Blebbistatin: use as inhibitor of muscle contraction

Abstract

Blebbistatin (BLEB) is a recently discovered compound that inhibits myosin-II ATPase activity. In this study, we tested BLEB in intact and skinned isolated rat cardiac trabeculae, rat intact myocytes, and single rabbit psoas myofibrils. BLEB (10 μM) reduced twitch force and cell shortening that was reversed by exposure to light. BLEB treatment of skinned trabeculae in the dark (1 hr) reduced Ca²⁺-activated force $\left( {{\text{EC}}_{50} = 0.38 \pm 0.03\;{\text{ $ \mu $ M}}} \right)$. Rapid (<5 ms) BLEB application in Ca²⁺-activated rabbit myofibrils reduced force proportional to [BLEB]. Two-photon Indo1-AM ratio-metric confocal line-scan microscopy revealed no impact of BLEB on the cytosolic Ca²⁺ transient. BLEB reduced contractile force in skinned trabeculae without affecting tension-dependent myofilament ATPase activity. We conclude that BLEB specifically uncouples cardiac myofilament activation from Ca²⁺ activation without affecting EC coupling or cross-bridge cycling parameters. This agent could be useful to uncouple myofilament contractility from electrical events that lead to sarcoplasmic reticulum Ca²⁺ release in the cardiac myocyte (uncoupling agent) However, the compound is very sensitive to light, a property that limits its application to mechanistic physiological studies.