Comparison of elementary steps of the cross-bridge cycle in rat papillary muscle fibers expressing α- and β-myosin heavy chain with sinusoidal analysis.

In mammalian ventricles, two myosin heavy chain (MHC) isoforms have been identified. Small animals express α-MHC, whereas large animals express β-MHC, which contribute to a large difference in the heart rate. Sprague-Dawley rats possessing ~99% α-MHC were treated with propylthiouracil to result in 100% β-MHC. Papillary muscles were skinned, dissected into small fibers, and used for experiments. To understand the functional difference between α-MHC and β-MHC, skinned-fibers were activated under the intracellular ionic conditions: 5 mM MgATP, 1 mM Mg2+ , 8 mM Pi, 200 mM ionic strength, pH 7.00 at 25 °C. Small amplitude sinusoidal length oscillations were applied in the frequency range 0.13-100 Hz (corresponding time domain: 1.6-1200 ms), and effects of Ca2+ , Pi, and ATP were studied. The results show that Ca2+ sensitivity was slightly less (10-15%) in β-MHC than α-MHC containing fibers. Sinusoidal analysis at pCa 4.66 (full Ca2+ activation) demonstrated that, the apparent rate constants were 2-4× faster in α-MHC containing fibers. The ATP study demonstrated that, in β-MHC containing fibers, K1 (ATP association constant) was greater (1.7×), k2 and k-2 (cross-bridge detachment and its reversal rate constants) were smaller (×0.6). The Pi study demonstrated that, in β-MHC containing fibers, k4 (rate constant of the force-generation step) and k-4 were smaller (0.75× and 0.25×, respectively), resulting in greater K4 (3×). There were no differences in active tension, rigor stiffness, or K2 (equilibrium constant of the cross-bridge detachment step). Our study further demonstrated that there were no differences in parameters between fibers obtained from left and right ventricles, but with an exception in K5 (Pi association constant).