The Role of Thin Filament Cooperativity in Cardiac Length-Dependent Calcium Activation

Abstract

Length-dependent activation (LDA) is a prominent feature of cardiac muscle characterized by decreases in the Ca(2+) levels required to generate force (i.e., increases in Ca(2+) sensitivity) when muscle is stretched. Previous studies have concluded that LDA originates from the increased ability of (strong) cross-bridges to attach when muscle is lengthened, which in turn enhances Ca(2+) binding to the troponin C (TnC) subunit of the troponin complex. However, our results demonstrate that inhibition of strong cross-bridge attachment with blebbistatin had no effect on the length-dependent modulation of Ca(2+) sensitivity (i.e., EC(50)) or Ca(2+) cooperativity, suggesting that LDA originates upstream of cross-bridge attachment. To test whether LDA arises from length dependence of thin-filament activation, we replaced native cTnC with a mutant cTnC (DM-TnC) that is incapable of binding Ca(2+). Although progressive replacement of native cTnC with DM-TnC caused an expected monotonic decrease in the maximal force (F(max)), DM-TnC incorporation induced much larger increases in EC(50) and decreases in Ca(2+) cooperativity at short lengths than at long lengths. These findings support the conclusion that LDA arises primarily from the influence of length on the modulation of the Ca(2+) cooperativity arising from interaction between adjacent troponin-tropomyosin complexes on the thin filament.

Historical perspective on heart function: the Frank–Starling Law

“…They observed that blebbistatin (which prevents strong-binding formation) had no effect on the Hill coefficient (Sun et al 2009). Farman et al (2010) reached similar conclusions, because, in their experiments, blebbistatin decreased Ca 2+ sensitivity and force, but did not affect the Hill coefficient. Importantly, the authors reconstituted rat cardiac thin filaments with a cTnC mutant incapable of binding Ca 2+ and observed that both Ca 2+ sensitivity and nH were decreased (Farman et al 2010).…”

“…Farman et al (2010) reached similar conclusions, because, in their experiments, blebbistatin decreased Ca 2+ sensitivity and force, but did not affect the Hill coefficient. Importantly, the authors reconstituted rat cardiac thin filaments with a cTnC mutant incapable of binding Ca 2+ and observed that both Ca 2+ sensitivity and nH were decreased (Farman et al 2010). In addition, they observed that the effects of the cTnC mutant were greater at short (2.0 μm) than at longer (2.2 μm) sarcomere lengths.…”

“…Two recent studies support the idea that Ca 2+ cooperative effects are independent of myosin-binding and are strongly associated with the thin filaments (Sun et al 2009;Farman et al 2010). Sun et al (2009) reconstituted cardiac thin filaments with fluorescent-labeled TnC and analyzed changes in the orientation of the structure of troponin.…”

Effects of aging, exercise, and disease on force transfer in skeletal muscle

“…It would therefore appear that titin does not affect muscle contraction by directly altering the sarcomeric structure. Other mechanisms suggested within the cardiac literature have ranged from altering the Ca 2ϩ affinity of troponin, interfilament spacing, myosin structure, and thin filament activation (23,27). The last possibility may reflect the fact that titin, within the thick filament, could affect nebulin, within the thin filament, and alter muscle contraction and subsequent longitudinal force transfer (see below).…”

Electromechanical coupling in the cardiac myocyte; stretch-arrhythmia feedback

“…Furthermore, the sensitivity of the contractile response, at submaximal [Ca 2+ ] i levels, is strikingly sarcomere length dependent: a phenomenon known as length-dependent activation (LDA) [24,34,45,75]. This remarkable phenomenon is found in all striated muscle but is stronger in cardiac muscle than in fast skeletal muscle, which again is more length dependent than slow skeletal muscle [47].…”