Calcium channel inactivation in frog (Rana pipiens and Rana moctezuma) skeletal muscle fibres (original) (raw)

Fast gating kinetics of the slow Ca2+ current in cut skeletal muscle fibres of the frog

The Journal of physiology, 1990

1. Calcium currents and intramembrane charge movements were measured in cut twitch muscle fibres of the frog and the time course of activation of the current was studied using various conditioning pulse protocols. 2. When a conditioning activation was produced by a depolarizing pulse which ended before inactivation occurred, a subsequent depolarization led to a faster onset of activation, indicating that the system had not completely returned to the initial state during the interval between the two pulses. 3. The interval between conditioning and test pulse was varied at different subthreshold potentials to study the time course of restoring the steady-state conditions. Complete restoration required a waiting period of about 1 min at the holding potential of -80 mV due to a very slow process but partial recovery was reached within 100 ms. This initial recovery process was strongly voltage dependent and became considerably slower when the interval potential approached the threshold f...

Kinetic properties of calcium channels of twitch muscle fibres of the frog

The Journal of physiology, 1983

Calcium currents (ICa) were recorded in frog skeletal muscle fibres using the three-micro-electrode voltage-clamp technique. The sartorius muscle was bathed in TEA methanesulphonate saline with 350 mM-sucrose. 5 mM-3,4-diaminopyridine was added to the saline to minimize K+ currents. The I-V relationship for peak Ca2+ currents showed that ICa was detected at -40 mV and reached a maximum value at ca. -10 mV. No net inward current was recorded at potentials positive to ca. +40 mV. Remaining K+ currents (IK) were recorded by replacing 10 mM-Ca2+ with 5.5 mM-Co2+. They were not noticeably time-dependent up to +20 mV and would tend to diminish the amplitude of ICa without greatly affecting its time course. ICa tail currents could be separated from non-linear capacity currents. Tail currents were measured 5 msec after repolarization and extrapolated to the end of the pulse. ICa tail-current amplitudes at EK were measured with pulses of different durations. The envelope of tail-current ampl...

Decay of the slow calcium current in twitch muscle fibers of the frog is influenced by intracellular EGTA

The Journal of General Physiology, 1989

The mechanism(s) of the decay of slow calcium current (/ca) in cut twitch skeletal muscle fibers of the frog were studied in voltage-clamp experiments using the double vaseline-gap technique./ca decay followed a single exponential in 10 mM external Ca 2 § and 20 mM internal EGTA solutions in all pulse protocols tested: single depolarizing pulses (activation protocol), two pulses (inactivation protocol), and during a long pulse preceded by a short prepulse (400 ms) to 80 mV (tail protocol). In single pulses the rate constant oflc~ decay was ~0.75 s -~ at 0 mV and became faster with larger depolarizations. Ic~ had different amplitudes during the second pulses of the inactivation protocol (0 mV) and of the tail protocol (-20 to 40 mV) and had similar time constants of decay. The time constant of decay did not change significantly at each potential after replacing 10 mM Ca 2 § with a Ca2+-buffered solution with malate. With 70 mM intracellular EGTA and 10 mM external Ca 2+ solutions,/ca also decayed with a single-exponential curve, but it was about four times faster (~3.5 s -~ at 0 mV pulse). In these solutions the rate constant showed a direct relationship with Ic~ amplitude at different potentials. With 70 mM EGTA, replacing the external 10 mM Ca ~+ solution with the Ca ~+buffered solution caused the decay of Ic~ to become slower and to have the same relationship with membrane potential and/ca amplitude as in fibers with 20 mM EGTA internal solution. The mechanism of Ic~ decay depends on the intracellular EGTA concentration: (a) internal EGTA (both 20 and 70 mM) significantly reduces the voltage dependence of the inactivation process and (b) 70 mM EGTA dramatically increases the rate of tubular calcium depletion during the flow of ~c~.

Calcium currents during contraction and shortening in enzymatically isolated murine skeletal muscle fibres

The Journal of Physiology, 1999

Calcium currents (ICa) were monitored in enzymatically isolated murine toe muscle fibres using the two-microelectrode voltage-clamp technique. ICa was recorded (i) in hypertonic solution to suppress contraction, and (ii) in actively shortening fibres in isotonic solution.In hypertonic solution the threshold potential for ICa was about −30 mV for both 2 and 10 mM external Ca2+ solution. Maximum peak currents measured −12.6 ± 2.3 nA (mean ± s.d.; n = 4) in 2 mM Ca2+ and −65 ± 15 nA (n = 7) in 10 mM Ca2+. The time to peak (TTP) ICa was 96 ± 22 ms (n = 4) in 2 mM Ca2+ and 132 ± 13 ms (n = 7) in 10 mM Ca2+. The exponential decay of ICa was similar in 2 and 10 mM Ca2+ with rate constants (τ−1(V)) of 3.7 s−1 (2 mM) and 3.8 s−1 (10 mM) at +10 mV.ICa in isotonic 10 mM Ca2+ solution was recorded by inserting the micropipettes exactly opposite to each other close to the centre of mass of the fibre where negligible contraction-induced movement occurs.In isotonic 10 mM Ca2+ solution ICa had a smaller peak amplitude (−45 ± 5 nA; n = 7) and faster TTP (82.8 ± 22.1 ms; n = 7) than in hypertonic solution. The exponential decay of ICa showed a significantly larger τ−1(V) of 6.4 s−1 at +10 mV (P < 0.03).To test for calcium depletion, extracellular Ca2+ was buffered by malic acid in isotonic solution to 9 mM. The decay of ICa had a time constant of 348 ± 175 ms (n = 14) vs. 107 ± 24 ms (n = 12; P < 0.001) at 0 mV in unbuffered 10 mM Ca2+ solution.We conclude that calcium depletion from the transverse tubular system contributes significantly to the decay of calcium currents in murine toe muscle fibres under hypertonic as well as isotonic conditions. In the latter, depletion is even more prominent.

Activation of L-type calcium channel in twitch skeletal muscle fibres of the frog

The Journal of physiology, 1996

1. The activation of the L-type calcium current (ICa) was studied in normally polarized (-100 mV) cut skeletal muscle fibres of the frog with the double Vaseline-gap voltage-clamp technique. Both external and internal solutions were Ca2+ buffered. Solutions were made in order to minimize all but the Ca2+ current. 2. The voltage-dependent components of the time course of activation were determined by two procedures: fast and slow components were evaluated by multiexponential fitting to current traces elicited by long voltage pulses (5 s) after removing inactivation; fast components were also determined by short voltage pulses having different duration (0.5-70 ms). 3. The components of deactivation were evaluated after removing the charge-movement current from the total tail current by the difference between two short (50 and 70 ms) voltage pulses to 10 mV, moving the same intramembrane charge. Two exponential components, fast and slow (time constants, 6 +/- 0.3 and 90 +/- 7 ms at -10...

Voltage dependence of the pattern and frequency of discrete Ca2+ release events after brief repriming in frog skeletal muscle

Proceedings of the National Academy of Sciences, 1997

Applying a brief repolarizing pre-pulse to a depolarized frog skeletal muscle fiber restores a small fraction of the transverse tubule membrane voltage sensors from the inactivated state. During a subsequent depolarizing test pulse we detected brief, highly localized elevations of myoplasmic Ca 2؉ concentration (Ca 2؉ "sparks") initiated by restored voltage sensors in individual triads at all test pulse voltages. The latency histogram of these events gives the gating pattern of the sarcoplasmic reticulum (SR) calcium release channels controlled by the restored voltage sensors. Both event frequency and clustering of events near the start of the test pulse increase with test pulse depolarization. The macroscopic SR calcium release waveform, obtained from the spark latency histogram and the estimated open time of the channel or channels underlying a spark, exhibits an early peak and rapid marked decline during large depolarizations. For smaller depolarizations, the release waveform exhibits a smaller peak and a slower decline. However, the mean use time and mean amplitude of the individual sparks are quite similar at all test depolarizations and at all times during a given depolarization, indicating that the channel open times and conductances underlying sparks are essentially independent of voltage. Thus, the voltage dependence of SR Ca 2؉ release is due to changes in the frequency and pattern of occurrence of individual, voltage-independent, discrete release events.

Modulation of calcium current gating in frog skeletal muscle by conditioning depolarization

The Journal of physiology, 1992

1. Ca2+ inward currents were measured by voltage clamping cut skeletal muscle fibres of the frog (Rana esculenta) in a double-Vaseline-gap system. 2. In order to study the basis of the previously described fast gating mode induced in the Ca2+ inward current by a conditioning depolarization we quantitatively analysed the response to differing features of the conditioning prepulse. 3. The faster activation seen during the second of two depolarizations was confined to the component of the inward current which could be blocked by 5 to 10 microM nifedipine. 4. By applying depolarizing conditioning pulses of gradually increasing length the time course of the transition to the fast gating mode could be determined. 5. Both the transition to the fast gating mode (point 4) caused by a depolarization and the slow inward current activated during the same depolarization showed similar voltage-dependent kinetics. 6. The kinetic change of the test current appeared to be equal when the same fractio...

Calcium-channel gating in frog skeletal muscle membrane: effect of temperature

The Journal of Physiology, 1983

1. Voltage-clamp experiments using the three micro-electrode method were performed to study the temperature dependence of the calcium current ICa in intact twitch skeletal muscle fibres of the frog. Contraction was blocked by recording in hypertonic sucrose solutions. 2. For depolarizations smaller than 0 mV the decay of the transient, slow, inward current, recorded in the presence of external tetraethylammonium (TEA+) and by replacing Cl-for CH3SO3-, followed a complex time course. For larger depolarizations, after the initial inward current, there was a prominent, slow, outward current which showed two phases: after reaching a peak (time to peak 1-0 sec, peak amplitude 20-50 #sA/cm2 at 20 mV) it slowly declined to a steady level in about 2-3 see at 23 'C. 3. The inward current was greatly reduced or abolished by the adding of2 mM-Cd2+ or by replacing external Ca2+ with Mg2+. The amplitude and time course of slow, outward currents were not obviously modified by replacing Ca2+ with Mg2+, having the two described phases. However, in the presence of Cd2+ the first transient phase of the outward current was not detected and only outward currents slowly increasing to a steady level were observed. 4. Reliable ICa records were obtained by further blocking K+ outward currents by incubating the muscles in a K+-free TEA+and Cs+-containing solution prior to experiments. Tubular space clamp was improved by recording ICa from small fibres with 20-30 jsm radius. 5. The decay phase of ICa under a maintained depolarization in incubated muscles was fitted by a single exponential. The corresponding rate constant determined between 12 and 24 OC strongly depended on temperature, as expected for a gating process. The values for the activation energy and the corresponding Q10 (calculated for a 10-20 0C transition) were respectively: 17-5 + 1'0 kcal/mole and 2-9 + 0-2 at 0 mV, and 18-0 + 1-5 kcal/mole and 3 0 + 0-3 at-20 mV. 6. The activation phase of Ica, analysed following the ma2h Hodgkin-Huxley kinetic model, showed a similar temperature dependence with a Q10 of 3-0 + 0 3. The * Recipient of a fellowship from CONACyT, Mexico.