Abstract

The interaction between the effects of the endogenous cannabinoid receptor agonist anandamide and ethanol on the function of homomeric α₇-nicotinic acetylcholine (nACh) receptors expressed in Xenopus oocytes were investigated using the two-electrode voltage-clamp technique. Anandamide and ethanol reversibly inhibited currents evoked with 100 μM acetylcholine in a concentration-dependent manner. Coapplication of anandamide and ethanol caused a significantly greater inhibition of α₇-nACh receptor function than anandamide or ethanol alone. The IC₅₀ value of 238 ± 34 nM for anandamide inhibition decreased significantly to 104 ± 23 nM in the presence of 30 mM ethanol. The inhibition of α₇-mediated currents by coapplication of anandamide and ethanol was not altered by phenylmethylsulfonyl fluoride, an inhibitor of anandamide hydrolyzing enzyme, or N-(4-hydroxyphenyl)-arachidonylamide, an anandamide transport inhibitor. Analysis of oocytes by matrix-assisted laser desorption/ionization technique indicated that ethanol treatment did not alter the lipid profile of oocytes, and there is negligible, if any, anandamide present in these cells. Results of studies with chimeric α₇-nACh-5-HT₃ receptors comprised of the amino-terminal domain of the α₇-nACh receptor and the transmembrane and carboxyl-terminal domains of 5-HT₃ receptors suggest that although ethanol inhibition of the α₇-nACh receptor is likely to involve the N-terminal region of the receptor, the site of action for anandamide is located in the transmembrane and carboxyl-terminal domains of the receptors. These data indicate that endocannabinoids and ethanol potentiate each other's inhibitory effects on α₇-nACh receptor function through distinct regions of the receptor.