The present study represents the first characterization of calcium-activated chloride currents (I_ClCa) in single smooth muscle cells from a murine vascular preparation (portal veins). I_ClCa were recorded using the perforated patch version of the whole cell voltage-clamp technique and evoked by membrane depolarization. Generation of I_ClCa was reliant upon Ca²⁺ entry through dihydropyridine-sensitive Ca²⁺ channels as I_ClCa was abolished by 1 µM nicardipine and enhanced by raising external [Ca²⁺] or by application of BAY K 8644. I_ClCa was characterized by the sensitivity to chloride channel blockers and the effect of altering the external anion on reversal potential. Activation of I_ClCa following membrane depolarization was dependent upon Ca²⁺-release from intracellular stores. Thus, the amplitude of I_ClCa was diminished by the SR-ATPase inhibitor cyclopiazonic acid (CPA), the IP₃-receptor antagonist 2-aminoethoxydiphenyl borate (2-APB) and the ryanodine receptor blocker tetracaine. The degree of inhibition produced by the application of 2-APB and tetracaine together was significantly greater than the effect of each agent applied alone. In current clamp mode, injection of depolarizing current elicited a biphasic action potential with the later depolarization being sensitive to niflumic acid (10 µM). In isometric tension recordings niflumic acid inhibited spontaneous contractions. These data support a role for this conductance in portal vein excitability.