EFFECTS OF PKC ACTIVATION AND RECEPTOR DESENSITIZATION ON NEUROSTEROID MODULATION OF GABA(A) RECEPTORS

The effect of calcium-phospholipid-dependent protein kinase (PKC) acti vation on neurosteroid modulation of the GABA(A) receptor was examined in Xenopus oocytes expressing human recombinant alpha 1 beta 2 gamma 2L GABA(A) receptors. GABA-gated chloride currents were measured using the two-electrode voltage-clamp technique. The peak amplitude of GABA -gated chloride currents was reduced by the PKC activator phorbol 12-m yristate 13-acetate (PMA), but not by the inactive analog phorbol 12-m ono-myristate (PMM). This effect of PMA was inhibited by the protein k inase inhibitor staurosporine. To investigate whether the activation o f PKC could alter neurosteroid modulation of the GABA(A) receptor, the effect of PMA was studied on the positive allosteric modulatory stero id 3 alpha,21-dihydroxy-5 alpha-pregnan-20-one (THDOC) and the negativ e modulatory neurosteroid pregnenolone sulfate (PS). THDOC potentiatio n of GABA-gated chloride currents was found to be increased by approxi mately 120% following PMA treatment, while PS inhibition was not affec ted. The increase in THDOC potentiation by PMA was blocked by staurosp orine. No change in THDOC potentiation was observed following PMM trea tment. The enhancement of THDOC potentiation following PMA treatment w as not due to a shift in the GABA EC50. In addition to inhibiting the peak amplitude of the GABA response, PMA treatment resulted in non-des ensitizing GABA responses. Similarly, GABA responses of receptors whic h had been desensitized with prolonged GABA application also showed a reduction in peak amplitude and reduced desensitization. THDOC potenti ation of desensitized receptors was enhanced approximately 70% with re spect to non-desensitized receptors. The present results demonstrate t hat protein phosphorylation and receptor desensitization alter modulat ion of the GABA(A) receptor complex by some neurosteroids. (C) 1997 El sevier Science B.V.