UBC Theses and Dissertations
An investigation into the potential involvement of vasopressin in the regulation of amniotic fluid circulation Plath, Susan Marie
The mechanisms which control the accumulation and circulation of the amniotic fluid are poorly understood. Earlier studies had made the tentative suggestion that vasopressin, which is important in the control of water balance in adults, might play a role in the regulation of amniotic fluid turnover. In the work reported here, attempts were made to detect and measure the vasopressin levels in the amniotic fluid of guinea pigs, sheep and humans. Unconcentrated amniotic fluid samples from the guinea pig, sheep and human were assayed using the water-loaded, alcohol-anesthetized rat antidiuretic preparation. If vasopressin is present in the fluid of these animals, it would be at a concentration below the detectable limits of the assay, which were found to be 10 to 15 microunits/ml. Only 4 out of 36 experiments on guinea pig amniotic fluid gave even a suggestion of a response, with 3 of these being too small to be quantified accurately. No sheep or human samples produced an antidiuresis. An attempt was made to dehydrate the maternal guinea pigs, as it. was thought that this would create an osmotic stress in the fetus which might result in increased output of vasopressin. In 29 experiments with fluid of fetuses whose mothers had been without water for 24 or 48 hours prior to collection, 27 gave no response. The responses to the two injections that did indicate activity were too low for accurate quantification. However, these experiments were not considered conclusive, and more extended investigation is needed. Seven human amniotic fluid samples were concentrated on CM-25 Sephadex columns, along with three standard vasopressin loads. The standards showed recoveries of 80% to 90% under optimal conditions. An antidiuretic substance was found in fractions from all of these samples, and it eluted at a similar pH and molarity to the standard vasopressin. This material measured approximately 200 micro-units/ml, and was relatively consistent in all of the samples tested. The amniotic fluid antidiuretic substance (AFAS) appeared to respond with an antidiuresis similar to that of vasopressin on the assay preparation. However, further work suggested that the AFAS could not be identified as vasopressin. Although the pattern of antidiuretic responses were similar, there were marked changes in the sensitivity of the bioassay preparation to the unknown material during periods of little change in the responses to vasopressin itself. A similar contrast was found between different experiments. Sodium thioglycollate incubation also failed to deactivate the active AFAS fractions, whilst control volumes of vasopressin lost apparently all activity following this procedure. The stability characteristics were also dissimilar to those of vasopressin, as the fractions remained active for up to five weeks when stored at neutral pH at 4° C. Optimum pH conditions for the storage of vasopressin are between 3.0 and 5.0. An attempt was made to identify the AFAS as angiotensin II. However, the response pattern on the records of the antidiuretic assay were dissimilar. Further comparisons with combinations of vasopressin and angiotensin II in varying concentrations also failed to mimic the responses to the AFAS. It was concluded that the antidiuretic activity found in the human amniotic fluid was not attributable to vasopressin, angiotensin II, or to a combination of these two hormones, at the levels tested. The nature of the AFAS is at present unknown. Further studies are needed to identify the unknown antidiuretic agent extracted from human amniotic fluid, and the possible mechanisms of its release, and its potential physiological functions, are still unknown.
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