Effects of perchlorate on the molecules of excitation-contraction coupling of skeletal and cardiac muscle.
To understand the nature of the transmission process of excitation-contraction (EC) coupling, the effects of the anion perchlorate were investigated on the voltage sensor (dihydropyridine receptor, DHPR) and the Ca release channel (ryanodine receptor, RyR) of the sarcoplasmic reticulum (SR). The molecules, from rabbit skeletal muscle, were either separated in membrane vesicular fractions or biochemically purified so that the normal EC coupling interaction was prevented. Additionally, the effect of ClO4- was investigated on L-type Ca2+ channel gating currents of guinea pig ventricular myocytes, as a native DHPR not in the physiological interaction of skeletal muscle. At 20 mM, ClO4- had minor effects on the activation of ionic currents through Ca channels from skeletal muscle transverse tubular (T) membranes fused with planar bilayers: a +7-mV shift in the midpoint voltage, V, with no change in kinetics of activation or deactivation. This is in contrast with the larger, negative shift that ClO4- causes on the distribution of intramembrane charge movement of skeletal muscle. At up to 100 mM it did not affect the binding of the DHP [3H]PN200-110 to triad-enriched membrane fractions (TR). At 8 mM it did not affect the kinetics or the voltage distribution of gating currents of Ca channels in heart myocytes. These negative results were in contrast to the effects of ClO4- on the release channel. At 20 mM it increased several-fold the open probability of channels from purified RyR incorporated in planar bilayers and conducting Ba2+, an effect seen on channels first closed by chelation of Ca2+ or by the presence of Mg2+. It significantly increased the initial rate of efflux of 45Ca2+ from TR vesicles (by a factor of 1.75 at 20 mM and 4.5 at 100 mM). ClO4- also increased the binding of [3H]ryanodine to TR fractions. The relative increase in binding was 50-fold at the lowest [Ca2+] used (1 microM) and then decayed to much lower values as [Ca2+] was increased. The increase was due entirely to an increase in the association rate constant of ryanodine binding. The chaotropic ions SCN- and I- increased the association rate constant to a similar extent. The binding of ryanodine to purified RyR protein reconstituted into liposomes had a greater affinity than to TR fractions but was similarly enhanced by ClO4-. The reducing agent dithiothreitol (5 mM) did not reduce the effect of ClO4-, and 5% polyethylene glycol, with an osmolarity equivalent to 20 mM ClO4-, did not change ryanodine binding.(ABSTRACT TRUNCATED AT 400 WORDS)
Ma
J
J
Department of Physiology, Rush University School of Medicine, Chicago, Illinois 60612.
Anderson
K
K
Shirokov
R
R
Levis
R
R
González
A
A
Karhanek
M
M
Hosey
M M
MM
Meissner
G
G
Ríos
E
E
D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002132:Calcium Radioisotopes; D002412:Cations / Q000378:metabolism; D004594:Electrophysiology; D006168:Guinea Pigs; D006321:Heart / Q000187:drug effects* / Q000502:physiology; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects; D017275:Isradipine / Q000378:metabolism; D009119:Muscle Contraction / Q000187:drug effects; D009132:Muscles / Q000187:drug effects* / Q000502:physiology; D009206:Myocardium / Q000166:cytology; D009469:Neuromuscular Junction / Q000187:drug effects* / Q000502:physiology; D010472:Perchlorates / Q000494:pharmacology*; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000187:drug effects; D017670:Sodium Compounds / Q000494:pharmacology*
D015220:Calcium Channels; D002132:Calcium Radioisotopes; D002412:Cations; D010472:Perchlorates; D017670:Sodium Compounds; D012433:Ryanodine; C031068:sodium perchlorate; D017275:Isradipine