The current study examined the role of extracellular Ca2+, calmodulin and myosin light-chain kinase (MLCK) in pervanadate-induced constriction of cannulated, pressurized rat cremaster arterioles. Pervanadate (0.03-100 microM) induced a concentration-dependent constriction of arterioles that was significantly attenuated (P<0.05) by the tyrosine kinase inhibitor tyrphostin 47 (30 microM). The L-type voltage-sensitive Ca2+ channel antagonists verapamil (10 microM) and nifedipine (1 microM) dilated vessels possessing myogenic tone but had no demonstrable effect on pervanadate constriction, while a higher concentration of nifedipine (10 microM) reduced constriction by approximately 50%. Pervanadate-induced contractions were reduced by the calmodulin inhibitor W-7 (N-(6-aminohexyl)-chloro-1-naphtalene sulphonamide, 50 microM) and the MLCK inhibitor ML-7 (1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine, 10 or 30 microM). Removal of extracellular Ca2+ abolished the contractile effect of pervanadate. Measurement of changes in arteriolar wall [Ca2+] using the Ca2+ sensitive dye Fura-2 showed that pervanadate did not increase [Ca2+] during arteriolar constriction. These observations suggest that pervanadate-induced contraction of smooth muscle in the cremaster arteriole involves Ca2+/calmodulin-dependent myosin phosphorylation and possibly sensitization of the contractile apparatus to Ca2+.