Effect of NCA on excitation-contraction coupling of cardiac muscle from phospholamban knockout mice

AIM: Nitroxyl(HNO) increases myofilament Ca²⁺ responsiveness relative to increases in intracellular Ca²⁺ in cardiac muscle. In this study, we further investigated this effect of HNO on trabecular muscles from phospholamban knockout(PLB-KO) and wide-type(WT) mice using a novel HNO donor, 1-nitrosocyclohexyl acetate(NCA). METHODS: Trabecular muscles were dissected from the right ventricles of the rat hearts and mounted between a force transducer and a motor arm. The muscles were superfused with K-H solution(pH 7.4) at room temperature. Fura-2 was loaded into the trabecular muscles via electrophoresis. The length of the sarcomere was set to 2.2~2.3μm. During steady-state activations, the maximal Ca²⁺-activated force and Ca²⁺ required for 50% activation were measured. RESULTS: The intracellular Ca²⁺ transients and force of the PLB-KO muscles at baseline were higher than those of the WT muscles and exhibited a negative force-frequency relationship(FFR). NCA(2.5μmol/L) increased systolic force in both PLB-KO group and WT group at any givenCa²⁺]_o. However, there was more dramatic increase in the force development due to moderate increases in the intracellular Ca²⁺ transients in the WT muscles when external Ca²⁺ increased from 1.5 to 4.5 mmol/L under NCA. NCA did not affect the negative FFR in PLB-KO muscle. Steady-state force-Ca²⁺ relations obtained from skinned muscles were not different between the 2 groups, while NCA increased Ca²⁺ responsiveness in skinned muscles from both PLB-KO and WT mice.CONCLUSION: HNO increases force development in both PLB-KO and WT muscles as a result of increases in myofilament Ca²⁺ responsiveness. The increased intracellular Ca²⁺ transients are accompanied by greater force development in WT mice, suggesting that HNO improves Ca²⁺ activation and establishes HNO as a positive inotropic agent with novel mechanisms.