Angol nyelvű Tudományos Szakcikk (Folyóiratcikk)
Megjelent: JOURNAL OF GENERAL PHYSIOLOGY 0022-1295 1540-7748 102 (3) pp. 373-421 1993
      The effects of the anion perchlorate (present extracellularly at 8 mM) were studied on functional skeletal muscle fibers from Rana pipiens, voltage-clamped in a Vaseline gap chamber. Established methods were used to monitor intramem-branous charge movement and flux of Ca release from the sarcoplasmic reticulum (SR) during pulse depolarization. Saponin permeabilization of the end portions of the fiber segment (Irving, M., J. Maylie, N. L. Sizto, and W. K. Chandler. 1987. Journal of General Physiology. 89:1-41) substantially reduced the amount of charge moving during conventional control pulses, thus minimizing a technical error that plagued our previous studies. Perchlorate prolonged the ON time course of charge movement, especially at low and intermediate voltages. The OFFs were also made slower, the time constant increasing twofold. The hump kinetic component was exaggerated by ClO4− or was made to appear in fibers that did not have it in reference conditions. ClO4− had essentially no kinetic ON effects at high voltages (>10 mV). ClO4− changed the voltage distribution of mobile charge. In single Boltzmann fits, the midpoint potential V was shifted -20 mV and the steepness parameter K was reduced by 4.7 mV (or 1.78-fold), but the maximum charge was unchanged (n = 9). Total Ca content in the SR, estimated using the method of Schneider et al. (Schneider, M. F., B. J. Simon, and G. Szucs. 1987. Journal of Physiology. 392:167-192) for correcting for depletion, stayed constant over tens of minutes in reference conditions but decayed in ClO4− at an average rate of 0.3 (xmol/liter myoplasmic water per s. ClO4− changed the kinetics of release flux, reducing the fractional inactivation of release after the peak. ClO4− shifted the voltage dependence of Ca release flux. In particular, the threshold voltage for Ca release was shifted by about -20 mV, and the activation of the steady component of release flux was shifted by > 20 mV in the negative direction. The shift of release activation was greater than that of mobile charge. Thus the threshold charge, defined as the minimum charge moved for eliciting a detectable Ca transient, was reduced from 6 nC/µF (0.55, n = 7) to 3.4 (0.53). The average of the paired differences was 2.8 (0.33, P > 0.01). The effects of ClO4− were then studied in fibers in modified functional situations. Depletion of Ca in the SR, achieved by high frequency pulsing in the presence of intracellular BAPTA and EGTA, simplified but did not eliminate the effects of ClO4−. ON humps were not observed in the depleted fibers but the slowing effect of ClO4−, both in ON and OFF, was still present. The shift in V was reduced to -15 mV, the change in steepness was reduced to ~ 15%, and Qmax was unchanged. The threshold charge was reduced by ClO4− regardless of depletion. In fibers inactivated by prolonged depolarization ClO4− did not change the kinetics of charge movement (charge 2) but changed its voltage distribution, shifting V by -14 mV (n = 6). The Cl channel blockers A9C, SITS, and DIDS shifted threshold depolarization for Ca release to more negative potentials, but lacked other effects of ClO4−. The results are evidence that ClO4− has a complex set of effects including: (1) a negative voltage shift that at least in part is shared with other Cl channel blockers, (2) a large increase in the steepness of the charge vs. voltage distribution, which is probably mediated by Ca2+ released from the SR, (3) a slowing of charge movement, and (4) an improvement in the transmission from voltage sensor to release channel, manifested in a reduction of threshold charge. Effects 3 and, of course, 4 have a specific requirement, that the interaction between sensor and release channel be functional. Two possible mechanisms are considered for effects 3 and 4: a binding to the voltage sensor that is sensitive to its functional state and a binding to the release channel that affects the voltage sensor secondarily through a mechanical link. These possibilities are further explored in the following articles in this series.
      Hivatkozás stílusok: IEEEACMAPAChicagoHarvardCSLMásolásNyomtatás
      2021-05-15 19:19