Strong Binding of Myosin Modulates Length-Dependent Ca²⁺ Activation of Rat Ventricular Myocytes

From the Department of Physiology, University of Wisconsin School of Medicine, Madison.

Correspondence to Dr Daniel P. Fitzsimons, Department of Physiology, University of Wisconsin Medical School, 1300 University Ave, Madison, WI 53706. E-mail fitzsimo{at}facstaff.wisc.edu

Abstract—Reductions in sarcomere length (SL) and concomitant increases in interfilament lattice spacing have been shown to decrease the Ca²⁺ sensitivity of tension in myocardium. We tested the idea that increased lattice spacing influences the SL dependence of isometric tension by reducing the probability of strong interactions of myosin crossbridges with actin, thereby decreasing cooperative activation of the thin filament. Single ventricular myocytes were isolated by enzymatic digestion of rat hearts and were subsequently rapidly skinned. Maximal tension and Ca²⁺ sensitivity of tension (ie, pCa₅₀) were measured in the absence and presence of N-ethylmaleimide–modified myosin subfragment 1 (NEM-S1) at both short and long SLs. NEM-S1, a strong-binding non–tension-generating derivative of the myosin head, was applied to single skinned myocytes to cooperatively promote strong binding of endogenous myosin crossbridges. Compared with control myocytes at SL of 1.90 µm, application of NEM-S1 markedly increased submaximal Ca²⁺-activated tensions and thereby increased Ca²⁺ sensitivity; ie, pCa₅₀ increased from 5.40±0.02 to 5.52±0.02 pCa units in the presence of NEM-S1. Furthermore, NEM-S1 treatment reversibly eliminated the SL dependence of the Ca²⁺ sensitivity of tension, in that the pCa₅₀ between short and long lengths was 0.02±0.01 pCa units in the presence of NEM-S1 compared with a pCa₅₀ of 0.10±0.01 pCa units in control myocytes. From these results we conclude that the decrease in the Ca²⁺ sensitivity of tension at short SL results predominantly from decreased cooperative activation of the thin filament due to reductions in the number of strong-binding crossbridges.

Key Words: Ca²⁺ sensitivity • muscle length • ventricular myocyte

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