Effect of sarcoplasmic reticulum calcium depletion on intramembranous charge movement in frog cut muscle fibers.

Abstract

Cut muscle fibers from Rana temporaria (sarcomere length, 3.3-3.5 microns; temperature, 13-16 degrees C) were mounted in a double Vaseline-gap chamber and equilibrated for at least an hour with an internal solution that contained 20 mM EGTA and phenol red and an external solution that contained predominantly TEA-gluconate; both solutions were nominally Ca-free. The increase in total myoplasmic concentration of Ca (delta[CaT]) produced by sarcoplasmic reticulum (SR) Ca release was estimated from the change in pH produced when the released Ca was complexed by EGTA (Pape, P.C., D.-S. Jong, and W.K. Chandler. 1995. Journal of General Physiology. 106:259-336). The resting value of SR Ca content, [CaSR]R (expressed as myoplasmic concentration), was taken to be equal to the value of delta[CaT] obtained during a step depolarization (usually to -50 to -40 mV) that was sufficiently long (200-750 ms) to release all of the readily releasable Ca from the SR. In ten fibers, the first depolarization gave [CaSR]R = 839-1,698 microM. Progressively smaller values were obtained with subsequent depolarizations until, after 30-40 depolarizations, the value of [CaSR]R had usually been reduced to < 10 microM. Measurements of intramembranous charge movement, Icm, showed that, as the value of [CaSR]R decreased, ON-OFF charge equality held and the amount of charge moved remained constant. ON Icm showed brief initial I beta components and prominent I gamma "humps", even after the value of [CaSR]R was < 10 microM. Although the amplitude of the hump component decreased during depletion, its duration increased in a manner that preserved the constancy of ON charge. In the depleted state, charge movement was steeply voltage dependent, with a mean value of 7.2 mV for the Boltzmann factor k. These and other results are not consistent with the idea that there is one type of charge, Q beta, and that I gamma is a movement of Q beta caused by SR Ca release, as proposed by Pizarro, Csernoch, Uribe, Rodriguez, and Rios (1991. Journal of General Physiology. 97:913-947). Rather, our results imply that Q beta and Q gamma represent either two distinct species of charge or two transitions with different properties of a single species of charge, and that SR Ca content or release or some related event alters the kinetics, but not the amount of Q gamma. Many of the properties of Q gamma, as well as the voltage dependence of the rate of SR Ca release for small depolarizations, are consistent with predictions from a simple model in which the voltage sensor for SR Ca release consists of four interacting charge movement particles.

Kinetic isoforms of intramembrane charge in intact amphibian striated muscle.

“…Hui and Chen, 1992). When applied to data from fibers exposed to RyR antagonists, the successive iterations yielded progressively diminishing values of one of the Qma~ terms, increasingly positive values of its corresponding transition potential, V*, and increasing values of the standard error of the slope factor k. This resembles recent reports for gluconate-containing solutions (Jong et al, 1995), the similar convergences of which toward single Boltzmann functions were attributed to the smaller proportion of q~ charge or closer values of V* shown by q~ and qv in gluconate-treated fibers. The latter was confirmed in the pharmacological analysis below; this would lead to convergences to a single Boltzmann function whose value of k fell between those of q~ and qv, respectively.…”

“…Yet, the present studies demonstrate that RyR modifications alter the kinetics particularly of q~ transients, even though the RyR resides in membranes of the terminal cisternae, from which the tubules are electrically isolated (see review in Huang, 1993). Recent depletions or loadings of sarcoplasmic reticular Ca 2+ have similarly accomplished kinetic changes while preserving the steady-state characteristics and individual identities of the component charge species (Jong et al, 1995;Pape et al, 1996). These suggest that altered conditions of intracellular Ca 2+ resulting from Ca 2+ release channel modification might cause such changes.…”

“…Thus, Table I Table II ofJong et al, 1995) when test agents were absent. This marked voltage dependence was attributed to steadystate contributions from a distinct species of q~ charge in both intact (Adrian and Peres, 1979;Huang, 1982;Hui, 1983) and cut Chandler, 1990, 1991) muscle fibers in this recent work (Jong et al, 1995). It contrasts with the more gradual variations (k ~ 14 to 15 mV; see Table IV below) attributed to the q~ charge.…”

“…Additionally, RyR modification also preserved the maximum charge in, and the steepness (k -~ 7-9 mV) of, inactivation curves. Jong et al (1995) and Pape et al (1996) similarly reported persistently steep charge-voltage relationships (k ~ 7 mV) characteristic of a q~ contribution, whether fibers were depleted of, or loaded with, Ca z+.…”

“…The charge-voltage plots also altered to a form that more closely resembled a single Boltzmann plot than did the control results obtained in the absence of RyR antagonists. Accordingly, the primary aim of the analysis in Table I was simply to draw overall comparisons with earlier results in cut fibers in the same solutions (Jong et al, 1995). Thus, Table I Table II ofJong et al, 1995) when test agents were absent.…”