The concentration of free Ca2+ in the sarcoplasmic reticulum of frog cut twitch skeletal muscle fibers estimated with tetramethylmurexide

One aim of this article was to determine the resting concentration of free Ca2+ in the sarcoplasmic reticulum (SR) of frog cut skeletal muscle fibers ([Ca2+]SR,R) using the calcium absorbance indicator dye tetramethylmurexide (TMX). Another was to determine the ratio of [Ca2+]SR,R to TMX's apparent dissociation constant for Ca2+ (Kapp) in order to establish the capability of monitoring [Ca2+]SR(t) during SR Ca2+ release – a signal needed to determine the Ca2+ permeability of the SR. To reveal the properties of TMX in the SR, the surface membrane was rapidly permeabilized with saponin to rapidly dissipate myoplasmic TMX. Results indicated that the concentration of Ca-free TMX in the SR was 2.8-fold greater than that in the myoplasm apparently due to binding of TMX to sites in the SR. Taking into account that such binding might influence Kapp as well as a dependence of Kapp on TMX concentration, the results indicate an average [Ca2+]SR,R ranging from 0.43 to 1.70 mM. The ratio [Ca2+]SR,R/Kapp averaged 0.256, a relatively low value which should not depend on factors influencing Kapp. As a result, the time course of [Ca2+]SR(t) in response to electrical stimulation is well determined by, and approximately linearly related to, the active TMX absorbance signal.

► The resting concentration of free Ca2+ in the SR of muscle was measured with TMX.
► The time course of [Ca2+] in the SR of skeletal muscle is reliably recorded with TMX.
► The ability to measure [Ca2+]SR should help future studies on Ca signalling in muscle.