The effect of extracellular ATP on steroidogenesis in primary cultured bovine adrenocortical fasciculata cells was investigated. I observed that in the presence of extracellular Ca2+, ATP caused a dose-dependent elevation of intracellular Ca2+ ([Ca2+]i) and induced steroidogenesis concentration- and time-dependently. However, in the absence of extracellular Ca2+, ATP had no effect on steroidogenesis. In the presence of extracellular Ca2+, calmodulin inhibitors inhibited the ATP steroidogenesis, but dihydropyridine calcium channel blockers did not. Furthermore, ATP did not cause an elevation of cyclic AMP in bovine adrenocortical fasciculata cells even if extracellular Ca2+ existed. These results suggest that extracellular ATP might have an influence on bovine adrenocortical cells via the purinoceptor (P2Y) in connection with calcium mobilization, open the non-selective calcium channel and induce steroidogenesis by means of an elevation of [Ca2+]i via the calcium-calmodulin system.