Title page for ETD etd-06062008-151126

Type of Document Dissertation
Author Ward, Christopher W.
URN etd-06062008-151126
Title The role of the apparent rate constant of cross-bridge transition from the strong binding state (g{u0333}a{u0333}p{u0333}p) in skeletal muscle force production
Degree PhD
Department Veterinary Medical Sciences
Advisory Committee
Advisor Name Title
Lee, John C. Committee Co-Chair
Williams, Jay H. Committee Co-Chair
Eng, Ludeman A. Committee Member
Klug, Gary A. Committee Member
McGrath, Charles J. Committee Member
  • muscle
  • cross-bridges
  • calcium
  • contraction
  • myosin
Date of Defense 1996-07-01
Availability unrestricted
Force regulation at the level of the actin-myosin cross-bridge (XB) can be described by a 2 state model in which the XB's cycle between a strongly bound (SB), force generating state and a weakly bound (WB), non-force generating state. This cycle can be characterized by the apparent rate constants for transition into the SB state (ƒapp) and returning to the WB state (gapp). Increases in XB force can be accounted for by an increase in ƒapp a decrease in gapp or both. While effort towards understanding XB force regulation has focused on the notion that force production is primarily regulated by ƒapp the purpose of this investigation was to determine if gapp contributes to force regulation at the XB and to determine whether gapp differs in, muscles with differing contractile characteristics.

Specifically, gapp is an apparent rate constant which represents the sum of the biochemical rates which characterize the transition from the SB state to the WB state. Estimates of gapp were experimentally derived by determining the ratio of myosin ATPase activity to force across a physiological range of calcium (Ca2+) in skinned muscle fibers. Insights into g., were gained by comparing the [Ca2+]50 (concentration of Ca1+ required to elicit 50% of maximal force or ATPase activity) of ATPase activity and force. For example, if g., contributes to XB force regulation it would be expected that gapp would be sensitive to increasing [Ca2+] which would manifest in differing Ca1+ sensitivities between ATPase and force.

Measurements of [Ca2+]50 for force and ATPase in fast fibers from the frog (20°C 2.0mM [MgATP]) were significantly different (2.64±0.14 vs. 1.58±0.08 μM Ca2+ p<.05). The separation of these curves resulted in a decreasing of A/F by nearly four fold over submaximal levels of Ca2+. At maximal Ca2+ activation gapp was 2.90 s-1.

To further explore the role of gapp in XB cycling, measurements of AIF were made at reduced [MgATP](l.0, 0.5, 0.25 mM) in frog fibers. Decreased [MgATP] would be expected to slow the SB to WB transition (breaking of the rigor bond) thus reducing g.,. Reduced (MgATP] resulted in progressively smaller differences in the [Ca2+ so of force and ATPase. This lead to a reduction in the Ca2+ sensitivity of gapp.

Fast fibers are known to be less sensitive to Ca2+ ([Ca2+]50) and contract with faster kinetics compared to slow fibers. Since differences in gapp should exist between muscles with differing contractile characteristics, gapp was examined between soleus (slow) and extensor digitorum longus (EDL, fast) fibers of the rat While no differences in Fmax (maximal Ca2+ activated force) were seen between the fiber types, differences in A/F ratios were seen between both fiber types. With increasing Ca2+ activation, this estimate of gapp shows a 5.5-fold decrease in EDL fibers and a 2.5-fold decrease in SOL fibers. At maximal Ca2+ activation gapp was 3.13±0.35s·1 and l.50±0.04s·1 for fast and slow fibers respectively.

To determine to what extent differences in gapp account for the differences in the [Ca2+]50 between fast and slow fibers, reductions in [MgATP] to 0.5 mM were used. Slow fibers exhibited no differences in the [Ca2+]50 between force and ATPase. In fast fibers, [Ca2+]so of force was different at 2.0 mM, this difference was abolished at 0.5 mM [MgATP]. This suggests that differences in calcium sensitivity of force between fast and slow muscle are due in part to gapp.

Based on the findings that gapp is calcium sensitive and that altering gapp can alter contractile kinetics (i.e., [Ca2+]50 so for force) it appears that gapp plays a role in the modulation of contractile characteristics at the skeletal muscle XB.

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