Calcium Dependence of Titin-Regulated Passive Forces in Skeletal Muscle Fibers

Abstract

There is evidence that the passive forces produced by titin in skeletal muscles may be regulated by Ca2+. Studies have shown an upwards shift in the passive force-sarcomere length (SL) relation when muscle fibres are tested with a high concentration of Ca2+ and the myosin-actin interaction is abolished. In this study we tested the hypothesis that there is a direct relation between the concentrations of Ca2+ and the cross-bridge independent increase in passive forces. Single fibres were isolated from the rabbit psoas muscle and transferred into an experimental chamber, between a force transducer and a motor arm. Fibres were activated in a range of Ca2+ concentrations (pCa2+ between 4.5 and 9.0), before and after administration of the myosin inhibitor blebbistatin, which caused the maximal isometric force to decrease by 93.5%. After blebbistatin administration, the fibres were submitted to a protocol in which they underwent consecutive step-stretches, starting at an initial SL of 2.5μm (amplitude of stretch: 5% initial SL, duration 300 ms, pauses between stretches: 30 sec). We observed an upwards shift in the passive force-SL relation when pCa2+ was lower than 6.4 (increase from 1.06 mN/mm2 to 4.28 mN/mm2 at SL of 2.8μm and from 14.99 mN/mm2 to 27.85 mN/mm2 at SL of 3.8μm). Decreasing the pCa2+ below 6.4 did not change the forces further. The results suggest that there is not a direct relation between passive forces and Ca2+ concentrations; instead there is a threshold for Ca2+ regulation of passive forces via titin (in the condition investigated, a pCa2+ of 6.4), beyond which force does not change significantly.