxide (mmHg) raw data for each participant.NX1 CO2 reactivity protocol.Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 43.4 44.0 37.8 42.0 44.3 47.9 51.7 2 44.3 45.6 38.3 42.1 45.6 48.6 53.0 3 - - - - - - - 4 42.9 43.1 35.1 39.6 44.9 46.6 50.3 5 - - - - - - - 6 40.0 40.5 29.6 34.7 41.8 43.7 46.8 7 - - - - - - - 8 41.1 41.3 35.4 37.5 42.1 46.6 48.7 9 - - - - - - - 10 42.9 43.7 37.3 39.4 44.1 46.5 49.6 11 37.0 39.1 31.0 34.4 38.7 42.6 45.9 12 36.6 39.2 29.9 34.0 39.6 42.1 45.8 13 37.2 38.2 31.2 34.3 38.9 42.5 46.0 14 40.9 38.1 31.5 34.8 39.8 42.5 46.3 15 38.0 38.0 31.0 34.7 38.6 41.2 45.0 16 42.8 41.2 36.4 40.4 42.9 45.9 50.8 17 39.2 41.4 32.9 36.2 41.7 44.3 48.3 18 36.1 35.6 28.1 31.1 36.5 39.3 42.3 MEAN 40.2 40.6 33.3 36.8 41.4 44.3 47.9 SEM 0.7 0.7 0.8 0.8 0.7 0.7 0.7  196   Table C.55. Partial pressure of arterial carbon dioxide (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 42.3 44.9 38.5 42.5 45.1 48.8 52.2 2 43.4 46.1 38.4 42.8 46.6 49.5 54.2 3 - - - - - - - 4 42.1 44.1 38.0 40.2 44.3 47.9 50.3 5 - - - - - - - 6 41.4 40.7 30.2 34.4 41.7 43.9 46.7 7 - - - - - - - 8 43.0 42.2 34.1 38.0 42.9 44.9 47.2 9 - - - - - - - 10 42.4 43.7 35.1 39.7 44.5 48.1 51.4 11 37.6 39.8 30.2 36.2 39.7 43.3 47.5 12 37.7 39.8 29.6 33.9 38.9 43.0 46.2 13 37.1 38.7 30.9 35.8 37.9 42.7 45.6 14 40.0 39.2 30.0 35.6 38.4 43.9 44.4 15 38.4 39.0 30.1 34.8 39.1 41.4 43.8 16 44.1 43.5 34.6 38.7 43.7 46.0 49.5 17 39.5 38.7 32.6 36.8 41.4 44.9 48.5 18 35.5 36.4 28.2 31.9 36.6 39.4 43.1 MEAN 40.3 41.2 32.9 37.2 41.5 44.8 47.9 SEM 0.7 0.7 0.9 0.8 0.8 0.7 0.8  197   Table C.56. Partial pressure of arterial carbon dioxide (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    SUBJECT AB AB-DEF -8 -4 0 4 8 1 42.2 42.9 37.5 39.9 43.7 47.8 51.5 2 43.5 44.0 37.8 41.0 43.7 46.6 51.3 3 38.0 38.3 27.6 33.8 38.3 41.4 45.1 4 43.7 43.7 34.8 38.7 42.5 46.6 50.4 5 - - - - - - - 6 39.3 38.2 30.1 33.2 38.0 41.4 45.6 7 - - - - - - - 8 38.8 39.4 32.2 35.1 39.7 41.7 46.1 9 - - - - - - - 10 42.3 42.7 34.1 39.5 43.0 46.3 48.9 11 36.8 37.3 31.0 34.7 37.7 41.3 45.0 12 38.2 38.4 30.5 34.7 38.2 41.7 45.1 13 - - - - - - - 14 40.0 38.7 30.9 34.6 39.2 42.8 47.5 15 38.0 37.4 31.3 35.4 38.2 40.6 43.8 16 43.1 42.6 32.5 39.0 41.7 45.7 49.0 17 39.5 38.8 31.0 35.5 39.5 43.2 47.3 18 36.5 37.1 29.9 33.8 37.3 40.7 45.5 MEAN 40.0 40.0 32.2 36.3 40.0 43.4 47.3 SEM 0.6 0.6 0.7 0.6 0.6 0.6 0.6  198   Table C.57. Partial pressure of arterial carbon dioxide (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 42.2 44.0 34.5 40.0 44.5 47.7 52.3 2 43.5 44.7 36.9 40.9 44.0 48.7 52.5 3 38.0 38.4 29.2 34.7 38.2 42.0 45.8 4 43.7 44.1 35.0 38.8 43.1 46.7 51.1 5 - - - - - - - 6 39.3 37.5 28.7 33.4 37.4 41.7 45.2 7 - - - - - - - 8 38.8 39.6 33.1 33.0 40.9 43.8 46.7 9 - - - - - - - 10 42.3 44.2 36.1 40.7 43.0 46.0 50.7 11 36.8 38.2 29.5 34.6 38.3 42.5 46.0 12 38.2 39.3 30.9 35.6 39.2 43.2 47.0 13 - - - - - - - 14 40.0 40.3 32.6 35.1 40.0 44.7 49.1 15 38.0 37.4 31.4 35.8 39.0 41.4 45.3 16 43.1 41.2 34.1 38.2 42.5 45.3 48.7 17 39.5 39.3 31.5 35.7 39.8 43.1 47.3 18 36.5 36.7 28.5 33.2 36.9 41.6 46.2 MEAN 40.0 40.3 32.3 36.4 40.5 44.2 48.1 SEM 0.6 0.7 0.7 0.7 0.6 0.6 0.6 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system     199   Table C.58. Partial pressure of arterial oxygen (mmHg) raw data for each participant. NX1 CO2 reactivity protocol.SUBJECT AB AB-DEF -8 -4 0 4 8 1 82.6 83.3 80.1 80.3 83.8 86.3 89.3 2 86.0 88.1 93.2 82.2 83.0 87.4 85.6 3 - - - - - - - 4 87.2 87.8 85.2 91.3 89.8 90.5 91.4 5 - - - - - - - 6 86.5 82.9 92.4 87.7 81.6 90.6 88.3 7 - - - - - - - 8 84.2 92.5 86.9 92.5 95.8 83.9 90.3 9 - - - - - - - 10 85.1 84.1 81.3 88.4 81.8 89.1 88.6 11 97.7 98.4 98.7 98.1 104.1 100.8 102.3 12 78.9 87.9 88.1 86.7 80.5 89.5 88.1 13 98.2 93.7 93.6 99.1 95.2 94.7 95.1 14 87.2 91.9 91.2 92.2 89.9 92.3 94.1 15 93.9 91.1 91.1 89.0 93.0 94.2 92.9 16 93.1 98.5 92.4 85.4 93.6 94.3 93.9 17 91.3 90.8 90.3 88.6 86.2 90.1 92.9 18 86.5 99.6 96.2 95.6 83.9 96.4 98.1 MEAN 88.5 90.8 90.1 89.8 88.7 91.4 92.2 SEM 1.4 1.4 1.3 1.4 1.7 1.1 1.1 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    200   Table C.59. Partial pressure of arterial oxygen (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 89.7 84.0 82.9 86.8 82.4 86.3 88.7 2 91.8 85.7 83.3 87.8 88.4 89.7 88.0 3 - - - - - - - 4 88.0 86.0 90.8 87.2 85.9 86.6 94.9 5 - - - - - - - 6 83.9 84.3 87.8 89.7 85.8 84.9 90.4 7 - - - - - - - 8 81.8 89.8 86.3 87.6 88.0 86.3 84.0 9 - - - - - - - 10 85.1 85.3 92.2 87.8 85.0 91.6 90.6 11 95.1 99.4 102.1 97.8 99.6 102.5 100.3 12 84.5 84.6 84.5 86.0 86.6 85.4 87.4 13 95.5 92.4 96.6 97.5 95.7 93.3 104.7 14 85.4 89.9 91.2 88.6 92.1 87.0 96.4 15 91.6 85.1 101.8 86.0 89.2 92.9 95.5 16 88.7 91.1 89.5 92.5 79.8 90.6 93.3 17 85.5 85.7 88.8 93.1 91.6 92.1 91.8 18 90.4 93.0 98.3 95.8 93.1 96.2 99.6 MEAN 88.3 88.3 91.1 90.3 88.8 90.4 93.3 SEM 1.0 1.1 1.6 1.0 1.3 1.2 1.4 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    201   Table C.60. Partial pressure of arterial oxygen (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 86.4 46.8 42.3 44.4 45.7 46.6 47.7 2 90.9 45.4 41.9 44.4 47.3 47.8 47.4 3 89.7 43.6 50.0 44.8 39.8 43.7 41.6 4 83.3 40.3 44.7 44.4 42.1 42.7 44.4 5 - - - - - - - 6 86.7 42.3 44.0 45.1 41.7 43.9 43.1 7 - - - - - - - 8 77.7 41.5 41.0 42.9 39.4 43.7 43.3 9 - - - - - - - 10 89.1 44.5 46.6 43.3 43.6 45.8 47.3 11 97.3 44.7 43.9 42.9 44.1 44.3 45.1 12 85.0 43.2 43.0 44.3 43.3 43.7 43.5 13 - - - - - - - 14 85.4 45.6 45.8 46.4 44.6 44.8 43.3 15 88.4 44.2 44.8 44.0 45.5 43.7 43.1 16 89.4 45.1 48.3 43.6 48.0 46.4 44.4 17 88.1 43.6 46.2 45.0 45.8 46.7 47.0 18 86.2 46.3 46.1 45.4 45.2 46.0 46.2 MEAN 87.4 44.1 44.9 44.3 44.0 45.0 44.8 SEM 1.1 0.4 0.6 0.2 0.6 0.3 0.5  202   Table C.61. Partial pressure of arterial oxygen (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 86.4 48.5 47.8 45.8 46.0 46.4 49.4 2 90.9 46.4 45.2 43.8 46.6 45.9 45.9 3 89.7 42.7 42.7 39.9 42.9 42.5 45.0 4 83.3 43.1 44.4 41.1 40.6 41.9 42.3 5 - - - - - - - 6 86.7 43.3 45.1 41.2 43.1 41.9 43.3 7 - - - - - - - 8 77.7 40.7 40.6 48.5 41.0 41.8 45.8 9 - - - - - - - 10 89.1 46.9 47.9 42.6 43.8 46.3 47.7 11 97.3 44.3 44.7 41.8 44.5 45.4 44.6 12 85.0 43.2 44.1 43.1 42.8 43.4 43.8 13 - - - - - - - 14 85.4 44.3 45.0 47.6 46.8 43.4 44.3 15 88.4 45.1 45.0 41.3 43.1 42.0 43.2 16 89.4 48.4 49.8 46.3 47.1 47.8 46.7 17 88.1 46.0 43.3 44.9 45.9 45.9 46.9 18 86.2 47.3 48.6 46.5 47.6 47.1 47.0 MEAN 87.4 45.0 45.3 43.9 44.4 44.4 45.4 SEM 1.1 0.5 0.6 0.7 0.5 0.5 0.5 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    203   Table C.62. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant NX1 CO2 reactivity protocol.SUBJECT AB AB-DEF -8 -4 0 4 8 1 -0.0 -0.2 0.0 0.7 -0.7 -0.9 -1.3 2 0.8 0.4 1.7 0.7 0.5 -0.4 -0.1 3 - - - - - - - 4 1.2 0.3 0.7 1.2 2.3 0.1 -0.4 5 - - - - - - - 6 2.2 0.6 -1.2 -1.4 1.3 -0.3 -0.7 7 - - - - - - - 8 0.7 -0.7 1.6 -0.6 -0.2 0.5 -1.3 9 - - - - - - - 10 -0.8 -0.4 1.5 -0.8 -0.2 -1.8 -2.5 11 0.2 0.1 0.0 -0.4 -0.3 -0.4 -1.2 12 -1.5 -0.2 -0.0 -1.5 0.1 -1.0 -1.3 13 -0.4 -0.6 0.2 -0.5 0.6 -0.4 -1.0 14 2.0 -0.6 0.4 -0.5 -0.5 -1.3 -1.5 15 0.6 0.0 0.8 0.6 0.6 -1.0 -1.1 16 2.3 -1.3 1.1 1.6 0.4 -0.5 -0.0 17 1.2 -0.1 0.7 -0.5 0.9 -0.3 -0.2 18 1.0 -1.6 -0.6 -1.5 -0.8 -1.7 -2.4 MEAN 0.6 -0.3 0.5 -0.2 0.3 -0.7 -1.1 SEM 0.2 0.1 0.2 0.2 0.2 0.1 0.1 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    204   Table C.63. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 1.3 -0.6 1.7 1.6 0.7 -0.3 -1.0 2 0.4 -0.0 1.1 1.2 0.4 -0.5 -0.2 3 - - - - - - - 4 1.5 1.1 3.4 1.4 1.2 0.7 -1.5 5 - - - - - - - 6 2.8 1.6 -1.4 -0.4 1.3 0.1 -0.8 7 - - - - - - - 8 1.6 0.1 0.4 -0.1 1.1 -1.3 -4.0 9 - - - - - - - 10 0.8 -1.4 -1.8 -1.1 -0.9 -1.7 -2.3 11 -0.7 -0.4 -1.6 0.2 -0.5 -1.3 -1.0 12 0.0 -0.2 -0.8 -1.2 -0.6 -1.1 -1.6 13 -1.2 -1.4 -0.4 -0.0 -1.7 -1.1 -2.9 14 1.7 -0.2 -0.6 0.2 -1.0 -0.1 -3.2 15 1.8 0.3 0.2 0.7 0.9 -1.2 -2.8 16 2.1 0.7 0.7 0.1 0.8 -0.7 -1.3 17 -0.7 -2.6 0.2 -0.0 -0.3 -0.6 -1.0 18 -0.2 -0.9 -0.2 -1.6 -1.1 -1.4 -2.8 MEAN 0.8 -0.2 0.0 0.0 0.0 -0.7 -1.9 SEM 0.3 0.2 0.3 0.2 0.2 0.1 0.2 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    205   Table C.64. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the HX1 protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.7 -0.1 2.4 0.9 0.7 0.7 0.4 2 1.1 0.5 2.5 1.6 -0.1 -0.6 -0.5 3 0.7 -0.1 -2.0 -0.3 0.2 -0.6 -1.1 4 2.7 1.1 0.3 0.0 -0.1 -0.1 -0.3 5 - - - - - - - 6 3.1 0.9 1.0 0.4 1.0 0.5 0.6 7 - - - - - - - 8 -0.2 -1.9 0.5 -1.6 -1.2 -3.1 -2.5 9 - - - - - - - 10 -0.2 -0.0 -0.8 0.1 -0.1 -0.5 -2.0 11 0.0 -0.3 1.4 1.0 0.3 -0.4 -0.6 12 -0.4 -0.8 0.3 -0.5 -1.0 -1.5 -1.9 13 - - - - - - - 14 1.7 -1.0 -0.2 -1.1 -0.6 -0.8 -0.0 15 0.3 -1.7 0.3 0.1 -0.9 -2.6 -3.4 16 1.9 1.6 -1.1 1.1 -0.5 0.1 -0.7 17 1.5 -0.1 0.4 0.4 0.4 -0.0 0.1 18 -0.4 -0.6 0.3 -0.0 -0.3 -0.9 -0.2 MEAN 0.9 -0.1 0.3 0.1 -0.1 -0.7 -0.8 SEM 0.3 0.2 0.3 0.2 0.1 0.2 0.3 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    206   Table C.65. End-tidal-to-arterial carbon dioxide gradient (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 0.7 0.7 1.3 1.0 1.4 0.6 1.3 2 1.1 1.0 1.6 1.6 0.2 0.3 1.2 3 0.7 -0.1 -0.3 -0.7 -0.4 -0.6 -1.2 4 2.7 1.0 0.2 0.4 -0.0 -0.0 -0.1 5 - - - - - - - 6 3.1 0.2 0.6 0.5 0.2 0.6 0.2 7 - - - - - - - 8 -0.2 -1.5 1.1 -3.0 -0.5 -1.8 -2.6 9 - - - - - - - 10 -0.2 -0.5 0.5 0.3 -0.3 -1.8 -1.5 11 0.0 -0.2 -0.4 0.3 -0.1 -0.3 -0.8 12 -0.4 -1.3 0.4 -0.5 -0.6 -0.8 -1.2 13 - - - - - - - 14 1.7 -0.5 0.5 -1.9 -0.6 0.1 0.2 15 0.3 -1.7 0.7 0.9 -0.1 -2.3 -2.3 16 1.9 -0.9 0.7 0.5 0.9 0.5 -1.3 17 1.5 0.0 1.1 0.5 0.4 -0.0 -0.4 18 -0.4 -1.8 -0.8 -1.0 -1.9 -1.1 -0.5 MEAN 0.9 -0.4 0.5 -0.0 -0.1 -0.4 -0.6 SEM 0.3 0.2 0.1 0.3 0.2 0.2 0.3  207   Table C.66. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant NX1 CO2 reactivity protocol.SUBJECT AB AB-DEF -8 -4 0 4 8 1 6.4 6.8 8.4 8.7 5.9 4.4 -0.1 2 6.7 -0.4 -3.4 4.8 7.1 2.0 3.7 3 - - - - - - - 4 5.9 4.5 8.4 2.2 4.3 3.4 2.5 5 - - - - - - - 6 9.1 8.8 -0.6 0.4 8.6 1.3 3.9 7 - - - - - - - 8 8.4 0.7 2.6 -1.2 -1.0 9.1 2.5 9 - - - - - - - 10 3.4 3.3 7.6 0.4 6.3 -0.6 0.3 11 4.7 5.1 4.2 4.8 -0.9 2.3 0.1 12 14.2 3.7 6.8 6.7 10.4 4.3 5.7 13 2.3 5.5 2.9 -1.1 4.7 3.3 2.0 14 9.6 8.3 3.8 2.6 3.5 3.1 1.6 15 5.3 3.2 8.0 5.7 2.9 2.0 3.1 16 5.4 -2.4 1.4 5.3 -0.0 -0.3 -0.0 17 6.9 1.4 2.7 3.8 7.5 3.0 1.0 18 9.5 0.5 3.6 4.7 6.4 3.2 2.1 MEAN 7.0 3.5 4.0 3.4 4.7 2.9 2.0 SEM 0.7 0.8 0.9 0.7 0.9 0.6 0.4 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    208   Table C.67. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 6.4 5.3 8.2 -4.4 5.8 2.5 1.2 2 0.4 2.8 6.2 -0.4 0.4 -0.6 1.3 3 - - - - - - - 4 4.1 8.2 3.7 6.8 5.2 6.3 -1.3 5 - - - - - - - 6 8.1 8.2 3.1 2.3 6.5 6.2 1.8 7 - - - - - - - 8 8.9 4.0 5.1 2.5 6.4 4.7 7.1 9 - - - - - - - 10 4.9 2.8 -3.7 1.9 3.3 -1.9 -1.1 11 3.3 3.2 1.0 5.4 0.0 0.4 2.5 12 13.3 8.6 10.2 7.9 4.9 7.4 6.0 13 2.7 5.6 2.3 -0.2 4.3 4.4 -6.1 14 7.8 2.7 2.3 4.9 2.6 5.6 -1.6 15 8.6 11.4 -4.9 9.6 7.5 4.1 1.8 16 5.0 1.7 3.8 0.4 12.7 4.2 0.3 17 7.2 7.8 3.8 1.0 1.9 0.8 1.0 18 10.4 6.3 2.4 3.0 4.9 5.9 0.7 MEAN 6.5 5.6 3.1 2.9 4.7 3.6 0.9 SEM 0.8 0.7 1.0 0.9 0.8 0.7 0.8 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    209   Table C.68. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the HX1 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 5.7 3.6 6.0 5.8 4.5 3.6 2.3 2 5.8 5.2 7.9 6.1 2.7 3.6 3.5 3 5.9 7.2 2.6 6.6 11.0 7.3 9.4 4 10.0 10.7 6.2 6.6 8.7 8.0 6.7 5 - - - - - - - 6 10.7 8.3 5.8 5.1 9.1 7.1 7.8 7 - - - - - - - 8 16.6 8.7 10.0 9.4 11.8 7.3 7.9 9 - - - - - - - 10 1.7 5.9 3.5 6.1 6.6 4.9 2.9 11 2.5 6.4 7.1 7.4 7.1 6.7 6.4 12 8.6 8.0 6.1 7.2 7.4 6.5 7.0 13 - - - - - - - 14 7.8 5.1 4.5 4.2 6.1 6.0 7.4 15 4.3 6.9 5.7 7.0 5.5 7.4 8.2 16 5.1 6.3 3.3 7.1 2.9 4.8 6.8 17 6.5 6.7 5.6 6.4 5.6 4.1 4.0 18 5.4 3.6 4.0 4.4 4.7 4.3 4.0 MEAN 6.9 6.6 5.6 6.4 6.7 5.8 6.0 SEM 0.9 0.5 0.5 0.3 0.7 0.4 0.5 Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    210   Table C.69. End-tidal-to-arterial oxygen gradient (mmHg) raw data for each participant during the HX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 5.7 1.5 3.1 3.9 4.1 3.6 0.7 2 5.8 4.4 5.3 6.2 4.1 5.0 4.9 3 5.9 8.4 8.9 10.1 8.1 8.5 6.0 4 10.0 7.2 6.5 9.6 10.2 8.8 8.4 5 - - - - - - - 6 10.7 6.9 6.4 8.5 7.7 8.6 7.6 7 - - - - - - - 8 16.6 9.4 11.7 4.8 10.2 8.7 5.2 9 - - - - - - - 10 1.7 2.8 5.0 5.5 6.2 4.0 2.6 11 2.5 6.2 6.4 8.7 6.4 5.6 6.4 12 8.6 6.8 7.6 6.5 8.2 7.2 6.7 13 - - - - - - - 14 7.8 6.2 5.2 2.3 4.2 7.1 7.2 15 4.3 5.6 6.4 10.3 8.0 9.4 8.4 16 5.1 3.2 1.5 5.0 4.4 3.4 4.8 17 6.5 4.9 7.1 6.0 5.1 4.9 3.7 18 5.4 2.8 2.4 3.3 2.5 2.9 3.2 MEAN 6.9 5.5 6.0 6.5 6.4 6.3 5.4 SEM 0.9 0.5 0.6 0.6 0.6 0.5 0.5 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    211   Table C.70. Internal carotid artery flow (ml/min) raw data for each participant NX1 CO2 reactivity protocol. Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 423.5 362.4 315.8 362.7 403.0 512.2 681.9 2 256.2 253.9 178.3 204.9 251.4 379.6 457.1 3 - - - - - - - 4 241.3 234.7 183.0 176.7 239.9 302.8 333.9 5 - - - - - - - 6 271.4 271.6 176.9 238.5 292.8 315.1 472.9 7 - - - - - - - 8 - - - - - - - 9 - - - - - - - 10 250.6 253.8 196.5 222.3 229.2 270 370.8 11 248.5 263.2 176.4 224.1 330.9 392.9 476.7 12 221.8 271.0 195.6 199.2 266.1 273.9 347.4 13 257.9 303.1 226.8 264.1 298.0 351.0 459.9 14 - - - - - - - 15 198.3 242.7 166.5 209.4 241.7 253.6 305.3 16 315.3 362.0 258.9 285.2 360.7 352.2 482.9 17 234.5 266.9 180.8 214.0 262.0 329.3 407.7 18 265.6 246.0 186.3 209.6 295.4 285.9 347.0 MEAN 265.4 277.6 203.5 234.2 289.2 334.9 428.6 SEM 14.6 11.0 11.1 12.7 13.5 18.2 26.0  212   Table C.71. Internal carotid artery flow (ml/min) raw data for each participant during the NX2 CO2 reactivity protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 371.4 442.9 275.3 384.9 396.3 566.0 657.7 2 255.4 252.6 205.7 230.0 292.4 351.9 447.2 3 - - - - - - - 4 277.5 280.7 200.2 188.4 241.5 294.3 378.4 5 - - - - - - - 6 288.2 276.5 168.9 219.0 324.9 350.8 444.9 7 - - - - - - - 8 - - - - - - - 9 - - - - - - - 10 267.6 268.0 192.8 217.3 260.8 315.7 383.6 11 264.9 274.8 195.2 253.7 289.2 383.1 459.3 12 272.2 272.6 204.1 244.2 295.0 307.8 408.3 13 281.8 297.2 208.4 236.1 276.9 303.1 480.5 14 - - - - - - - 15 239.7 207.6 150.6 198.9 237.0 273.1 310.9 16 319.8 241.1 235.2 268.6 320.6 396.5 506.7 17 259.0 251.4 168.1 185.3 243.1 308.4 410.7 18 236.2 225.8 153.9 232.6 273.5 288.1 323.9 MEAN 277.8 274.2 196.5 238.3 287.6 344.9 434.3 SEM 9.4 15.0 8.9 13.4 11.4 20.3 23.4 Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    213   Table C.72. Internal carotid artery flow (ml/min) raw data for each participant during the HX1 CO2 reactivity protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system   SUBJECT AB AB-DEF -8 -4 0 4 8 1 350.6 378.3 285.4 340.8 447.4 604.5 730.0 2 - - - - - - - 3 358.8 358.8 231.3 313.4 392.5 466.1 543.6 4 - - - - - - - 5 - - - - - - - 6 303.4 303.4 250.4 237.9 333.9 395.9 571.3 7 - - - - - - - 8 - - - - - - - 9 - - - - - - - 10 233.4 259.3 236.2 262.3 274.7 342.8 364.7 11 319.1 319.1 203.3 242.0 292.6 383.2 475.8 12 276.6 317.0 218.4 270.0 310.7 366.1 478.8 13 - - - - - - - 14 - - - - - - - 15 245.3 232.3 186.6 225.0 261.6 298.9 403.3 16 359.9 359.9 266.4 259.9 348.8 472.1 577.1 17 243.1 243.1 165.6 228.8 261.7 336.6 443.5 18 349.2 349.2 250.0 301.6 340.7 404.0 541.1 MEAN 303.9 312.0 229.4 268.2 326.5 407.0 512.9 SEM 12.9 13.1 9.3 9.8 15.1 22.3 26.4  214   Table C.73. Internal carotid artery flow (ml/min) raw data for each participant during the HX2 CO2 protocol. SUBJECT AB AB-DEF -8 -4 0 4 8 1 435.8 435.8 229.2 324.9 480.8 513.5 724.1 2 - - - - - - - 3 310.5 381.3 259.3 318.6 372.4 456.6 553.7 4 - - - - - - - 5 - - - - - - - 6 278.4 276.2 200.1 257.0 305.2 331.8 522.1 7 - - - - - - - 8 - - - - - - - 9 - - - - - - - 10 326.4 326.4 210.8 268.1 255.4 321.3 463.3 11 267.5 348.8 238.7 260.8 331.7 415.8 414.9 12 341.7 341.7 225.5 281.6 341.1 418.8 478.5 13 - - - - - - - 14 - - - - - - - 15 248.3 248.3 186.8 244.8 289.9 347.9 406.7 16 341.1 330.8 278.6 292.9 343.7 435.8 626.6 17 193.8 248.7 173.1 193.8 240.9 328.8 435.4 18 283.4 307.5 231.6 273.6 365.0 418.1 546.3 MEAN 302.7 324.6 223.4 271.6 332.6 398.8 517.2 SEM 16.5 14.8 8.1 9.5 17.2 16.2 25.4 Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; AB = airbreathing; AB-DEF = isocapnic euoxia controlled by the dynamic end-tidal forcing system    215  Table C.74. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2 raw data for each participant NX1 CO2 protocol.SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -2.9 8.6 -2.8 4.2 1.9 2 -3.3 10.0 -2.7 4.2 5.3 3 - - - - - 4 -2.9 4.6 -3.4 - 1.4 5 - - - - - 6 -4.1 8.5 -3.0 2.8 3.3 7 - - - -  8 - - -1.9 6.1 4.2 9 - - - - - 10 -1.6 7.9 -2.0 3.2 1.8 11 -5.8 5.4 -3.0 4.2 1.6 12 -2.6 3.9 -2.8 3.0 1.3 13 -3.2 6.1 -2.8 3.9 4.5 14 - - -1.7 3.1 1.5 15 -4.0 3.1 -2.2 3.8 2.3 16 -3.9 4.1 -3.0 2.7 2.0 17 -3.5 7.0 -2.8 3.7 3.9 18 -4.3 2.3 -2.8 3.0 1.7 MEAN -3.5 6.0 -2.6 3.7 2.6 SEM 0.2 0.6 0.1 0.2 0.3 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.    216  Table C.75. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2 raw data for each participant during the NX2 CO2 protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -4.0 7.4 -2.9 4.0 2.6 2 - - - - - 3 -3.2 6.4 -2.6 3.8 4.9 4 -2.0 6.4 -2.0 3.2 1.3 5 - - - - - 6 -5.5 5.2 -3.3 3.3 2.7 7 - - - - - 8 - - -2.4 4.5 3.4 9 - - - - - 10 -3.0 5.6 -2.5 2.0 1.1 11 -3.8 7.0 -2.7 3.2 2.2 12 -3.4 4.4 -2.6 2.7 2.1 13 -2.9 8.4 -3.0 4.6 5.5 14 - - -1.7 3.1 1.4 15 -4.4 3.6 -2.4 3.8 2.7 16 -2.9 7.2 -2.9 3.2 1.7 17 -3.3 8.8 -2.9 3.3 3.2 18 -4.6 2.2 -2.4 3.6 2.1 MEAN -3.6 6.0 -2.6 3.4 2.6 SEM 0.2 0.5 0.1 0.1 0.3  217  Table C.76. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2 raw data for each participant during the HX1 CO2 protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -4.4 7.7 -3.2 2.7 3.2 2 - - -2.0 5.6 7.4 3 -4.8 4.7 -2.1 5.3 4.6 4 - - -3.0 2.9 1.2 5 - - - - - 6 -3.2 8.9 -3.1 5.0 3.9 7 - - - - - 8 - - -2.8 8.7 4.1 9 - - - - - 10 -1.7 4.0 -2.5 1.7 1.7 11 -3.8 7.5 -4.2 5.3 2.5 12 -3.3 6.9 -2.6 3.0 1.8 13 - - - - - 14 - - -1.9 4.1 1.9 15 -3.5 6.5 -2.0 3.8 5.0 16 -2.8 8.5 -3.0 4.4 2.6 17 -4.3 8.5 -3.1 4.4 4.1 18 -3.2 7.2 -2.6 5.6 6.8 MEAN -3.5 7.1 -2.7 4.5 3.7 SEM 0.2 0.5 0.1 0.4 0.4  218  Table C.77. Internal carotid artery, middle cerebral artery, and HCVR to end-tidal CO2 raw data for each participant during the HX2 CO2 protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -5.2 6.3 -3.5 3.6 3.1 2 - - -3.3 9.4 10.2 3 -3.2 5.8 -2.4 5.0 4.3 4 - - -3.3 2.8 2.7 5 - - - - - 6 -3.7 9.1 -3.1 4.0 3.2 7 - - - - - 8 - - -2.1 5.4 4.1 9 - - - - - 10 -2.5 9.1 -2.4 1.9 1.5 11 -3.2 3.0 -1.5 5.5 2.0 12 -3.5 4.7 -3.1 3.3 1.2 13 - - - - - 14 - - -2.4 3.1 2.7 15 -4.1 4.7 -2.8 3.8 4.7 16 -2.2 9.7 -2.6 3.7 2.9 17 -3.0 9.6 -2.9 4.2 3.3 18 -3.8 6.3 -2.6 5.3 6.7 MEAN -3.4 6.8 -2.7 4.4 3.8 SEM 0.2 0.7 0.1 0.4 0.5  219  Table C.78. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2 raw data for each participant NX1 CO2 protocol.SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -3.2 9.3 -3.1 4.5 2.1 2 -3.9 10.8 -3.1 4.7 5.9 3 - - - - - 4 -2.4 6.6 -2.8 - 2.0 5 - - - - - 6 -3.2 12.6 -2.4 4.0 4.7 7 - - - - - 8 - - -2.4 6.6 4.0 9 - - - - - 10 -1.8 11.5 -2.4 4.7 2.6 11 -6.1 6.0 -3.1 4.8 1.8 12 -2.8 5.0 -2.8 3.7 1.6 13 -3.0 7.6 -2.6 4.8 5.6 14 - - -1.9 3.5 1.7 15 -4.1 4.2 -2.2 4.9 3.0 16 -4.1 4.6 -3.2 2.8 2.1 17 -3.5 8.2 -2.7 4.3 4.5 18 -4.4 3.1 -2.8 3.9 2.2 MEAN -3.5 7.5 -2.7 4.4 2.6 SEM 0.2 0.8 0.0 0.2 0.3 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.    220  Table C.79. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2 raw data for each participant during the NX2 CO2 protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -4.8 9.2 -3.3 5.1 3.2 2 -3.5 6.9 -2.8 4.2 5.4 3 - - - - - 4 -3.0 9.1 -2.7 4.6 1.9 5 - - - - - 6 -4.1 7.4 -2.5 4.8 3.9 7 - - - - - 8 - - -2.2 9.9 7.6 9 - - - - - 10 -2.7 6.8 -2.2 2.4 1.3 11 -3.4 7.4 -2.3 3.3 2.3 12 -3.3 5.0 -2.5 3.0 2.4 13 -3.2 8.7 -3.4 5.2 6.1 14 - - -1.7 3.5 1.5 15 -4.0 6.5 -2.2 6.9 4.9 16 -2.9 10.0 -2.8 4.3 2.5 17 -3.5 9.7 -3.1 3.7 3.6 18 -5.1 2.8 -2.7 4.5 2.6 MEAN -3.6 7.5 -2.6 4.7 3.5 SEM 0.2 0.5 0.1 0.4 0.4  221  Table C.80. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2 raw data for each participant during the HX1 CO2 protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -5.9 8.0 -4.2 2.7 3.4 2 - - -3.0 5.9 7.8 3 -3.8 5.6 -1.6 6.4 5.6 4 - - -3.2 3.0 1.2 5 - - - - - 6 -3.3 9.4 -3.1 5.3 4.1 7 - - - - - 8 - - -3.3 9.7 4.7 9 - - - - - 10 -1.6 5.5 -2.4 2.3 2.3 11 -4.4 8.6 -4.9 6.1 2.9 12 -3.8 7.8 -3.1 3.4 2.1 13 - - - - - 14 - - -2.0 3.7 1.8 15 -4.1 9.6 -2.4 5.4 7.3 16 -2.0 9.0 -2.7 4.5 2.6 17 -4.3 8.9 -3.1 4.6 4.3 18 -3.6 7.2 -2.9 5.5 6.8 MEAN -3.7 8.0 -3.0 4.9 4.1 SEM 0.3 0.4 0.2 0.5 0.5  222  Table C.81. Internal carotid artery, middle cerebral artery, and HCVR to arterial CO2 raw data for each participant during the HX2 CO2 protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -5.2 6.6 -3.4 3.6 3.1 2 - - -3.8 8.5 9.1 3 -3.3 6.4 -2.4 5.5 4.8 4 - - -3.4 2.9 2.8 5 - - - - - 6 -3.9 8.8 -3.2 4.0 3.2 7 - - - - - 8 - - -2.0 7.5 5.7 9 - - - - - 10 -2.8 10.6 -2.7 2.3 1.8 11 -3.0 3.3 -1.5 6.0 2.2 12 -4.0 5.1 -3.5 3.5 1.3 13 - - - - - 14 - - -2.6 2.8 2.5 15 -4.6 6.2 -3.1 5.3 5.8 16 -2.2 13.4 -2.6 5.1 3.9 17 -3.3 10.8 -3.1 4.7 3.7 18 -4.2 5.3 -3.0 4.5 5.7 MEAN -3.7 7.6 -2.9 4.7 4.0 SEM 0.2 0.9 0.1 0.4 0.5  223  Table C.82. Internal carotid artery, middle cerebral artery, and HCVR to predicted arterial CO2 raw data for each participant NX1 CO2 protocol.SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -3.2 9.2 -3.1 4.5 2.1 2 -3.7 10.7 -2.9 4.5 5.8 3 - - - - - 4 -2.8 5.4 -3.3 - 1.6 5 - - - - - 6 -3.9 10.1 -2.9 3.2 3.9 7 - - - - - 8 - - -2.2 6.7 4.4 9 - - - - - 10 -1.8 9.2 -2.2 3.8 2.1 11 -6.1 5.8 -3.1 4.6 1.7 12 -2.8 4.4 -3.0 3.4 1.5 13 -3.3 6.8 -2.9 4.3 5.0 14 - - -1.8 3.4 1.6 15 -4.2 3.6 -2.3 4.3 2.6 16 -4.1 4.4 -3.2 2.9 2.1 17 -3.7 7.7 -2.9 4.0 4.3 18 -4.5 2.7 -2.9 3.4 1.9 MEAN -3.7 6.7 -2.8 4.1 2.9 SEM 0.2 0.7 0.1 0.2 0.3 Definition of abbreviations: NX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.    224  Table C.83. Internal carotid artery, middle cerebral artery, and HCVR to predicted arterial CO2 raw data for each participant during the NX2 CO2 protocol. Definition of abbreviations: NX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -4.5 8.3 -3.1 4.5 2.9 2 -3.5 6.9 -2.7 4.1 5.3 3 - - - - - 4 -2.4 7.5 -2.3 3.7 1.6 5 - - - - - 6 -5.1 6.0 -3.1 3.9 3.2 7 - - - - - 8 - - -2.4 5.7 4.4 9 - - - - - 10 -3.0 6.2 -2.5 2.2 1.2 11 -3.8 7.4 -2.7 3.4 2.3 12 -3.5 4.8 -2.7 2.9 2.3 13 -3.1 8.9 -3.3 5.0 6.0 14 - - -1.8 3.4 1.5 15 -4.5 4.4 -2.5 4.7 3.3 16 -3.0 8.3 -3.0 3.7 2.0 17 -3.5 9.5 -3.1 3.6 3.5 18 -5.0 2.5 -2.6 4.0 2.4 MEAN -3.7 6.7 -2.7 3.9 3.0 SEM 0.2 0.5 0.1 0.2 0.3  225  Table C.84. Internal carotid artery, middle cerebral artery, and HCVR to predicted arterial CO2 raw data for each participant during the HX1 CO2 protocol. Definition of abbreviations: HX1 = uncorrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response.   SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -5.2 8.1 -3.7 2.8 3.4 2 - - -2.4 6.0 7.8 3 -4.6 5.2 -2.0 5.9 5.1 4 - - -3.2 3.1 1.3 5 - - - - - 6 -3.4 9.5 -3.2 5.3 4.1 7 - - - - - 8 - - -3.1 9.5 4.6 9 - - - - - 10 -1.7 4.7 -2.6 2.0 2.0 11 -4.2 8.2 -4.6 5.8 2.8 12 -3.6 7.5 -2.9 3.2 2.0 13 - - - - - 14 - - -2.0 4.2 2.0 15 -3.1 7.7 -2.2 4.4 5.9 16 -2.6 9.1 -3.0 4.6 2.7 17 -4.5 9.0 -3.2 4.7 4.4 18 -3.5 7.5 -2.8 5.8 7.1 MEAN -3.6 7.6 -2.9 4.8 3.9 SEM 0.3 0.5 0.1 0.5 0.5  226  Table C.85. Internal carotid artery, middle cerebral artery, and HCVR to predicted arterial CO2 raw data for each participant during the HX2 CO2 protocol. Definition of abbreviations: HX2 = corrected CO2 reactivity protocol; ICA-HYPO = internal carotid artery reactivity to hypocapnia; ICA-HYPER = internal carotid artery reactivity to hypercapnia; MCA-HYPO = middle cerebral artery reactivity to hypocapnia; MCA-HYPER = middle cerebral artery reactivity to hypercapnia HCVR = hypercapnic ventilatory response SUBJECT ICA- HYPO (%MMHG) ICA-HYPER (%MMHG) MCA-HYPO (%MMHG) MCA-HYPER (%MMHG) HCVR (L/MIN/MMHG) 1 -5.4 6.7 -3.6 3.7 3.3 2 - - -3.6 9.4 10.2 3 -3.4 6.2 -2.5 5.4 4.7 4 - - -3.5 3.0 2.9 5 - - - - - 6 -4.0 9.4 -3.2 4.2 3.4 7 - - - - - 8 - - -2.3 6.3 4.7 9 - - - - - 10 -2.7 10.0 -2.6 2.1 1.7 11 -3.3 3.2 -1.6 5.9 2.2 12 -3.8 5.1 -3.3 3.5 1.3 13 - - - - - 14 - - -2.6 3.2 2.8 15 -4.5 5.3 -3.0 4.4 5.3 16 -2.3 11.2 -2.7 4.3 3.3 17 -3.3 10.4 -3.1 4.6 3.6 18 -4.1 6.2 -2.9 5.2 6.6 MEAN -3.7 7.4 -2.9 4.7 4.0 SEM 0.2 0.8 0.1 0.4 0.5 

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