Differences in maximal activation properties of skinned short- and long-sarcomere muscle fibres from the claw of the freshwater crustaceanCherax destructor

Summary

Single fibres of different sarcomere length at rest have been isolated from the claw muscle of the yabby (Cherax destructor), a decapod crustacean. Fibres of either long (SL > 6 μm) or short (SL < 4 μm) sarcomere length have been mechanically skinned and were maximally activated by Ca²⁺ and Sr²⁺ under various experimental conditions (ionic strength, in the presence of 2,3 butanedione monoxime (BDM)) to determine differences in their contractile properties. Isometric force was measured simultaneously with either myofibrillar MgATPase or fibre stiffness in both fibre types. The ultrastructure of individual long- and short-sarcomere fibres was also determined by electron microscopy. The long-sarcomere fibres developed greater tension (30.48±1.72 N cm⁻²) when maximally activated by Ca²⁺ compared with the short-sarcomere fibres (18.60±0.80 N cm⁻²). The difference in the maximum Ca²⁺-activated force can be explained by the difference in the amount of filament overlap between the two fibre types. The maximum Ca²⁺-activated myofibrillar MgATPase rate in the short-sarcomere fibres (1.60±0.27 mmol ATP l⁻¹ s⁻¹) was higher, but not significantly different from the ATPase rate in fibres with long-sarcomeres (1.09±0.14 mmol ATP l⁻¹ s⁻¹). As the concentration of myosin is estimated to be higher only by a factor of 1.22 in the short-sarcomere preparations there is no evidence to suggest that the myofibrillar MgATPase activity is different in the long- and short-sarcomere preparations. The maximum Ca²⁺-activated force (P ₀) of both short- and long-sarcomere fibres was quite insensitive to BDM compared with vertebrate muscle. Force decreased to 60.2±5.3% and 76.1±2.7% in the short- and long-sarcomere fibres respectively in the presence of 100 mmol l⁻¹ BDM. The difference in the force depression between the. long- and short-sarcomere fibres is statistically significant (p<0.05). Fibre stiffness during maximum Ca²⁺-activation expressed as percentage maximum force per nm per half sarcomere was higher by a factor of 3.5 in short-sarcomere fibres than in long-sarcomere fibres suggesting that the compliance of the filaments in the long-sarcomere fibres is considerably higher than in the short-sarcomere fibres. Sr²⁺ could not activate the contractile apparatus to the same level as that seen by Ca²⁺ in either fibre type: the maximum Sr²⁺-activated force was (20±3%) and (63±3%) of the maximum Ca²⁺-activated force response in short- and long-sarcomere fibres, respectively. The ratio between fibre stiffness in the maximum Sr²⁺-activating solution and the Ca²⁺-activating solution was very similar to the ratio between the maximum Sr²⁺-activated force and Ca²⁺-activated force in either type of fibres, suggesting that the number of attached crossbridges is lower in the fibres when maximally activated by Sr²⁺ than when maximally activated by Ca²⁺. The short-sarcomere fibres were also more sensitive to changes in ionic strength than long-sarcomere fibres. In conclusion these results indicate that while several important specific characteristics of the short- and long-sarcomere length fibres (ATPase, maximum Ca²⁺-activated force and fibre stiffness) can be explained solely on differences in the ultrastructure (length and density per cross-sectional area of myosin filaments) there are also differences in the properties of the proteins involved in the force production and regulation evidenced by the differential effect of Sr²⁺, BDM and ionic strength on contractile activation in the two fibre types.