Experiments were designed to determine whether contractility of trout smooth muscle in vitro varied with temperature and if changes occurred at the receptor or intracellular levels. The role of calcium in contractility at various temperatures was also investigated. Isolated trout intestinal segments, approximately 2 cm in length, were suspended isometrically under 2 g tension in 10-mL organ baths containing trout Ringer's solution aerated with O2 and CO2 (95:5). Contractions of trout intestine were not statistically different at 10 and 20 degrees C for carbachol, 5-hydroxytryptamine, and KCl. However, the efficacy, but not the potency, of each agonist was decreased at 2 degrees C. Receptor-induced contractions were reduced to a greater extent at 2 degrees C and did not recover to the same extent when returned to 10 degrees C in comparison with those induced by depolarization. The calcium source for contractility was also dependent on temperature. As temperatures increased, utilization of intracellular calcium increased, as indicated by increased contractility in the absence of extracellular calcium. Thus, low temperatures decrease smooth muscle contractility by affecting receptor-mediated events rather than the intracellular contractile mechanisms. Receptor-operated agonists appear to have a higher capability of using intracellular calcium than depolarizing agents.