Effects of extracellular calcium on calcium movements of excitation‐contraction coupling in frog skeletal muscle fibres.

1. Calcium transients were measured in fast-twitch rat skeletal muscle fibres stretched to 3 7-4 0 #sm per sarcomere, and voltage clamped at a holding potential of -80 mV using the double-seal Vaseline gap technique. Resting calcium was monitored with fura-2 and the calcium transients were measured with antipyrylazo III. The rate of release of calcium from the sarcoplasmic reticulum was calculated from the calcium transient records. The temperature was 14-17 'C.2. The steady-state calcium dependence of inactivation of release was studied with a two-pulse protocol in which 200 ms prepulses of different amplitudes elevated the internal calcium concentration to various levels. The inactivation of release was then measured in the test pulse that followed the prepulses. The calcium concentration at which the inactivation of release was half-maximal was -0-22 uM, the average number of bound calcium ions needed to cause inactivation was about three per release channel and the amount of release that could be inactivated was, on average, 2-48 times the steady level of release during the test pulses. 3. Procaine (03 mM) reversibly decreased the amplitude and the rate of rise of the calcium transient. Both the peak and the steady level of release were decreased by about 50%. The shape of the release waveform was not modified.

Section: Methodsmentioning

confidence: 99%

Suppression of calcium release by calcium or procaine in voltage clamped rat skeletal muscle fibres.

Garcı́a

Schneider

1995

“…Details of the technique for recording charge movements and calcium transients, as well as for pulse generation and data acquisition, have been given elsewhere (Kovacs, Rios & Schneider, 1983;Brum & Rios, 1987;. Calcium release flux was calculated from the recorded calcium transients by the method of Melzer, Rios & Schneider (1987) modified by Brum, Rios & Schneider (1988). Release flux records were corrected for depletion of calcium in the sarcoplasmic reticulum (SR) (Schneider, Simon & SzUcs, 1987).…”

Section: Methodsmentioning

confidence: 99%

Differential effects of tetracaine on two kinetic components of calcium release in frog skeletal muscle fibres.

Pizarro

Csernoch

Uribe

et al. 1992

Self Cite

SUMMARY1. Intramembrane charge movements and changes in intracellular calcium concentration were recorded simultaneously in voltage clamped cut skeletal muscle fibres of the frog in the presence and absence of tetracaine.2. Extracellular application of 20 /IM tetracaine reduced the increase in myoplasmic [Ca2"]. The effect on the underlying calcium release flux from the sarcoplasmic reticulum was to suppress the peak of the release while sparing the steady level attained at the end of 100 ms clamp depolarizations.3. While the peak of the release flux at corresponding voltages was reduced by 62 o after the addition of tetracaine, the rate of inactivation was the same when the pulses elicited release fluxes of similar amplitude.4. Higher concentrations of tetracaine, 0-2 mm, abolished the calcium signal in stretched fibres whereas in slack fibres this concentration left a non-inactivating calcium release flux.5. Lowering the extracellular pH antagonized the effect of the drug both on charge movements and on calcium signals. The permanently charged analogue tetracaine methobromide lacked effects on excitation-contraction coupling.6. These results imply that the two kinetic components of calcium release flux have very different tetracaine sensitivities. They are also consistent with an intracellular site of action of the drug at low concentration. Taken together they strongly suggest that the inactivating and non-inactivating components of calcium release correspond to different pathways: one that inactivates, is sensitive to tetracaine and is controlled by calcium, and another that does not inactivate, is much less sensitive to tetracaine and is directly controlled by voltage.

“…Troponin C sites were assumed to be present at 250 /M (cf. Baylor, Chandler & Marshall, 1983) and to have 'on' and 'off' rate constants of 1-3 x 108 M-1 s-1 and 103 s-1 Brum, Rios & Stefani, 1988b) in all fibres. The parvalbumin calcium/magnesium sites constitute the major group of slowly equilibrating sites.…”

Section: General Proceduresmentioning

confidence: 99%

“…The parvalbumin calcium/magnesium sites constitute the major group of slowly equilibrating sites. The 'on' rate constants for binding of calcium and of magnesium to the parvalbumin sites were assumed to be 1-6 x 108 M-1 s-1 and 4 x 104 M-1 s-1, respectively (Baylor et al 1983;Brum et al 1988b). The concentration of parvalbumin calcium/magnesium sites and the off rate constants for dissociation of calcium and of magnesium from parvalbumin were determined by a least-squares fit of the removal model prediction to the observed decay of A[Ca2+] after various pulses.…”

Section: General Proceduresmentioning

confidence: 99%

Effects of caffeine on calcium release from the sarcoplasmic reticulum in frog skeletal muscle fibres.

Klein

Simon

Schneider

1990

3. The voltage dependence of the peak value ofRrei and of the steady level ofRrei at the end of a 60-120 ms pulse were both shifted towards more negative voltages in caffeine. For relatively small pulses the voltage at which a given release waveform was observed was also shifted to more negative voltages.4. Intramembrane charge movements measured in the same fibres in which the above changes in Rrei were observed showed no significant changes in caffeine.5. In caffeine calcium release continued for many milliseconds after the end of a short (10 ms) pulse. Continued release after a pulse was not observed without caffeine and was probably due to positive feedback of elevated [Ca2+] on calcium release resulting from calcium-induced calcium release in caffeine.6. Intramembrane charge movements after short pulses showed no change in caffeine that could account for the continued calcium release after the pulse.7. Continued release after short pulses in caffeine decreased as the pulse duration was increased and was absent for pulses of 60 ms or longer. Rre, also inactivated during such pulses. 8. Relatively large and long conditioning pulses in caffeine suppressed both the peak Rrei and the continued release after short pulses. Peak release and continued release after short pulses recovered in parallel with increasing recovery time following suppression by a conditioning pulse in caffeine. 9. These results indicate that in the presence of caffeine, charge movement and calcium-induced calcium release both contribute significantly to the activation of