Biochemical characterization of ethanol actions on dihydropyridine-sensitive calcium channels in brain synaptosomes

This study was undertaken to investigate the biochemical events underlying the inhibitory action of ethanol on dihydropyridine-sensitive voltage-dependent Ca2+ channels in brain synaptosomes. The binding of radiolabeled dihydropyridine was used to determine functional Ca2+ channels in synaptosomes following exposure to ethanol. No effect on [3H]PN 200-110 binding was found when disrupted synaptosomal membranes were incubated with ethanol concentrations as high as 300 mM, suggesting that ethanol did not interact directly with sites on or near the Ca2+ channels. However, when intact synaptosomes were first incubated with ethanol (100 mM) at 37 degrees and then disrupted, a significant reduction in membrane binding of [3H]PN 200-110 was found. Ethanol incubation of synaptosomes at 0 degree was ineffective. It appears that metabolic processes involving intracellular factors were required in the ethanol action. In examining this possibility, [3H]PN 200-110 binding was activated by incubation of disrupted membranes with MgATP and Ca(2+)-calmodulin, and ethanol was found to inhibit the activation in a concentration-dependent manner (50-200 mM). [3H]PN 200-110 binding to membranes was also activated by incubation with MgATP and cyclic AMP-dependent protein kinase, but this activation was not inhibited by ethanol. These findings are consistent with the interpretation that ethanol acts on Ca2+ channels by inhibiting calmodulin-dependent activation of the channels.