Increased ATP-sensitive potassium (KATP) channel activity in cardiac muscle during hypertension and myocardial hypertrophy may be induced by the release of endogenous proteases, altering inhibitory binding sites for intracellular ATP. To test this hypothesis, we studied the effects of trypsin (1.5 mg/ml) on channel sensitivity to ATP in myocytes from control (WKY) and spontaneously hypertensive rats (SHR). Trypsin increased channel activity in 63% of membrane patches from WKY rats, but in only 29% from SHR. Pre-treatment with trypsin decreased sensitivity to the inhibitory effects of ATP in both groups. These results support the possibility that KATP channel modification during chronic metabolic stress is caused by intracellular proteolysis, which decreases sensitivity to [ATP]i.