Regulation of Extrasynaptic GABAA α4 Receptors by Ethanol-Induced Protein Kinase A, but Not Protein Kinase C Activation in Cultured Rat Cerebral Cortical Neurons.
Ethanol produces changes in GABAA receptor trafficking and function that contribute to ethanol dependence symptomatology. Extrasynaptic γ-aminobutyric acid A receptors (GABAA-R) mediate inhibitory tonic current and are of particular interest because they are potentiated by physiologically relevant doses of ethanol. Here, we isolate GABAA α4δ 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 GABAA α4 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 α4δ receptors. In contrast, prolonged ethanol exposure (4 hours) reduced α4δ 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 α4δ subunit abundance or activity. We conclude that ethanol alters extrasynaptic α4δ 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.
Bowles Center for Alcohol Studies and Departments of Psychiatry and Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina.
Stephen L
SL
Carlson
Bowles Center for Alcohol Studies and Departments of Psychiatry and Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina.
J Peyton
JP
Bohnsack
Bowles Center for Alcohol Studies and Departments of Psychiatry and Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina.
Vraj
V
Patel
Bowles Center for Alcohol Studies and Departments of Psychiatry and Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina morrow@med.unc.edu.
A Leslie
AL
Morrow
D000818:Animals; D001640:Bicuculline / Q000494:pharmacology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D020536:Enzyme Activators / Q000494:pharmacology*; D000431:Ethanol / Q000494:pharmacology*; D005260:Female; D018756:GABA Antagonists / Q000494:pharmacology; D007555:Isoxazoles / Q000494:pharmacology; D008297:Male; D009474:Neurons / Q000187:drug effects*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011963:Receptors, GABA-A / Q000187:drug effects*; D013569:Synapses / Q000187:drug effects / Q000201:enzymology
D020536:Enzyme Activators; D018756:GABA Antagonists; C546262:Gabra4 protein, rat; D007555:Isoxazoles; D011963:Receptors, GABA-A; D015240:Phorbol 12,13-Dibutyrate; D000431:Ethanol; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; C015542:gaboxadol; D001640:Bicuculline