Scorpion toxins targeted against the sarcoplasmic reticulum Ca(2+)-release channel of skeletal and cardiac muscle.

Abstract

We report the purification of two peptides, called "imperatoxin inhibitor" and "imperatoxin activator," from the venom of the scorpion Pandinus imperator targeted against ryanodine receptor Ca2+-release channels. Imperatoxin inhibitor has a Mr of 10,500, inhibits [3Hjryanodine binding to skeletal and cardiac sarcoplasmic reticulum with an EDso of 10 nM, and blocks openings of skeletal and cardiac Ca2+-release channels incorporated into planar bilayers. In whole-cell recordings of cardiac myocytes, imperatoxin inhibitor decreased twitch amplitude and intracellular Ca2+ transients, suggesting a selective blockade of Ca2+ release from the sarcoplasmic reticulum. Imperatoxin activator has a Mr of -8700, stimulates [3H]ryanodine bind in skeletal but not cardiac sarcoplasmic reticulum with an ED50 of -6 nM, and activates skeletal but not cardiac Ca2+-release channels. These ligands may serve to selectively "turn on" or "turn off" ryanodine receptors in fragmented systems and whole cells.Activation of muscle, neurons, and secretory cells by voltage, neurotransmitters, or hormones can evoke a release of Ca2+ from intracellular Ca2+ stores (1). Two types of Ca2+ channels, namely ryanodine receptors and inositol 1,4,5-trisphosphate (InsP3) receptors, have been shown to control the intracellular Ca2+ permeability of many cells. In striated muscle, these channels transduce membrane voltage, sarcolemmal Ca2+ entry, and other external stimuli into an increase in the Ca2+ permeability of the sarcoplasmic reticulum (SR) (2). Elucidation of the mechanism of Ca2+ release from intracellular stores depends critically on the specificity of pharmacological agents to selectively alter a single intracellular Ca2+ channel type. The alkaloid ryanodine is presently the only ligand available to dissect the contribution of ryanodine receptors to intracellular Ca2+ release in situ. However, the usefulness of this compound is limited by the fact that it has an extremely slow association and dissociation kinetics that makes the onset of the pharmacological effect slow and essentially irreversible (3). To accelerate the onset, micromolar instead of nanomolar levels of ryanodine are used, but at micromolar concentrations the alkaloid inhibits other Ca2+ channels (4). Furthermore, certain concentrations of ryanodine may open the Ca2+-release channel while others may block it (5), leading to ambiguous results (6). Alternative ligands that act fast, reversibly, and in a simple manner should be more helpful in establishing the contribution of ryanodine receptors to intracellular Ca2+ signals.Scorpion venoms have traditionally represented an invaluable source ofpeptide toxins specific for a single channel type (7). In the present report, we screened venom from several genera of scorpions with the hope of finding peptides specific for ryanodine receptors. We purified two peptides from the venom of Pandinus imperator that selectively blocked [imperatoxin inhibitor (IpTxi)] or activated [imperatoxin activator (IpTxa)] ryanodine receptors of skel...

Cyclic ADP‐ribose induced Ca2+ release in rabbit skeletal muscle sarcoplasmic reticulum

“…2 by fusion of rabbit junctional SR to a planar bilayer. The identification of this large unit conductance channel as the ryanodine receptor has been documented extensively [9,17,21]. In this case, the control channel activity at 0 mV was elicited by Ca" which is present in the myoplasmic-equivalent cis solution as a contaminant and is typically 1 to 3 PM free Ca".…”

“…A separate class of intracellular Ca" channels, namely ryanodine receptors, has been described in muscle and brain cells where they are believed to mediate the ubiquitous Ca"-mobilizing mechanism known as Ca"-induced Ca7+ release [7]. In skeletal and cardiac muscle, ryanodine receptors are abundant in the junctional sarcoplasmic reticulum [8] and release Ca" during excitation-contraction coupling in response to cell membrane depolarization [9]. Ryanodine receptors are activated by Ca'+ at physiological concentrations, and are modulated by a variety of ligands that affect Ca"-induced Ca7+ release such as Mg'+, adenine nucleotides and caffeine Lee [15] showed that cADPR like caffeine.…”

Maurocalcine and Peptide A Stabilize Distinct Subconductance States of Ryanodine Receptor Type 1, Revealing a Proportional Gating Mechanism

“…Imperatoxin A (IpTx a ), a 33-amino acid peptide isolated from the scorpion Pandinus imperator, was shown to be a high affinity agonist of RyR1 channels (20,21). Nanomolar IpTx a increased high affinity [ 3 H]ryanodine-binding and induced rapid Ca 2ϩ release from SR vesicles.…”

Myocarditis with Hypotension after a Scorpion Sting

“…Cardiac damage might be enhanced by the depressive effect of cytokines upon myocardial cells. Hyperglycemia may also contribute to myocardial injury [10]. Valdivia et al [11] reported a series of 32 children with scorpion bites who developed cardiac complications.…”