| Abstract No.: | B-B2037 |
| Country: | Canada |
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| Title: | EVIDENCE THAT THE PHOSPHORYLATION OF GABAA RECEPTORS REGULATES NEUROSTEROID EFFICACY AFTER KINDLING |
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| Authors/Affiliations: | 1 Mirhadi-Arash Kia*; 1 Michael Poulter;
1 Robarts Research Institute and the Department of Physiology and Pharmacology, University of Western Ontario, London, ON, Canada
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| Content: | In our previous report we showed that tetra-hydro-deoxy-corticosterone (THDOC), a neurosteroid that normally enhances GABAA receptor mediated currents, has reduced efficacy after the induction of stage 5 amygdala kindled seizures. As phosphorylation of GABAA receptors has been implicated in regulating THDOC pharmacology, we hypothesized that perhaps kindling induces (long term) changes in the phosphorylation state of GABAA receptors. In order to test this hypothesis we have manipulated phosphorylation state of GABAA receptors by employing agents that activate kinases (PMA, 100 nM) or inhibit phosphatase (FK-506,100 nM). We performed whole-cell patch clamp recordings in layers 1-3 of the piriform cortex prepared from non-kindled male Sprague Dawley rats (200-250 gm). GABAA synaptic responses were isolated by applying 200 nM TTX, 50 µM APV, and 20 µM DNQX. We also carefully monitored the holding current to see if the extrasynaptic transmission may be affected. We applied THDOC (100 nM) for a minimum of 10 minutes, a protocol that has been shown previously to enhance currents by as much as 100 %. First, we found that PMA prolonged the deactivation of the mIPSCs. The subsequent application of THDOC reduced the amplitude of mIPSCs, but THDOC prolonged mIPSCs further. However, the net affect was that THDOC did not enhance charge transfer, an outcome that was identical to those found in kindled tissue. Similar results were also found with FK-506 except the reduction in charge transfer occurred in only a subset of recordings. Both the inactive phorbol ester (4a-PMA) and the PKC antagonist (BIS I, 100 nM) had no effect on the THDOC modulation. In presence of PMA the small change in holding current that normally occurs due to THDOC application was reduced or induced an outward shift while FK-506 was without effect. These data suggest that phosphorylation has profound effects on THDOC and that kindling may reduce the THDOC efficacy by inducing the phosphorylation of GABAA receptors.
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