Abstract 1627: Increasing the Rate of Cardiac Contraction by High Calcium Binding Affinity Troponin C Mutant
Calcium (Ca2+) binding to cardiac troponin C (cTnC) triggers structural changes in the thin filament that allow myosin crossbridges to bind to actin and generate force. Cardiac muscle contraction, on a beat-to-beat basis, is modulated by the interplay between the thin and thick filament activation. We assessed the hypothesis that direct modulation of thin filament Ca2+ sensitivity can alter the rate of contraction at different levels of Ca2+ activation. We reconstituted genetically engineered cTnC mutants with high and low Ca2+-binding affinity in the troponin complex, V44Q cTnCF27W and F20Q cTnCF27W respectively, into rat skinned cardiac trabeculae at 15°C. All recombinant cTnCs had fluorescent F27W substitution to measure Ca2+ affinity in solution. When force reached a plateau of activation, crossbridges were mechanically disrupted by a rapid slack (20% of muscle length)-restretch protocol, which allowed crossbridges to re-attach and redevelop force. Kinetics of contraction is assessed as the rate of tension redevelopment (Ktr) as fitted with a monoexponential relationship. For endogenous cTnC, control cTnCF27W, V44Q cTnCF27W and F20Q cTnCF27W, Ktr values were not significantly different when muscle was maximally activated in pCa 4.0 (9.3/s ± 0.83, 10.53/s ± 1.27, 10.51/s ± 0.88, 8.97/s ± 0.75, n=6, respectively). When the muscle was generating between 10 – 60% of maximal force, Ktr was significantly faster for the high Ca2+-binding affinity V44Q cTnCF27W compared to control cTnCF27W (at pCa 6.2: 3.61/s ± 0.23 vs. 1.98/s ± 1.91, at pCa 6.0: 5.51/s ± 0.67 vs. 2.41/s ± 0.2 and at pCa 5.8: 8.83/s ± 0.97 vs. 4.94/s ± 0.52, n=6). For the low Ca2+-binding affinity F20Q cTnCF27W, Ktr was significantly slower compared to V44Q cTnCF27W but similar to control. In conclusion, we showed that increasing the thin filament Ca2+ sensitivity using cTnC with high Ca2+ binding affinitiy could speed up the rate of contraction. That this effect is prevalent only at submaximal levels of activation may have important physiological implications for contraction of the heart in vivo: it might be possible to improve systolic dysfunction in heart failure by increasing the Ca2+-binding affinity of cTnC.