Mechanical Load Effects on Cardiac Action Potential and Arrhythmogenic Ca2+Activitiesrevealed by a Novel Patch-Clamp-In-Gel Technology

Abstract

Cardiomyocytes are under mechanical load when the heart pumps blood against peripheral resistance. However, the mechanical load effects on cells have been missed in previous patch-clamp experiments because the cells were bathed in solution under load-free condition. We have developed an innovative Patch-Clamp-in-Gel technique to embed single cells in a 3-D hydrogel made of viscoelastic polymer matrix. Experiments were performed when mouse ventricular myocytes were contracting in-gel under mechanical load. (1) Compared to load-free cells, the myocytes in-gel under mechanical load showed prolonged action potential duration (long APD), early afterdepolarization (EAD), delayed afterdepolarization (DAD), and triggered AP, indicating that mechanical load increases arrhythmogenic AP activities. (2) Simultaneous triple-signal recordings of AP, Ca2+, and contraction revealed that load-induced DAD occurs in conjunction with spontaneous Ca2+ wave. (3) The EADs, DADs, and Ca2+ tides were abolished by blocking the late Na+ current, suggesting changes in the Na+ and Ca2+ homeodynamics. (4) The load-induced arrhythmogenic activities were abolished by specific inhibition of nitric oxide synthase 1 (NOS1 or nNOS), indicating a critical role of nNOS in the mechano-chemo-electro-transduction. Our data show that mechanical load significantly affects action potential, Ca2+ signaling, and myocyte contraction. The Patch-Clamp-in-Gel technology provides a new tool to control mechanical load on single myocytes, which enables studying mechanotransduction effects on the three dynamic systems - electrical, Ca2+ signaling, and contractile systems - that control cardiac function and arrhythmogenesis.