The ability of alcohols to regulate InsP3-receptor activity was examined in permeabilized hepatocytes. Incubation with 30-300 mM ethanol decreased the sensitivity to InsP3 for Ca2+ release, with little effect on the size of the Ca2+ store that could be released with maximal concentrations of InsP3. Ethanol (300 mM) increased the EC50 for InsP3 from a control value of 134.0 +/- 13.5 nM to 220.0 +/- 25.9 nM. Although ethanol also caused a partial depletion of the total pool of stored Ca2+, the ethanol-induced shift in InsP3 sensitivity was not secondary to this alteration in Ca2+ loading. Partial depletion of the Ca2+ stores with low doses of ionomycin and thapsigargin did not cause a shift in InsP3 sensitivity. Furthermore, measurements of InsP3 receptor channel activity using retrograde flux of Mn2+ to quench the fluorescence of Fura-2 within the Ca2+ stores demonstrated that ethanol inhibited InsP3-activated channel activity in the absence of stored Ca2+. Other short chain alcohols (methanol, 1-propanol and 1-butanol) also decreased the efficacy of InsP3 to release Ca2+. Measurements of 3H-InsP3 binding demonstrated that ethanol decreased the total number of InsP3 binding sites without changing the KD. The effect of ethanol on InsP3 binding was apparent in the presence or absence of Ca2+ and was observed when the cells were pre-incubated with ethanol at either 37 degrees C or 4 degrees C. The initial rate of InsP3-induced Mn2+ quenching of compartmentalized Fura-2 was reduced by ethanol at all doses of InsP3. These data suggest that ethanol decreases the sensitivity of the intracellular Ca2+ store to release by InsP3, by reducing the number of channels that can be activated by InsP3.