Simultaneous Measurement of Force and Lattice Spacing in Skinned Cardiac Fibers

Abstract

This study explores the relationship between Ca2+-activated force and myofilament lattice spacing (LS) in skinned cardiac muscle fibers. LS measurements were obtained using synchrotron x-ray diffraction while isometric force was simultaneously monitored with a force transducer. Dextran T500 (3%) compressed the lattice to physiological LS of intact muscle, and sarcomere length was set to 2.1μm at pCa8 using HeNe-laser diffraction. Fibers were isolated from left-ventricle of wild-type (WT) or troponin C knock-in mice bearing the homozygous mutation Ala8Val, which causes hypertrophic cardiomyopathy (KI-cTnC-A8V+/+). Cardiac fibers from KI-TnC-A8V+/+ have been reported to have 2.5x greater myofilament Ca2+-sensitivity compared to WT. Since WT and KI fibers generate different levels of force at the same [Ca2+], they provide a powerful tool for investigating whether LS is tightly coupled to force-development. Compared to WT, KI-TnC-A8V+/+ fibers tended toward smaller LS at all [Ca2+] tested (pCa8 to 5.4). However, at pCa6.2 and 5.8, both WT and KI-TnC-A8V+/+ fibers maintained LS similar to that recorded at pCa8 in each muscle. Interestingly, WT fibers developed ∼15% of the maximal force at pCa5.8 while KI-TnC-A8V+/+ fibers developed ∼70%. Because the lattice spacing vs pCa curves are approximately parallel, these data suggest that at submaximal [Ca2+], skinned fiber force does not strictly determine the distance between thin and thick filaments and that force levels can reach up to 70% of the maximal force without shortening of LS. At higher [Ca2+] (pCa5.6, 5.4) both WT and KI-TnC-A8V+/+ fibers tended toward reduced LS, despite the markedly different levels of force in each muscle. There were no differences in TnI-PSer23,24, MyBPC-Pser282 or mitochondrial content between both muscles. Future experiments will aim at determining whether radial stiffness differs between WT and KI, and measuring kinetics and number of cross-bridges at submaximal [Ca2+].