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-induced 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.