Exogenous carbon monoxide (0.2%) increases L-type calcium (Ca2⁢) current in human jejunal circular smooth muscle cells. The stimulatory effect of carbon monoxide (CO) on L-type Ca2⁢ current is inhibited by pre-application of L-NNA, a classical competitive inhibitor of nitric oxide synthase (NOS) with no significant isoform selectivity (Lim, 2003). In the present study, we investigated which isoform of NOS affected CO induced stimulation of L-type Ca2⁢ current in human jejunal circular smooth muscle cells. Cells were voltage clamped by whole-cell mode patch clamp technique, and membrane currents were recorded with 10 mM barium as the charge carrier. Before the addition of CO, cells were pretreated with each inhibitor of three NOS isoforms for 15 minutes. CO-stimulating effect on L-type Ca2⁢ current was partially blocked by N-(3-(Amino-methyl) benzyl) acetamidine·2HCl (1400W, an iNOS inhibitor). On the other hand, 3-bromo-7-nitroindazole (BNI, a nNOS inhibitor) or N5-(1-Iminoethyl)-L-ornithine dihydrochloride (L-NIO, an eNOS inhibitor) completely blocked the CO effect. These data suggest that low dose of exogenous CO may stimulate all NOS isoforms to increase L-type Ca2⁢ channel through nitric oxide (NO) pathway in human jejunal circular smooth muscle cells.