Abstract
Enhanced cardiac contractile function with increased sarcomere length (SL) is, in part, mediated by a decrease in the radial distance between myosin heads and actin. The radial disposition of myosin heads relative to actin is modulated by cardiac myosin binding protein-C (cMyBP-C), suggesting that cMyBP-C contributes to the length-dependent activation (LDA) in the myocardium. However, the precise roles of cMyBP-C in modulating cardiac LDA are unclear. To determine the impact of cMyBP-C on LDA, we measured isometric force, myofilament Ca2+-sensitivity (pCa50) and length-dependent changes in kinetic parameters of cross-bridge (XB) relaxation (krel), and recruitment (kdf) due to rapid stretch, as well as the rate of force redevelopment (ktr) in response to a large slack-restretch maneuver in skinned ventricular multicellular preparations isolated from the hearts of wild-type (WT) and cMyBP-C knockout (KO) mice, at SL's 1.9 μm or 2.1 μm. Our results show that maximal force was not significantly different between KO and WT preparations but length-dependent increase in pCa50 was attenuated in the KO preparations. pCa50 was not significantly different between WT and KO preparations at long SL (5.82 ± 0.02 in WT vs. 5.87 ± 0.02 in KO), whereas pCa50 was significantly different between WT and KO preparations at short SL (5.71 ± 0.02 in WT vs. 5.80 ± 0.01 in KO; p < 0.05). The ktr, measured at half-maximal Ca2+-activation, was significantly accelerated at short SL in WT preparations (8.74 ± 0.56 s−1 at 1.9 μm vs. 5.71 ± 0.40 s−1 at 2.1 μm, p < 0.05). Furthermore, krel and kdf were accelerated by 32% and 50%, respectively at short SL in WT preparations. In contrast, ktr was not altered by changes in SL in KO preparations (8.03 ± 0.54 s−1 at 1.9 μm vs. 8.90 ± 0.37 s−1 at 2.1 μm). Similarly, KO preparations did not exhibit length-dependent changes in krel and kdf. Collectively, our data implicate cMyBP-C as an important regulator of LDA via its impact on dynamic XB behavior due to changes in SL.
Highlights
Length-dependent activation (LDA) is the mechanism by which force production in the heart becomes more sensitive to Ca2+ as the sarcomere length (SL) is increased (Allen and Kentish, 1985)
SDS gels loaded with ventricular samples from WT and KO hearts were stained with Coomassie blue or Pro-Q Diamond stain to assess the effects of cardiac myosin binding protein-C (cMyBP-C) KO on sarcomeric protein isoform expression and phosphorylation levels (Figures 2B,C, respectively)
P1 was significantly decreased (P = 0.032) at long SL in KO compared to WT (Figure 6; Table 2). These results suggest that a decrease in the muscle fiber stiffness contributed, at least in part, to the acceleration of krel observed at short SL compared to long SL in WT, and at long SL in KO compared to long SL in WT (Figure 5A)
Summary
Length-dependent activation (LDA) is the mechanism by which force production in the heart becomes more sensitive to Ca2+ as the sarcomere length (SL) is increased (Allen and Kentish, 1985). The cooperative effect between neighboring troponin-tropomyosin (Tn-Tm) complexes impacts the Ca2+ binding properties of the thin-filament (Butters et al, 1997; Farman et al, 2010) and influence the LDA in cardiac muscle (for details on LDA refer to reviews by Konhilas et al, 2002; Hanft et al, 2008; De Tombe et al, 2010; Campbell, 2011)
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