Addition of thrombin (Thr) or adenosine triphosphate (ATP) to rat aortic smooth muscle cells in culture (A-10, ATCC CRL 1476) induced rapid formation of inositol phosphates and release of intracellular calcium. These responses depended on the concentration of Thr and ATP used. The Thr effect was blocked by the Thr antagonist, hirudin. ADP was almost as effective as ATP, but AMP was ineffective in mediating the effects. Pretreatment of the cells with pertussis toxin (PT) resulted in partial (60-70%) inhibition of Thr- and ATP-mediated calcium release, suggesting that in smooth muscle cells, Thr and ATP (purinergic P2) receptors are coupled to phosphoinositide specific PLC (PI-PLC) through PT-sensitive guanine nucleotide binding (G) proteins.