Sulphydryl reagents trigger Ca2+ release from the sarcoplasmic reticulum of skinned rabbit psoas fibres.

Abstract

SUMMARY1. By analogy with studies on sarcoplasmic reticulum (SR) vesicles, Ca2+ release induced by heavy metals and mercaptans (e.g. cysteine) was investigated in rabbit skinned psoas fibres through measurements of isometric tension.2. Heavy metals (at 2-5 /tM) elicited phasic contractions by triggering Ca2+ release from the SR and had the following order of potency: Hg2+ > Cu2+ > Cd2+ > Ag+ > Ni2+. Higher concentrations produced tonic contractions due to maintained high Ca2+ permeability of SR membranes.3. Contractions induced by heavy metals required a functional and Ca2+-loaded SR, were dependent on [Ca2+]free, blocked by Ruthenium Red (RR), inhibited by free Mg2+ and reduced glutathione (GSH) but not by oxidized glutathione (GSSG). Such contractions were not elicited through direct interactions) of heavy metals with the myofilaments.4. In the presence of catalytic concentrations of Hg2+ or Cu2+ (2-5 /UM), additions of cysteine (25-100 /LM) elicited rapid twitches, producing 70 % of maximal force with a time to half-peak of 2 s. Contractions induced by cysteine plus a catalyst required a functional SR network, were dependent on free [Mg2+] and were blocked by RR or GSH but not by GSSG.5. In the presence of Hg2+ (2-5 ,uM), low concentrations of cysteine (10 /uM) elicited tonic contractures, but subsequent or higher additions of cysteine (50-100 uM) caused further SR Ca2+ release and tension, followed by rapid and full relaxation.6. High cysteine (200-250 /tM, without Cu2+ or Hg2+) blocked contractions elicited by Cl-induced depolarization of sealed T-tubules. High cysteine probably acted as a sulphydryl reducing agent which promoted rapid relaxation of the fibres through the closure of Ca2+-release channels and ATP-dependent re-uptake of Ca2+ by the SR. 7. In some batches of skinned fibres (-10%), cysteine (5-50 /tM) alone (without Hg2+ or Cu2+ catalyst) produced rapid twitches. This implied that the catalysts) necessary to promote the sulphydryl oxidation reaction with exogenously added cysteine may be present in intact fibres but is usually lost by the skinning procedure.

Further Characterization of the Type 3 Ryanodine Receptor (RyR3) Purified from Rabbit Diaphragm

“…It has been demonstrated that a number of sulfhydryl-oxidizing reagents activate RyR channels (40,41,43). Sulfhydrylreducing reagents such as glutathione and DTT have been shown to reduce the RyR channel activity (39,42).…”

Oxidant stress activates a non‐selective cation channel responsible for membrane depolarization in calf vascular endothelial cells.

“…This observation initially suggests that the oxidant-activated channel may not require the presence of an intracellular messenger to remain active. Ca2+ release channels from the sarcoplasmic reticulum of skeletal muscle are activated by the oxidation of critical thiol-containing residues (Salama, Abramson & Pike, 1992). Further, thiol oxidation has been shown to modulate the activity of non-selective channels in adipocytes (Koivisto, Siemen & Nedergaard, 1993).…”

Sarcoplasmic reticulum Ca2+release and muscle fatigue

“…It has also been proposed that endogenous sulfhydryl (SH) groups present on the RYR protein may be responsible for regulating the gating of the SR Ca 2ϩ -release channel (1,111,112). It has been demonstrated that oxidation of the SH groups stimulates SR Ca 2ϩ release in vesicle experiments (2) and muscle fibers (112).…”

“…It has been demonstrated that oxidation of the SH groups stimulates SR Ca 2ϩ release in vesicle experiments (2) and muscle fibers (112). Whereas no clear role for the SH regulation has been established, it remains a viable mechanism to modulate channel gating.…”