Calcium sensitivity of myosin cross-bridge activation in striated muscles commonly varies during ontogeny and in response to alterations in muscle usage, but the consequences for wholeorganism physiology are not well known. Here we show that the relative abundances of alternatively spliced transcripts of the calcium regulatory protein troponin T (TnT) vary widely in flight muscle of Libellula pulchella dragonflies, and that the mixture of TnT splice variants explains significant portions of the variation in muscle calcium sensitivity, wing-beat frequency, and an index of aerodynamic power output during free flight. Two size-distinguishable morphs differ in their maturational pattern of TnT splicing, yet they show the same relationship between TnT transcript mixture and calcium sensitivity and between calcium sensitivity and aerodynamic power output. This consistency of effect in different developmental and physiological contexts strengthens the hypothesis that TnT isoform variation modulates muscle calcium sensitivity and whole-organism locomotor performance. Modulating muscle power output appears to provide the ecologically important ability to operate at different points along a tradeoff between performance and energetic cost. S triated muscles from a variety of taxa show substantial interand intra-specific variation in the sensitivity of myosin crossbridge activation by calcium (1-6). Within individual animals, calcium sensitivity of muscle activation varies during ontogeny, training, and disease and appears to be one of the primary ways that striated muscles adjust to changes in contractile regimes (7-9). It generally is thought that varying the calcium sensitivity of muscle activation affects recruitment of force-generating cross-bridges during a calcium transient, thereby modulating the rate and amount of force and power output (7-12). However, little is presently known regarding the effects of variation in calcium sensitivity on whole-muscle contractile performance, locomotor ability, and energetics.Variation in muscle calcium sensitivity often involves changes in the molecular composition of the troponin-tropomyosin complex (7-9, 13). This group of molecules constitutes the molecular ''switch'' that turns myosin cross-bridge activity on and off in response to neurally induced calcium signals. Troponin T (TnT), one of the components of the troponin-tropomyosin complex, varies in isoform composition during development (14-17), training (18), and human heart failure (19,20). Changes in TnT isoform composition frequently correlate with variation in the calcium sensitivity of myosin cross-bridge activation (1-9), and experimental manipulations of TnT isoform composition have been shown to affect the calcium sensitivity of actomyosin ATPase (21). Point mutations in human cardiac TnT alter both calcium sensitivity and myosin cross-bridge kinetics (22,23). The emerging picture is that many regions of TnT interact in functionally significant ways with the other troponins, tropomyosin (7-9, 13), and perhaps with m...

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Maximum lift production and the thermal sensitivity of lift production increase dramatically during adult maturation of Libellula pulchella dragonflies. Here, we report that the mechanistic basis for this transition appears to involve a developmental change in protein expression, which alters the Ca2+-sensitivity of muscle activation and twitch contraction kinetics. The alternatively spliced Ca2+ regulatory protein troponin T (TnT) undergoes an isoform shift during adult maturation. Skinned (demembranated) fibers of mature flight muscle are up to 13 times more sensitive to activation by Ca2+ than skinned fibers from teneral (newly emerged adult) flight muscle, and their Ca2+-sensitivity is more strongly affected by temperature. Intact muscle from mature individuals has a shorter time to peak tension and longer time to half-relaxation during twitch contractions, which is consistent with a greater Ca2+-sensitivity of mature muscle. Because it becomes activated more quickly and relaxes more slowly, mature flight muscle is able to generate, with each twitch, more force per unit area than teneral muscle; this difference in force becomes greater at high temperatures. There do not appear to be any age-related differences in actomyosin crossbridge properties, since teneral and mature flight muscles do not differ in shortening velocity, tetanic tension or instantaneous power output during isotonic contraction. Thus, variation in TnT expression appears to affect the temperature-dependent Ca2+-sensitivity of muscle activation, which in turn affects the kinetics and force production of the twitch contractions used by dragonflies during flight. This cascade of effects suggests that maturational changes in the expression of TnT isoforms may be a key determinant of overall muscle and organismal performance.

SUMMARYThe flight muscles of Libellula pulchella dragonflies contain a mixture of six alternatively spliced transcripts of a single troponin T (TnT) gene. Here, we examine how intraspecific variation in the relative abundance of different TnT transcripts affects the Ca2+ sensitivity of skinned muscle fibers and the performance of intact muscles during work-loop contraction regimes that approximate in vivo conditions during flight. The relative abundance of one TnT transcript, or the pooled relative abundance of two TnT transcripts, showed a positive correlation with a 10-fold range of variation in Ca2+ sensitivity of skinned fibers (r2=0.77, P<0.0001) and a threefold range in peak specific force (r2=0.74, P<0.0001), specific work per cycle (r2=0.54; P<0.0001) and maximum specific power output (r2=0.48, P=0.0005) of intact muscle. Using these results to reanalyze previously published data for wing kinematics during free flight, we show that the relative abundances of these particular transcripts are also positively correlated with wingbeat frequency and amplitude. TnT variation alone may be responsible for these effects, or TnT variation may be a marker for changes in a suite of co-regulated molecules. Dragonflies from two ponds separated by 16 km differed significantly in both TnT transcript composition and muscle contractile performance, and within each population there are two distinct morphs that showed different maturational trajectories of TnT transcript composition and muscle contractility. Thus, there is broad intraspecific variability and a high degree of population structure for contractile performance phenotypes, TnT ribotypes and ontogenetic patterns involving these traits that affect locomotor performance.