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Role of the Ryanodine Receptor of Skeletal Muscle in Excitation‐Contraction Coupling
Author(s) -
FILL MICHAEL,
MA JIANJIE,
KNUDSON C. MICHAEL,
IMAGAWA TOSHIAKI,
CAMPBELL KEVIN P.,
CORONADO ROBERTO
Publication year - 1989
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.1989.tb24092.x
Subject(s) - annals , physiology , medicine , gerontology , classics , history
In skeletal muscle, contraction is initiated by a depolarization of the transverse tubular membrane (t-tubule), which in turn signals the release of Ca from the sarcoplasmic reticulum (SR). A key protein involved in this process is the ryanodine receptor, an SR membrane protein of MW 450,000 that binds the alkaloid ryanodine with nanomolar affinity and is present exclusively at the junction between t-tubule and SR membranes. The ryanodine receptor plays a dual role: Functionally, it is the putative Ca-release channel of the SR, and structurally, it is the major protein responsible for forming “bridges” or “feet” that anatomically connect t-tubule and SR. Here we demonstrate that the ryanodine receptor is steeply gated by both voltage and protons, and for the first time in vitro, we measured nonlinear capacitance (charge movement) that may be involved in the gating of this channel protein. We recently identified the 450,000-Da ryanodine receptor-feet protein (FIG. 1A) as the Ca-release channel of native SR. This was achieved using the planar bilayer recording technique and by comparing ligand-dependent gating, ionic selectivity, and pharmacology of purified ryanodine receptors to that of native Ca-release channels. Voltage dependence was a gating property notoriously absent in our study and in studies by others that followed. Its inconspicuousness in our earlier work is related to the effect of protons. At pH 7.4 (FIG. 1B) the channel dwells in a fast gating mode (p = 0.38). A drop to pH 7.2 drives the channel into an almost closed condition (p = 0.08) and at pH 7.0 the channel never opens (p < 0.01). Reversibility is shown in the last record of FIGURE 1 where alkalinization from pH 7.0 to pH 7.6 reopens the channel, resulting in a higher level of activity (p = 0.72) than seen at pH 7.4. Over this narrow range of pH, slope conductance is not affected (FIG. 2C, inset), and the kinetics remain fast, with a mean open event duration of approximately 100 μsec. Thus for all practical purposes, a change of 0.6 unit in solution pH from 7.6 to 7.0 units is sufficient to make the channel switch from an almost all-open to an almost all-closed conformation. The fitted Hill coefficient for data in FIGURE 1 was n = 6.8 and the apparent pK a was 7.5. FIGURE 2 describes the ensuing changes in voltage