Mechanism of contracture on cooling of caffeine-treated frog skeletal muscle fibres

Abstract

1. In order to clarify the mechanism of contracture on cooling of caffeine-treated intact muscle fibres, the temperature dependence of a calcium (Ca2+) release mechanism, 'Ca2+-induced Ca2+ release', of the sarcoplasmic reticulum (SR) was examined in skinned frog muscle fibres. 2. Skinned fibres in a solution containing 1.2 mM-caffeine and 0.7 mM-EGTA (Mg2+, 1.5 mM, Mg-ATP, 3.5 mM, pH 7), contracted on cooling (from 22 to 2 degrees C) due to Ca2+ release from the SR. 3. The rate of Ca2+ release from skinned fibre SR in a medium which contained Ca2+ ions (with 10 mM-EGTA) and no ATP salts, was determined under various conditions using the 'caffeine method.' 4. In the absence of Mg2+ ions, adenine nucleotides and caffeine, the rates at room temperature (21-22 degrees C) were 3-4 times greater than those at a lower temperature (1.5-3 degrees C), at any concentrations of Ca2+ ions external to the SR. 5. In the presence of Mg2+ ions (1.5 mM) and beta,gamma-methylene ATP (1 mM), the effect of temperature on the rates disappeared in Ca2+-containing media, although the effect remained in Ca2+-free medium. 6. When caffeine (1.2 mM), which is a potentiator of the Ca2+-induced Ca2+ release, was added to the test medium with Mg2+ and beta,gamma-methylene ATP, the resulting potentiating effect was several times greater than that at lower temperature. 7. In order to examine the temperature dependence of the Ca2+ pump activity of the SR, the initial rate of Ca2+ uptake by the empty SR was determined under various conditions in the presence of Mg2+ ions (1.5 mM) and Mg-ATP (3.5 mM). The Q10 of the pump activity was around 2.0 at the Ca2+ ion concentrations examined (less than 10(-6) M). 8. A numerical model based on the results obtained, together with some reasonable assumptions, suggested that both suppression of the Ca2+ pump and enhancement of the Ca2+ release contribute to the cooling contracture of caffeinized fibres.