The effects of angiotensin (Ang) II, arginine vasopressin (AVP), and serotonin (5-HT) on inositol trisphosphate (IP3) production, cytosolic Ca2+ concentration [( Ca2+]i), and the phosphorylation of 20 kilodalton myosin light chain (MLC) were examined in cultured rat vascular smooth muscle cells (VSMC). These vasoconstrictors immediately induced dose-dependent and concomitant increases in IP3 production, [Ca2+]i, and phosphorylation of 20 kDa MLC. The agonist-induced increases in IP3 production, [Ca2+]i, and phosphorylation of 20 kDa MLC in VSMC were completely blocked by pretreatment with the receptor antagonists for Ang II, V1, and 5-HT2, respectively. These data suggest that receptor-mediated increases in IP3 production, [Ca2+]i, and phosphorylation of 20 kDa MLC by these vasoconstrictors are closely interrelated.