For a study of the underlying mechanisms of a possible interaction between ethanol and nicotinic receptors during ethanol dependence, the aim of this work was to investigate the effect of chronic ethanol exposure on nicotinic receptor subtypes in a transfected fibroblast cell line (M10 cells) stably expressing alpha4beta2 nicotinic receptor subtype and an SH-SY5Y neuroblastoma cell line expressing alpha3, alpha5, alpha7, beta2, and beta4 nicotinic acetylcholine receptor (nAChR) subunits. A significant dose-related decrease (-30-80%) in number of [3H]nicotine binding sites was observed in ethanol-treated (25-240 mM) compared with untreated M10 cells. Similarly, 4-day treatment with ethanol in concentrations relevant to chronic alcoholism (100 mM) decreased the number of nicotinic receptor binding sites in the SH-SY5Y cells when measured using [3H]epibatidine. When M10 cells were chronically treated with nicotine, ethanol partly inhibited the up-regulation of nicotinic receptors when present in the cells together with nicotine. Chronic treatment for 4 days with 100 mM ethanol significantly decreased the mRNA level for the alpha3 nAChR subunit (-39%), while the mRNA levels for the alpha7 (+30%) and alpha4 (+22%) subunits were significantly increased. Chronic ethanol treatment did not affect the mRNA levels for the beta2 nAChR subunit. Changes in the levels of nAChR protein and mRNA may have adaptive significance and be involved in the development of dependence, tolerance, and addiction to chronic ethanol and nicotine exposure. They also may be targets for therapeutic strategies in the treatment of ethanol and nicotine dependence.