Coronary blood flow in the subendocardium is preferentially increased by adenosine but is redistributed to the subepicardium during ischemia in association with coronary pressure reduction. The mechanism for this flow redistribution remains unclear. Since adenosine is released during ischemia, it is possible that the coronary microcirculation exhibits a transmural difference in vasomotor responsiveness to adenosine at various intraluminal pressures. Although the ATP-sensitive K⁺ (K_ATP) channel has been shown to be involved in coronary arteriolar dilation to adenosine, its role in the transmural adenosine response remains elusive. To address these issues, pig subepicardial and subendocardial arterioles (60-120 µm) were isolated, cannulated, and pressurized to 20, 40, 60, or 80 cmH₂O without flow for in vitro study. At each of these pressures, vessels developed basal tone and dilated concentration dependently to adenosine and the K_ATP channel opener pinacidil. Subepicardial and subendocardial arterioles dilated equally to adenosine and pinacidil at 60 and 80 cmH₂O luminal pressure. At lower luminal pressures (i.e., 20 and 40 cmH₂O), vasodilation in both vessel types was enhanced. Enhanced vasodilatory responses were not affected by removal of endothelium but were abolished by the K_ATP channel inhibitor glibenclamide. In a manner similar to reducing pressure, a subthreshold dose of pinacidil potentiated vasodilation to adenosine. In contrast to adenosine, dilation of coronary arterioles to sodium nitroprusside was independent of pressure changes. These results indicate that coronary microvascular dilation to adenosine is enhanced at lower intraluminal pressures by selective activation of smooth muscle K_ATP channels. Since microvascular pressure has been shown to be consistently lower in the subendocardium than in the subepicardium, it is likely that the inherent pressure gradient in the coronary microcirculation across the ventricular wall may be an important determinant of transmural flow in vivo during resting conditions or under metabolic stress with adenosine release.