Isometric force redevelopment of skinned muscle fibers from rabbit activated with and without Ca2+

Abstract

Fiber isometric tension redevelopment rate (kTR) was measured during submaximal and maximal activations in glycerinated fibers from rabbit psoas muscle. In fibers either containing endogenous skeletal troponin C (sTnC) or reconstituted with either purified cardiac troponin C (cTnC) or sTnC, graded activation was achieved by varying [Ca2+]. Some fibers were first partially, then fully, reconstituted with a modified form of cTnC (aTnC) that enables active force generation and shortening in the absence of Ca2+. kTR was derived from the half-time of tension redevelopment. In control fibers with endogenous sTnC, kTR increased nonlinearly with [Ca2+], and maximal kTR was 15.3 +/- 3.6 s-1 (mean +/- SD; n = 26 determinations on 25 fibers) at pCa 4.0. During submaximal activations by Ca2+, kTR in cTnC reconstituted fibers was approximately threefold faster than control, despite the lower (60%) maximum Ca(2+)-activated force after reconstitution. To obtain submaximal force with aTnC, eight fibers were treated to fully extract endogenous sTnC, then reconstituted with a mixture of a TnC and cTnC (aTnC:cTnC molar ratio 1:8.5). A second extraction selectively removed cTnC. In such fibers containing aTnC only, neither force nor kTR was affected by changes in [Ca2+]. Force was 22 +/- 7% of maximum control (mean +/- SD; n = 15) at pCa 9.2 vs. 24 +/- 8% (mean +/- SD; n = 8) at pCa 4.0, whereas kTR was 98 +/- 14% of maximum control (mean +/- SD; n = 15) at pCa 9.2 vs. 96 +/- 15% (mean +/- SD; n = 8) at pCa 4.0. Maximal reconstitution of fibers with aTnC alone increased force at pCa 9.2 to 69 +/- 5% of maximum control (mean + SD; n = 22 determinations on 13 fibers) and caused a small but significant reduction of kTR to 78 +/- 8% of maximum control (mean +/- SD; n = 22 determinations on 13 fibers); neither force nor krR was significantly affected by Ca>2(pCa 4.0). Taken together, we interpret our results to indicate that kTR reflects the dynamics of activation of individual thin filament regulatory units and that modulation of kTR by Ca> is effected primarily by Ca>+ binding to TnC.

Contractile properties of muscle fibers from the deep and superficial digital flexors of horses

“…Freshly dissected muscle fibers from the DDF, SDF, and SOL muscles were prepared for mechanical experimentation using published methods (11,12). Fiber fascicles were treated with a skinning solution containing 0.5% Brij-58 detergent (Pierce Ultrapure; Pierce Biotechnology, Rockford, IL) for 1 h on ice and then glycerinated with a 50% glycerol-skinning solution and stored at Ϫ20°C.…”

“…Mechanical experiments on mammalian muscle fibers have most often been performed on skinned (permeabilized) fibers from rabbits (11)(12)(13)35) and even smaller mammals, such as rats and mice (2,3), to determine fundamental measurements of physiological capacity [e.g., maximum unloaded shortening velocity (V 0 ) or velocity of unloaded shortening (V US )]. A thorough investigation of DDF and SDF contractile properties using single, skinned fibers would provide a valuable contribution toward understanding each muscle's functional capacity for contraction velocity and thus work and power performance, as suggested by their respective architectures.…”

Basal myosin light chain phosphorylation is a determinant of Ca2+ sensitivity of force and activation dependence of the kinetics of myocardial force development

“…ktr was determined by linear transformation of the half-time (t1/2) of force recovery [ktr ϭ Ϫln 0.5 ϫ (t1/2) Ϫ1 ], as described previously (7,38).…”

Modulation of the rate of cardiac muscle contraction by troponin C constructs with various calcium binding affinities

“…One approach has been developed by Brenner (3, 4). According to this protocol, a rapid shorteningrestretch maneuver mechanically detaches the cross bridges, and the rate at which the cross bridges reattach and generate force is a measure of the rate of contraction, known as the rate of force (tension) redevelopment (k tr ).In both cardiac and skeletal muscle, it is well established that k tr becomes faster with increasing levels of activation by Ca 2ϩ (5,6,13,28,46). Many studies have investigated the role Ca 2ϩ plays in the activation dependence of k tr (for review, see Ref.…”

“…In both cardiac and skeletal muscle, it is well established that k tr becomes faster with increasing levels of activation by Ca 2ϩ (5,6,13,28,46). Many studies have investigated the role Ca 2ϩ plays in the activation dependence of k tr (for review, see Ref.…”