Relaxation Kinetics Following Sudden Ca2+ Reduction in Single Myofibrils from Skeletal Muscle

Abstract

To investigate the roles of cross-bridge dissociation and cross-bridge-induced thin filament activation in the time course of muscle relaxation, we initiated force relaxation in single myofibrils from skeletal muscles by rapidly (approximately 10 ms) switching from high to low [Ca(2+)] solutions. Full force decay from maximal activation occurs in two phases: a slow one followed by a rapid one. The latter is initiated by sarcomere "give" and dominated by inter-sarcomere dynamics (see the companion paper, Stehle, R., M. Krueger, and G. Pfitzer. 2002. Biophys. J. 83:2152-2161), while the former occurs under nearly isometric conditions and is sensitive to mechanical perturbations. Decreasing the Ca(2+)-activated force preceding the start of relaxation does not increase the rate of the slow isometric phase, suggesting that cycling force-generating cross-bridges do not significantly sustain activation during relaxation. This conclusion is strengthened by the finding that the rate of isometric relaxation from maximum force to any given Ca(2+)-activated force level is similar to that of Ca(2+)-activation from rest to that given force. It is likely, therefore, that the slow rate of force decay in full relaxation simply reflects the rate at which cross-bridges leave force-generating states. Because increasing [P(i)] accelerates relaxation while increasing [MgADP] slows relaxation, both forward and backward transitions of cross-bridges from force-generating to non-force-generating states contribute to muscle relaxation.

Calcium regulation of troponin and its role in the dynamics of contraction and relaxation

“…Similar to, as indicated in de Tombe's paper (see Tables 1 and 2 in Ref. 6), studies on skeletal myofibrils (33), cardiac trabeculae (23), and cardiac myofibrils (29,30,26) revealed, that under all conditions investigated, k act is similar to the rate constant of force redevelopment (k tr ) following a mechanical slack-restretch maneuver during steady Ca 2ϩ activation. This implies that the Ca 2ϩ -induced switch on of the regulatory system occurs before the rise in force and does not rate limit Ca 2ϩ -induced force development.…”

“…Reduced inhibitory function of hcTnI by the R145G mutation and likewise by the truncation of hcTnI to hcTnI1-192, the predominant degradation product in the stunned myocardium, inevitably slows down relaxation kinetics manifesting in a delayed onset and decreased rate constant of the fast relaxation phase (12,22). That these two relaxation parameters are very sensitively affected by only slight, persistent activation is in agreement with the similar effects in native myofibrils following a slight incomplete Ca 2ϩ removal (33).…”

“…However, it remains unclear in which order these events should be reversed during relaxation. The rate constant of the initial, slow myofibrillar force decay (k rel,slow ) following Ca 2ϩ removal suggests that no force-generating cross-bridges become newly formed after Ca 2ϩ removal (26,27,29,30,32,33). This is corroborated by the findings of de Tombe et al (6) that k rel,slow and t rel,slow are insensitive to interventions that decrease the off rate of Tn.…”

“…Comparison of the kinetics of the slow force decay (k rel,slow ) with force development kinetics suggests that k rel,slow represents the turnover rate by which cross bridges leave forcegenerating states (27,29,33). The isometric, slow-relaxation phase lasts until the moment when the first, mechanically weakest, sarcomere in myofibril elongates.…”

“…This perturbs the strain of cross-bridges in all sarcomeres, thereby catalyzing their detachment from actin and accelerating the force decay (29). This complex process of muscle relaxation is independent of the level of thin-filament activation prior to Ca 2ϩ removal, but rather depends on the final [Ca 2ϩ ] after rapid solution switch (27,33). As de Tombe's results indicate, the different TnI isoforms do not alter the observed rate by which cross bridges leave force-generating states under isometric conditions (k rel,slow ) and the strain-induced accelerated cross-bridge detachment rates by sarcomere dynamics (k rel,fast ).…”