Effects of Verapamil and Gadolinium on Caffeine-Induced Contractures and Calcium Fluxes in Frog Slow Skeletal Muscle Fibers

Abstract

In this work, we tested whether L-type Ca(2+ )channels are involved in the increase of caffeine-evoked tension in frog slow muscle fibers. Simultaneous net Ca(2+) fluxes and changes in muscle tension were measured in the presence of caffeine. Isometric tension was recorded by a mechanoelectrical transducer, and net fluxes of Ca(2+) were measured noninvasively using ion-selective vibrating microelectrodes. We show that the timing of changes in net fluxes and muscle tension coincided, suggesting interdependence of the two processes. The effects of Ca(2+)channel blockers (verapamil and gadolinium) were explored using 6 mM: caffeine; both significantly reduced the action of caffeine on tension and on calcium fluxes. Both caffeine-evoked Ca(2+) leak and muscle tension were reduced by 75% in the presence of 100 microM: GdCl(3), which also caused a 92% inhibition of net Ca(2+) fluxes in the steady-state condition. Application of 10 microM: verapamil to the bath led to 30% and 52% reductions in the Ca(2+)leak caused by the presence of caffeine for the peak and steady-state values of net Ca(2+) fluxes, respectively. Verapamil (10 microM): caused a 30% reduction in the maximum values of caffeine-evoked muscle tension. Gd(3+)was a more potent inhibitor than verapamil. In conclusion, L-type Ca(2+) channels appear to play the initial role of trigger in the rather complex mechanism of slow fiber contraction, the latter process being mediated by both positive Ca(2+)-induced Ca(2+ )release and negative (Ca(2+) removal from cytosol) feedback loops.

Effects of Cannabinoids on Caffeine Contractures in Slow and Fast Skeletal Muscle Fibers of the Frog

“…We used caffeine to induce muscle tension since it is well known that caffeine evokes maintained tension in fast and slow fibers, mainly by calcium release from the sarcoplasmic reticulum (Caputo 1966 ; Klein et al 1990 ; Muñiz et al 1992 ). Moreover, in slow fibers Shabala et al ( 2008 ) suggest that caffeine also opens the L-type Ca 2+ channels from the sarcolemma. To investigate the possible direct effect of cannabinoids on skeletal muscle contraction, we induced caffeine contractures in the presence of WIN 55,212-2, causing a diminution in maximum tension and tension-time integral compared to the control.…”

“…Thus, the functional significance of the cannabinoid receptors in skeletal muscle would be the modulation of tension. This force modulation by cannabinoids would be done by acting on the extracellular binding site of the membrane and/or regulating the Ca 2+ release from the sarcoplasmic reticulum (Hoock et al 1996 ; Huerta et al 1986 ; Krippeit-Drews and Schmidt 1992 ; Muñiz et al 1992 ; Shabala et al 2008 ). However, further research is necessary to elucidate these mechanisms.…”

Glibenclamide increases post-fatigue tension in slow skeletal muscle fibers of the chicken

“…To examine whether or not the effects of glibenclamide were mediated by an increased Ca 2+ release from SR, the effect of this drug on contractures induced by caffeine, an alkaloid that mainly induces Ca 2+ release from the SR (Caputo 1966; Huerta and Stefani 1981; Muñiz et al 1992; Shabala et al 2008) was studied. Moreover, in slow muscle fibers caffeine can open L-type Ca 2+ channels in the sarcolemma (Huerta and Stefani 1981; Shabala et al 2008).…”

“…Moreover, in slow muscle fibers caffeine can open L-type Ca 2+ channels in the sarcolemma (Huerta and Stefani 1981; Shabala et al 2008). In caffeine-induced contractures (8 mM), the addition of glibenclamide (150 μM) caused an increase in amplitude of the contracture with respect to the control.…”

“…Caffeine-evoked-tension has been reported to be due to calcium release from the sarcoplasmic reticulum (SR) (Caputo 1966; Klein et al 1990; Huerta and Stefani 1981; Muñiz et al 1992), but in slow fibers, Huerta and Stefani (1981) and Shabala et al (2008) suggest that caffeine also opens the L-type Ca 2+ channels from the sarcolemma, assuring Ca 2+ entry from the external media. Thus, contractures in the ALD muscle were induced by applying 8 mM caffeine.…”