Regulation of extrasynaptic GABAA α4 receptors by ethanol-induced PKA, but not PKC activation in cultured rat cerebral cortical neurons.

Ethanol produces changes in GABA(A) receptor trafficking and function that contribute to ethanol dependence symptomatology. Extrasynaptic gamma-aminobutyric acid A receptors (GABA(A)-R) mediate inhibitory tonic current and are of particular interest because they are potentiated by physiologically relevant doses of ethanol. Here, we isolate GABA(A) alpha 4 delta receptors by western blotting in subsynaptic fractions to investigate protein kinase A (PKA) and protein kinase C (PKC) modulation of ethanol-induced receptor trafficking, while extrasynaptic receptor function is determined by measurement of tonic inhibition and responses evoked by 4,5,6,7-tetrahydroisoxazolo[5,4-c] pyridin-3-ol (THIP). Rat cerebral cortical neurons were grown for 18 days in vitro and exposed to ethanol and/or PKA/PKC modulators. Ethanol exposure (1 hour) did not alter GABA(A) alpha 4 receptor abundance, but it increased tonic current amplitude, an effect that was prevented by inhibiting PKA, but not PKC. Direct activation of PKA, but not PKC, increased the abundance and tonic current of extrasynaptic alpha 4 delta receptors. In contrast, prolonged ethanol exposure (4 hours) reduced alpha 4 delta receptor abundance as well as tonic current, and this effect was also PKA dependent. Finally, PKC activation by ethanol or phorbol-12,13-dibutyrate (PdBu) had no effect on extrasynaptic alpha 4 delta subunit abundance or activity. We conclude that ethanol alters extrasynaptic alpha 4 delta receptor function and expression in cortical neurons in a PKA-dependent manner, but ethanol activation of PKC does not influence these receptors. These results could have clinical relevance for therapeutic strategies to restore normal GABAergic functioning for the treatment of alcohol use disorders.