Glycolysis is slow in the heart, especially in the cardiomyopathic heart. Glycolysis is partially rate-limited by phosphofructokinase (PFK), an enzyme which is inhibited by calcium (Ca2+)i and hydrogen ions (H+)i and activated by cAMP. (H+)i and (Ca2+)i are augmented in cardiomyopathy. With glucose as the only substrate (NADH)/(NAD) the phosphorylation potential and developed pressure were significantly lower, and concentrations of phosphomonoester sugars and hydrogen ions (H+)i were significantly higher in isolated cardiomyopathic hearts as compared to healthy hamster hearts. Pyruvate lowered diastolic (Ca2+)i in cardiomyopathic hamster hearts. With pyruvate as the substrate (NADH)/(NAD), the phosphorylation potential and developed pressure increased significantly and concentrations of phosphomonoester sugars (PME), (H+)i and diastolic (Ca2+)i decreased significantly in myopathic hamster hearts. The results suggest that late heart failure in the myopathic hamster is associated with calcium and/or hydrogen ion-induced inhibition of glycolysis.