Initiation of active contraction by photogeneration of adenosine-5'-triphosphate in rabbit psoas muscle fibres.

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RESUMO

Mechanical and biochemical descriptions of the muscle cross-bridge cycle have been correlated. Skinned muscle fibres of rabbit psoas muscle in rigor were incubated in solutions containing approximately equal to 30 microM-Ca2+ ions and P3-1-(2-nitro)phenylethyladenosine-5'-triphosphate, 'caged ATP', an inert photolabile precursor of ATP. ATP was liberated from caged ATP within the fibres by pulses of 347 nm radiation from a frequency-doubled ruby laser. The mechanical responses of muscle fibres to the rapid increase of ATP concentration were monitored. Tension dropped briefly and then rose above the rigor value to the level characteristic of a steady active contraction. Liberation of ATP decreased in-phase stiffness (measured at 500 Hz) from the rigor level to a maintained value intermediate between rigor and relaxed values. Out-of-phase stiffness increased to a maintained level indicating a phase lead of tension with respect to imposed length oscillations. Rigor tension was varied prior to photolysis by slight alterations of fibre length. Tension traces starting at different rigor tensions converged to a common tension level at the same rate, whether or not Ca2+ was included in the medium. These data suggest that the rate of cross-bridge detachment by ATP from the rigor state is not influenced by Ca2+. Analysis of the tension records, in terms of sequential detachment and reattachment reactions, provided a measure of cross-bridge reattachment rate and an alternate measure of the detachment rate. Detachment from the rigor state was approximately proportional to the ATP concentration, with a second-order rate constant of at least 5 X 10(5) M-1 S-1. Reattachment with force generation had no detectable dependence on the concentration of ATP liberated by photolysis. A simple kinetic model of the cross-bridge cycle in terms of chemically defined intermediates was compatible with most of the experimental data. The ATP dependence of cross-bridge detachment, the kinetics of maintained cross-bridge reattachment in the presence of Ca2+, and transient reattachment and final relaxation in the absence of Ca2+ were explained. In this model, reversibility of cross-bridge attachment and the steps leading to force production allow the relatively high observed detachment rate to be accommodated with other data relating to active contraction. These data include the steady ATPase rate of active muscle fibres and the fewer attached cross-bridges in active contractions compared to rigor.

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