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Sarcoplasmic reticulum calcium release in frog skeletal muscle fibres estimated from Arsenazo III calcium transients


Sarcoplasmic reticulum calcium release in frog skeletal muscle fibres estimated from Arsenazo III calcium transients



Journal of Physiology 344: 625-666



ISSN/ISBN: 0022-3751

PMID: 6655593

DOI: 10.1113/jphysiol.1983.sp014959

Single twitch fibers, dissected from frog muscle, were injected with the metallochromic dye Arsenazo III. Changes in dye-related absorbance measured at 650 or 660 nm were used to estimate the time course of myoplasmic free [Ca2+] following either action potential stimation or voltage-clamp depolarization (temperature, 15.degree.-17.degree. C). Computer simulations were carried out to estimate the flux of Ca2+ between the sr [sarcoplasmic reticulum] and myoplasm (in fibers containing no more than 0.8 mM-dye). Following action potential stimulation 0.2-0.3 mM-Ca2+ enters the myoplasm from the sr. The wave form of sr Ca2+ release is early and brief compared with the wave form of free [Ca2+]. Neither the selection of troponin rate constants nor the inclusion of parvalbumin has much effect on the shape of the release wave form; the main effect of varying these parameters is to change the magnitude. After the initial, rapid phase of Ca2+ release from the sr there is a longer, maintained period of Ca2+ uptake. The peak magnitude of this uptake divided by the concentration of sr Ca2+ pump sites is used to estimate the turnover rate of the sites. The numbers so obtained, 2.4-6.3 s-1, are in good agreement with previoulsy published values based on biochemical experiments. The known biochemical properties of the sr Ca2+ pump can explain the final phase of the computed sr Ca2+ flux and therefore, within the framework of the model, the falling phase of the free [Ca2+] transient. Simulations, based on Ca2+ transients obtained during a train of action potentials spaced 10 ms apart, indicate that sr Ca2+ release associated with a 2nd or subsequent action potential is no more than 20-30% of the release associated with the 1st action potential. Similarly, simulations based on Ca2+ transients measured in voltage-clamp experiments show that during a step depolarization near threshold the sr Ca2+ release reaches an early peak which is followed by a decline to a smaller level. These observations of reduced Ca2+ release appear to be more consistent with a decrease in sr Ca2+ permeability than with a decrease in driving force on the ion. The simulations show that the presence of intracellular binding sites for Ca2+ makes the estimate of the voltage dependence of sr CA2+ release less steep than that observed for the peak of the Ca2+ transient. For example, two experiments which showed a 3 mV e-fold change in peak free [Ca2+] required a 4 mV e-fold change for the peak of Ca2+ release. Inferences about Ca2+ movements between sr and myoplasm from measurements of myoplasmic free [Ca2+] are aided by the use of a quantitative model that takes into account the kinetic and steady-state properties of the known Ca2+-binding sites in myoplasm.

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