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A role of protein kinase C (PKC) and photosphatidylinositol 3-kinase (PI 3-K) in motoneuron dysfunction

University of British Columbia

Amyotrophic lateral sclerosis (ALS), also known as motoneuron disease, is a neurodegenerative disorder of humans characterized by motoneuron loss in the spinal cord, brainstem, corticospinal tract neurons and other regions. Human postmortem data from spinal cord tissue of ALS patients revealed a significant increase in the activities of protein kinase C (PKC) and phosphatidylinositol 3- kinase (PI 3-K) compared to control subjects. Protein levels of PKC, PI 3-K, protein kinase B (PKB) and p70 S6 kinase (p70 S6K) were also elevated in spinal cord tissue from ALS patients compared to controls. The activities and protein levels of PI 3-K, PKC, PKB, p70 S6K, Extracellular signal-regulated kinase (Erk)1 and Erk2 were not different in brain regions such as the motor cortex and visual cortex between ALS patients and control subjects. This data indicate that alteration in the activities of PKC and PI 3-K is present in the spinal cord tissue from ALS patients, an area that is pathologically affected in ALS. One possible pathogenic mechanism of motoneuron death in ALS is the activation of glutamate receptors such as the NMDA receptor. To investigate the role of PKC in NMDA-mediated cell death in HEK 293 cells transfected with NR1A/NR2A subunits of the NMDA receptor, phorbol ester, a PKC agonist was added to the transfected HEK cells. Exposure to phorbol ester significantly augmented NMDA-mediated cell death and this augmentation could be antagonized by a specific PKC inhibitor (RO 320432) in the NR1A/NR2A transfected HEK cells. These data indicate that activation of PKC can potentiate neurotoxicity through NMDA receptor activation. The pmn/pmn mouse is considered an animal model of motoneuron disease. PI 3-K was significantly reduced in spinal cord tissue of pmn/pmn mice compared to control mice at the age when these mice show significant impairment in locomotion and neurological function. In the cerebellum and brainstem, PI 3-K activities and protein levels were not significantly different between affected and control mice. The neurotrophic factor, BDNF was exogenously applied to these mice and caused a significant increase in PI 3-K activity in spinal cord tissue from both pmn/pmn and control mice, with a greater increase in the pmn/pmn mice. The role of PI 3-K and MEK inhibitors in the retrograde transport of fluorochromes from transected sciatic nerve was studied in pmn/pmn and control mice. The MEK inhibitor PD98059 had no effect on the number of labelled motoneurons in pmn/pmn and control mice. The PI 3-K inhibitor LY 294002 reduced the number of labelled motoneurons in control mice, but had no effect on the labelling of motoneurons in the pmn/pmn mice. Only in the presence of BDNF did LY 294002 inhibit retrograde labelling of motoneurons in the pmn/pmn mice. This data indicated that at least some of the retrograde labelling of motoneuron is PI 3-Kdependent in control mice. In the pmn/pmn mice, the reduced PI 3-K activity requires administration of BDNF to stimulate PI 3-K activity in order to improve motoneuron labelling. Thus, PI 3-K is important in the uptake and/or retrograde transport of substances by motoneurons and PI 3-K activity is altered in mice with motoneuron dysfunction. Abnormal regulation of PI 3-K and PKC, together with processes such as excitotoxicity, could contribute to motoneuron death in ALS.

Thesis/Dissertation

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2009-09-25

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http://hdl.handle.net/2429/13169

Medical Genetics

University of British Columbia

UBCV

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