The effects of pharmacological activation and inhibition of protein kinase C (PKC) and temperature on the relationship between cytoplasmic Ca²⁺ and lumen diameter were studied in pressurized (50 mmHg) rat posterior cerebral arteries permeabilized with -toxin. Increasing Ca²⁺ concentrations (30 nM-10 µM, 22°C) induced stable, concentration-dependent constrictions with a half-maximal effective concentration (EC₅₀) of 112 nM. The maximal constriction was 80% of baseline diameter and 157% of that during depolarization of nonpermeabilized vessels with 124 mM KCl. Elevation of temperature to 37°C increased the EC₅₀ to 246 nM and enhanced the steepness of concentration-response curves. Exposure of permeabilized arteries to indolactam V, an activator of PKC, resulted in a significant myofilament Ca²⁺ sensitization (e.g., EC₅₀ at 5 µM = 126 nM) without changing efficacy. The effects of calphostin C, a PKC inhibitor, on Ca²⁺ sensitivity were minimal; however, the amplitude of Ca²⁺-induced constrictions in both control and indolactam-treated arteries was suppressed in a concentration-dependent manner. Thus 1) temperature is an important variable in studies of arterial Ca²⁺ sensitivity, and 2) changes in PKC activity can significantly alter both myofilament sensitivity to and constrictor efficacy of cytosolic Ca²⁺.