Negative inotropic mechanisms of beta-cardiotoxin in cardiomyocytes by depression of myofilament ATPase activity without activation of the classical beta-adrenergic pathway

Beta-cardiotoxin (beta -CTX) from the king cobra venom (Ophiophagus hannah) was previously proposed as a novel beta -adrenergic blocker. However, the involvement of beta -adrenergic signaling by this compound has never been elucidated. The objectives of this study were to investigate the underlying mechanisms of beta -CTX as a beta -blocker and its association with the beta -adrenergic pathway. The effects of beta -CTX on isolated cardiac myocyte functions, calcium homeostasis, the phosphorylation level of targeted proteins, and the myofibrillar ATPase activity were studied. Healthy Sprague Dawley rats were used for cardiomyocytes isolation. Like propranolol, beta -CTX attenuated the cardiomyocyte inotropy and calcium transient alterations as induced by isoproterenol stimulation. In contrast, these effects were not observed in forskolin-treated cells. Interestingly, cardiomyocytes treated with beta -CTX showed no changes in phosphorylation level at any PKA-targeted sites in the myofilaments as demonstrated in Western blot analysis. The skinned fibers study revealed no change in myofilament kinetics by beta -CTX. However, this protein exhibited the direct inhibition of myofibrillar ATPase activity with calcium de-sensitization of the enzyme. In summary, the negative inotropic mechanism of beta -CTX was discovered. beta -CTX exhibits an atypical beta -blocker mechanism. These properties of beta -CTX may benefit in developing a novel agent aid to treat hypertrophic cardiomyopathy.