Sympathetic nerve stimulation causes a constriction of submucosal arterioles that is mediated by ATP acting at P2 receptors. In the present study, we examined the P2 receptor subtype mediating this response. A computer-assisted video monitoring system measured drug-induced changes in arteriolar diameter (resting diameter approximately 40-80 microns) in pieces of submucosa in vitro. The rank-order potency for vasoconstriction caused by several ATP analogs was alpha,beta-methylene ATP (alpha,beta-MeATP) > beta,gamma-methylene ATP = 2-methylthioATP > ATP > ADP. Constrictions caused by alpha,beta-MeATP were competitively antagonized by suramin (KB = 3.2 microM) and were noncompetitively blocked by pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (KB = 0.5 microM). Norepinephrine-induced constrictions were not affected by suramin (100 microM) or by pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (3 microM). Extracellular solutions with 0 added calcium prevented constrictions caused by alpha,beta-MeATP (0.1-1.0 microM), but not those caused by norepinephrine (3 and 10 microM). Nifedipine (0.3 and 1 microM) reduced constrictions caused by 40 and 60 mM extracellular potassium chloride, but not those caused by alpha,beta MeATP. These data indicate that P2X receptors mediate constriction of submucosal arterioles. P2X-mediated vasoconstriction is dependent on extracellular calcium, but calcium entry through nifedipine-sensitive calcium channels does not contribute to P2X-mediated vasoconstriction.