To determine the feasibility of measuring gastric mucosal blood flow by laser-Doppler velocimetry (LDV), we utilized two LDV flowmeters to monitor blood flow in mucosa and serosa of chambered canine stomach. In isolated, nonautoregulating gastric segments vasodilated with isoproterenol, LDV mucosal and muscularis blood flows were both linearly related to total electromagnetic blood flow during step increases in perfusion pressure. To assess the depth of the LDV measurement, we recorded reactive hyperemia following arterial occlusion. Reactive hyperemia was frequently registered in the mucosa but rarely in muscularis. Placing a layer of nonperfused mucosa-submucosa between the probe and the perfused mucosa abolished the resting LDV mucosal flow signal and attenuated the recording of peak hyperemia by 85%. Furthermore, intra-arterial infusions of both adenosine and isoproterenol frequently increased LDV mucosal flow and decreased LDV muscularis flow, although total flow was consistently increased. These findings indicate that our LDV instruments yield linear, superficial measurements of gastric blood flow in either mucosa or muscularis. Although calibration in absolute units remains to be achieved, our results demonstrate that LDV is a practical means of studying the gastric mucosal microcirculation.