Lipoprotein and protein binding of the calcium channel blocker diltiazem

The plasma protein binding properties of the calcium channel blocker diltiazem were studied using a three-chamber equilibrium dialysis system. Diltiazem is 81% bound to human sera with significant inter-individual variation. The relative binding of diltiazem by lipoproteins and alpha 1-acid glycoprotein was higher than by albumin. The binding to low density lipoprotein was strong and appeared not to be associated with the surface apoprotein.     

The role of region IVS5 of the human cardiac calcium channel in establishing inactivated channel conformation: use-dependent block by benzothiazepines

The role of inactivated channel conformation and use dependence for diltiazem, a specific benzothiazepine calcium channel inhibitor, was studied in chimeric constructs and point mutants created in the IVS5 transmembrane segment of the L-type cardiac calcium channel. All mutations, chimeric or point mutations, were restricted to IVS5, while the YAI-containing segment in IVS6, i.e. the primary interaction site with benzothiazepines, remained intact. Slowed inactivation rate and incomplete steady state inactivation, a behavior of some mutants, were accompanied by a reduced or by a complete loss of use-dependent block by diltiazem. Single channel properties of mutants that lost use dependence toward diltiazem were characterized by drastically elongated mean open times and distinctly slower time constants of open time distribution. Mutation of individual residues of the IVMLF segment in IVS5 did not mimic the complete loss of use dependence as observed for the replacement of the whole stretch. These results establish evidence that amino acids that govern inactivation and the drug-binding site and other amino acids that are located distal from the putative drug-binding site contribute significantly to the function of the benzothiazepine receptor region. The data are consistent with a complex "pocket" conformation that is responsive to a specific class of L-type calcium channel inhibitors. The data allow for a concept that multiple sites within regions of the alpha(1) subunit contribute to auto-regulation of the L-type Ca(2+) channel.     

Effect of verapamil on platelet aggregation

The influence of the calcium channel blocker verapamil on the aggregation of human blood platelets was studied in vitro in comparison with the calcium channel blocker diltiazem and with the 5-HT antagonist cyproheptadine. Verapamil inhibited the 5-HT-potentiated. ADP-induced aggregation more effectively than the aggregation induced by adrenaline, ADP and collagen. Verapamil antagonized the 5-HT effect in a noncompetitive manner. The same was true of cyprohepatadine which was by more than one order of magnitude more potent than verapamil in inhibiting the 5-HT-induced aggregation. Diltiazem was much less effective than verapamil.     

Calcium channel blockers and prostaglandin generation by gastric surface epithelial cells.

We investigated the effects of calcium channel blockers on generation of prostaglandin (PG) E2 and 6-keto PGF1 alpha by gastric mucosal surface epithelium. Surface epithelial cells (SEC) isolated from rat gastric mucosa were incubated with either verapamil (1 or 10 micrograms/ml), diltiazem (2.5 or 25 micrograms/ml) or nifedipine (2.5 or 25 micrograms/ml) for 30 min at 37 degrees C in calcium containing or calcium-free medium. Verapamil (both doses) significantly increased PGE2 and 6-keto PGF1 alpha generation by the surface epithelial cells but only in calcium containing medium. Diltiazem did not affect PG generation in calcium containing nor calcium-free medium. Nifedipine 25 micrograms/ml decreased PGE2 but increased 6-keto PGF1 alpha generation. The inhibitory effect of nifedipine on PGE2 generation was abolished in calcium-free medium, while the calmodulin antagonist did not affect verapamil-induced increase in PG generation.     

Effects of the calcium antagonist, diltiazem on in vitro and in vivo/in vitro bone resorption

The effect of the calcium antagonist diltiazem on bone resorption in organ culture has been investigated. It was found that diltiazem was ineffective alone but that in concentrations above 5 mumol/l it reduced mineral and organic resorption induced in vitro by 1.25 dihydroxycholecalciferol (1.25 (OH)2D3). No additivity with calcitonin effects was observed. Diltiazem did not significantly affect bone resorbing activity stimulated by 24,25(OH)2D3. Bone resorption was measured by an in vivo/in vitro technique using 45Ca prelabelled mice. Compared with 1.25(OH)2D3 alone treated group (0.480 pmol/g), it was found that diltiazem (100 nmol/g) reduced bone resorption without effect on calcium and phosphorus plasmatic concentrations at death. These data suggest that such a calcium antagonist is able to inhibit 1.25-(OH)2D3-increased-bone resorption either in vitro or in vivo/in vitro.     

Effect of diltiazem, a calcium antagonist, on renin secretion from rat kidney slices

The purpose of these experiments was to characterize the effects of diltiazem on renin secretion from rat renal cortical slices. Incubation of slices in 60 versus 4 mM K medium almost completely abolished renin secretion. Diltiazem antagonized the inhibitory effect in a concentration-dependent manner but had no effect on secretion of slices incubated in 4 mM K medium. Lowering extracellular Ca enhanced the efficacy of diltiazem. These observations demonstrate that Ca influx through voltage-sensitive Ca channels mediates the inhibitory effect of depolarization and further demonstrate that such channels are not open in the basal state of this preparation. In the presence of a concentration of diltiazem which blocked the inhibitory effects of depolarization, both angiotensin II and antidiuretic hormone (ADH) still inhibited secretion. Therefore, both these peptides inhibit secretion by mechanisms which are independent of the voltage-sensitive Ca channels. These observations confirm and extend previous observations suggesting that Ca plays an inhibitory coupling role in the control of renin secretion.     

Diltiazem at high concentration increases the ionic permeability of biological membranes

Summary The effects of diltiazem, a drug which inhibits the calcium channels in cardiac muscle as well as the light-sensitive channels in photoreceptor cells, were studied on ionic fluxes in both membrane and intact cell preparations. Diltiazem nonselectively increased the ionic permeability to both anions and cations in photoreceptor rod outer segment and synaptic membrane vesicles as well as in intact erythrocytes. Under our conditions, the estimated threshold for the diltiazem effect varied between 12.5 and 200 m. In each case the concentration dependence exhibited the sigmoidal shape characteristic of positive cooperativity. The effect of diltiazem on ionic fluxes from phospholipid vesicles were strongly influenced by phospholipid composition and membrane charge. By contrast, diltiazem inhibited the efflux of⁸⁶Rb from photoreceptor cells of intact aspartate-isolated retina, an effect opposite to that of diltiazem on ionic permeabilities in photoreceptor membrane vesicle preparations.These data raise serious doubts on the specificity of diltiazem as a calcium channel blocker or as a cGMP channel blocker when used at concentrations higher than 10 m.     

In Vivo and Ex Vivo Evaluation of L-Type Calcium Channel Blockers on Acid β-Glucosidase in Gaucher Disease Mouse Models

Gaucher disease is a lysosomal storage disease caused by mutations in acid β-glucosidase (GCase) leading to defective hydrolysis and accumulation of its substrates. Two L-type calcium channel (LTCC) blockers—verapamil and diltiazem—have been reported to modulate endoplasmic reticulum (ER) folding, trafficking, and activity of GCase in human Gaucher disease fibroblasts. Similarly, these LTCC blockers were tested with cultured skin fibroblasts from homozygous point-mutated GCase mice (V394L, D409H, D409V, and N370S) with the effect of enhancing of GCase activity. Correspondingly, diltiazem increased GCase protein and facilitated GCase trafficking to the lysosomes of these cells. The in vivo effects of diltiazem on GCase were evaluated in mice homozygous wild-type (WT), V394L and D409H. In D409H homozygotes diltiazem (10 mg/kg/d via drinking water or 50–200 mg/kg/d intraperitoneally) had minor effects on increasing GCase activity in brain and liver (1.2-fold). Diltiazem treatment (10 mg/kg/d) had essentially no effect on WT and V394L GCase protein or activity levels (<1.2-fold) in liver. These results show that LTCC blockers had the ex vivo effects of increasing GCase activity and protein in the mouse fibroblasts, but these effects did not translate into similar changes in vivo even at very high drug doses.     

The effects of calcium channel blockers on isoproterenol induced cyclic adenosine 3',5'-monophosphate generation in intact human lymphocytes

We have investigated the effects of clinically available calcium channel blockers (nifedipine, verapamil and diltiazem) on isoproterenol stimulated cyclic adenosine 3',5'-monophosphate (cyclic AMP) generation in intact human lymphocytes. After preincubation of various calcium antagonists with intact lymphocytes at 37 degrees C for 15 minutes, 10 microM nifedipine or verapamil partially inhibited isoproterenol induced cyclic AMP generation in the presence of cyclic AMP phosphodiesterase inhibitor (3-isobutyl-1-methylxanthine) while they alone had no effect on cyclic AMP level at a concentration of up to 100 microM. In contrast, 10 nM-1.0 microM nifedipine, verapamil or diltiazem potentiated cyclic AMP generation induced by isoproterenol in a dose dependent manner. Similar results were observed in the time course studies of cyclic AMP generation. These effects are somewhat similar to the effect of phenothiazine, a calmodulin inhibitor, which, at 10 microM (close to IC50), also potentiated the effects of isoproterenol. In contrast, lanthanum chloride (LaCl3), an extracellular inorganic calcium antagonist, at 1.0 mM, inhibited isoproterenol induced cyclic AMP generation. The biochemical mechanisms underlying these potentiating effects are unknown. It may be partly related to the effect of calcium channel blockers (at least for nifedipine) on preventing beta 2 adrenergic receptor desensitization. This may provide a potential mechanism for the synergistic effect between calcium channel blockers and beta 2 adrenoceptor agonists on bronchial dilatation.     

A new class of calcium entry blockers defined by 1,3-diphosphonates. Interactions of SR-7037 (belfosdil) with receptors for calcium channel ligands

Tetrabutyl-2(2-phenoxyethyl)-1,3-propylidene diphosphonate (SR-7037) completely displaced dihydropyridine [( 3H]PN200-110), phenylalkylamine [( 3H]D888), and benzothiazepine [( 3H]diltiazem) ligands from brain L-type calcium channels. Half-maximal inhibition of [3H]PN200-110 binding occurred at 19 nM with a Hill coefficient of 0.96. SR-7037 primarily decreased the affinity for [3H]PN200-110 with a small, but significantly, effect on the maximal binding capacity. Kinetic studies showed that this was due to an increased radioligand dissociation rate from 0.04 min-1 to 0.43 min-1 in the presence of the diphosphonate. Displacement of [3H]D888 by SR-7037 was biphasic with respective IC50 of 44 and 8400 nM. Likewise, unlabeled (-)-D888 identified two sites with IC50 values of 0.9 and 27 nM. Both SR-7037 (1000 nM) and D888 (200 nM) accelerated radioligand dissociation about 2-fold. [3H]Diltiazem binding was inhibited by SR-7037 with an IC50 value of 29 nM. The inhibition of dihydropyridine binding by SR-7037 is enhanced by most divalent cations at millimolar concentrations with the following potency: Mn2+ greater than Mg2+ greater than Ca2+ greater than Co2+. Barium has the opposite effect. The half-maximal effect of calcium occurred at 6 microM free ion. Specific binding of [3H]D888 was antagonized in the presence of 1 mM CaCl2. It is concluded that SR-7037 has allosteric interactions with the dihydropyridine receptor of the L-type calcium channel. The differential effect of Ca2+ on the potency of D888 and diltiazem relative to that of SR-7037 indicates that the three drugs may bind to nonequivalent sites. These results support specific calcium channel inhibition, possibly at a novel site, as the primary mechanism of the diphosphonate's pharmacological actions.     

Mechanism of inhibition of net ion transport across frog corneal epithelium by calcium channel antagonists

Summary In the isolated bullfrog cornea, three calcium channel antagonists had dose-dependent inhibitory effects on the Cl-originated short-circuit current (SCC). Their order of decreasing potency was bepridil, verapamil and diltiazem. One millimolar diltiazem inhibited the SCC by 98% and subsequent incubation with the calcium ionophore A23187 had no restorative effect. Increasing the bathing solution Ca concentration from 0.05 to 15mm, however, decreased diltiazem's inhibitory efficacy. This antagonist depolarized the intracellular potential differenceV _m from –54 to –18 mV (tear: reference) and the voltage divider ratioFR ₀ decreased from 0.58 to 0.30, suggesting an increase in basolateral membrane electrical resistance. Additional indication of a basolateral membrane effect by the drug was that preincubation with 10⁵ m amphotericin B in Cl-free Ringer's did not eliminate the inhibitory effect of the drug on the Na- and K-elicited SCC. In the absence of amphotericin B in Cl-free Ringer's (SCC=0), 1 ×10³ m diltiazem depolarized theV _m from –78 to –9 mV suggesting that the increase in basolateral membrane resistance was due to K channel blockade. Diltiazem (1×10³ m) significantly decreased cyclic AMP content; however, isoproterenol in the presence of the drug increased cyclic AMP fourfold without having any restorative effect on the inhibited SCC. Therefore, the inhibition of the Cl-originated SCC resulting from an increase in basolateral membrane K resistance is not caused by a decline in cyclic AMP content. In plasma membrane-enriched fractions prepared from broken cell preparations of bovine corneal epithelium, 1×10³ m diltiazem had no inhibitory effects on either Na,K-ATPase or Ca,Mg-ATPase activities. These latter effects further point to the selectivity of diltiazem as an inhibitor of K-channel activity, but do not preclude a Ca-channel blocker effect by the drug in the micromolar range.     

Effects of calcium channel blockers on bupivacaine-induced toxicity

The purpose of this study was to investigate the influence of calcium channel blockers on bupivacaine-induced acute toxicity. For each of the three tested calcium channel blockers (diltiazem, verapamil and bepridil) 6 groups of mice were treated by two different doses, i.e. 2 and 10 mg/kg/i.p., or an equal volume of saline for the control group (n=20); 15 minutes later, all the animals were injected with a single 50 mg/kg/i.p. dose of bupivacaine. The convulsant activity, the time of latency to convulse and the mortality rate were assessed in each group. The local anesthetic-induced mortality was significantly increased by the three different calcium channel blockers. The convulsant activity of bupivacaine was not significantly modified but calcium channel blockers decreased the time of latency to obtain bupivacaine-induced convulsions; this effect was less pronounced with bepridil.     

Effects of calcium channel blockers on phototaxis and motility of Chlamydomonas reinhardtii

The effects of the four calcium channel blockers flunarizine, verapamil, diltiazem and nimodipine on motility and phototaxis of Chlamydomonas reinhardtio have been tested with a fully automated and computerized population system. Flunarizine inhibits motility transiently by causing the detachement of the flagella which, however, are regenerated during some hours. Phototaxis is inhibited to the same extent, but this is simply the result of the decreased motility and, hence, a non-specific effect. Verapamil causes also a detachement of the flagella with following regeneration, but in addition motility and phototaxis are inhibited by this drug to different extents, indicating the involvement of calcium channels in both processes. Diltiazem and nimodipine inhibit phototaxis without impairing motility, indicating that both processes are regulated in different ways. If diltiazem and nimodipine are applied simultaneously, no additive inhibitory effect can be observed. However, the combination of both blockers with verapamil causes and additive inhibitory effect as if verapamil is applied alone. By increasing the external calcium concentration from 10^-4 M to 10^-3 M the optimum of positive phototaxis is shifted to higher fluence rates. This shifting occurs also in the presence of channel blockers, but the strength of the positive reaction is influenced. These results point to the involvement of calcium channels in both phototaxis and motility, but simultaneously demonstrate the different sensitivity of the two processes to these drugs.Abbreviations DIL diltiazem (=benzothiazepine) - FLU flunarizine (=(E)-1-(bis-(4-fluorophenyl(methyl)-4-(3-phenyl-2-propenyl)piperazinex2HCl) - NIM nimodipine (=1,4-dihydropyridine) - VER verapamil (=diphenylalkylamine) CaM, calmodulin - PDE phosphodiesterase - DMSO dimethylsulfoxide     

Unusual responses of proboscis muscles ofBusycon canaliculatum to some calcium antagonist agents

Summary K- and ACh-induced responses of the radular sac, odontophore retractor, and radular retractor muscles ofBusycon canaliculatum were found to be strongly dependent upon [Ca]₀. Diltiazem had strong positive inotropic and chronotropic actions on fast twitch activity in the odontophore retractor and radular protractor muscles. K-induced tonic force in these muscles was partly inhibited by diltiazem but only at very high concentrations. ACh responses in all muscles were eliminated by diltiazem. Nifedipine enhanced fast twitches and tonic force in response to high K, and induced persistent spontaneous fast twitch discharges. Nifedipine inhibited ACh-induced tonic force, but induced rhythmic bursts of fast twitches persisting long after nifedipine washout. Verapamil strongly inhibited K- and ACh-induced tonic force in all three muscles at high concentration, but stimulated fast twitch responses and converted ACh contractures into fast twitch activity. Sucrose gap studies showed that nifedipine and diltiazem reduced K- and ACh-induced tension and depolarization. Paradoxically, verapamil reduced K- and ACh-induced tension but significantly enhanced their induced depolarizations. Diltiazem, nifedipine and verapamil did not act like slow Ca channel antagonists in these muscles. This may reflect differences in channel structure between molluscs and mammals, or differences in the cellular calcium release pathways operated by such channels in molluscan and mammalian muscle. These Ca-ant-agonists appeared to act as agonists of fast twitch activity in these muscles and antagonists of the ACh-induced calcium release pathway for tonic force development.     

Cladribine inhibits a diltiazem-induced increase in red blood cell purine nucleotide concentrations in a zebrafish model

Minimizing drug interactions is paramount to improving the efficacy and tolerability of cancer therapy. The zebrafish represents an innovative cancer model due to highly conserved genetics and inherent capacity for high-throughput chemical screening. This pilot study extends the utility of the zebrafish to a preclinical model for pharmacodynamics by examining the interaction of the nucleoside analogue, cladribine with the calcium channel blocker, diltiazem. Cladribine (0.7–3.5?mM) and/or diltiazem (2.4?mM), was injected intraperitoneally into adult zebrafish and red blood cell (RBC) lysates were assayed by HPLC for levels of purine nucleotides (e.g. ATP), potential biomarkers of cardiovascular health. Diltiazem increased RBC ATP concentrations, which were inhibited by co-injection of cladribine. These results suggest a novel drug interaction and highlight the feasibility of the zebrafish as an in vivo model for pharmacodynamic studies.     

Lack of melatonin response to acute administration of nifedipine and diltiazem in healthy men

Calcium antagonists have been shown to influence some endocrinological processes in mammals. The use of calcium channel blockers in clinical practice is well documented. The current study monitored nocturnal melatonin, prolactin, and cortisol levels in 19 healthy volunteers before and after administration of calcium channel blockers. The effect of nifedipine was tested in 9 subjects, while diltiazem was administered in 10 men. The nocturnal profile of the given parameters was studied between 23:00 and 05:00 h. At midnight (zero time), the participants were given placebo, nifedipine (in a sublingual dose of 20 mg) or diltiazem (in a single dose of 90 mg). The hypothesis that calcium channel blockers decrease nocturnal melatonin secretion has not been confirmed. The mean nocturnal levels of melatonin between 01:00 and 05:00 h were: 78.1+/-8.8 (control study) vs. 82.4+/-10.2 ng/l (nifedipine study) and 73.0+/-5.3 ng/l (control study) vs. 75.1+/-5.1 ng/l (diltiazem study). In conclusion, the calcium channel blockers used in this study do not alter the nocturnal melatonin secretory process in healthy men.     

Effects of acute and chronic nicotine on elevated plus maze in mice: involvement of calcium channels

The current experiments examined the anxiety-related effects of acute and repeated nicotine administration using the elevated plus maze test in mice. Nicotine (0.1 mg/kg s.c., 5 and 30 min after injection; 0.5 mg/kg, s.c., 5 min after injection) had an anxiogenic effect, shown by specific decreases in the percentage of time spent on the open arms and in the percentage of open arm entries. Tolerance developed to this anxiogenic action after 6 days of daily nicotine administration (0.1 mg/kg, s.c.). Five minutes after the seventh injection, an anxiolytic effect was observed, i.e., specific increases in the percentage of time spent on the open arms and in the percentage of open arm entries. L-type voltage-dependent calcium channel antagonists nimodipine (5 and 10 mg/kg, i.p.), flunarizine (5 and 10 mg/kg, i.p.), verapamil (5, 10, 20 mg/kg) and diltiazem (5, 10, 20 mg/kg, i.p.) were also injected prior to an acute low dose of nicotine or to each injection of chronic nicotine. Our results revealed that calcium channel blockers dose-dependently attenuated both an anxiogenic effect of nicotine as well as the development of tolerance to this effect. Our results suggest that neural calcium-dependent mechanisms are involved in the anxiety-related responses to acute and chronic nicotine injection that may ultimately lead to addiction and smoking relapse in human smokers.     

Effects of diltiazem on the calcium accumulation and atp synthesis simultaneously sustained by isolated rat heart mitochondria

1. The effects of diltiazem have been investigated in isolated rat heart mitochondria exposed to conditions possibly attained in ischemia-damaged cells.2. The results obtained indicate that diltiazem, at the concentrations expected within cells following pharmacological treatment, does not significantly affect the mitochondrial calcium content.3. Diltiazem did not appear to modify ATP synthesis, and hence the capacity of mitochondria to sustain the ATP-requiring processes needed for the recovery of cardiac cells.     

Modulation of the Frequency of Spontaneous Sarcoplasmic Reticulum Ca2+ Release Events (Ca2+ Sparks) by Myoplasmic [Mg2+] in Frog Skeletal Muscle

The modulation by internal free [Mg²⁺] of spontaneous calcium release events (Ca²⁺ “sparks”) from the sarcoplasmic reticulum (SR) was studied in depolarized notched frog skeletal muscle fibers using a laser scanning confocal microscope in line-scan mode (x vs. t). Over the range of [Mg²⁺] from 0.13 to 1.86 mM, decreasing the [Mg²⁺] induced an increase in the frequency of calcium release events in proportion to [Mg²⁺]^−1.6. The change of event frequency was not due to changes in [Mg-ATP] or [ATP]. Analysis of individual SR calcium release event properties showed that the variation in event frequency induced by the change of [Mg²⁺] was not accompanied by any changes in the spatiotemporal spread (i.e., spatial half width or temporal half duration) of Ca²⁺ sparks. The increase in event frequency also had no effect on the distribution of event amplitudes. Finally, the rise time of calcium sparks was independent of the [Mg²⁺], indicating that the open time of the SR channel or channels underlying spontaneous calcium release events was not altered by [Mg²⁺] over the range tested. These results suggest that in resting skeletal fibers, [Mg²⁺] modulates the SR calcium release channel opening frequency by modifying the average closed time of the channel without altering the open time. A kinetic reaction scheme consistent with our results and those of bilayer and SR vesicle experiments indicates that physiological levels of resting Mg²⁺ may inhibit channel opening by occupying the site for calcium activation of the SR calcium release channel.     

Fast activation of dihydropyridine-sensitive calcium channels of skeletal muscle. Multiple pathways of channel gating

Dihydropyridine (DHP) receptors of the transverse tubule membrane play two roles in excitation-contraction coupling in skeletal muscle: (a) they function as the voltage sensor which undergoes fast transition to control release of calcium from sarcoplasmic reticulum, and (b) they provide the conducting unit of a slowly activating L-type calcium channel. To understand this dual function of the DHP receptor, we studied the effect of depolarizing conditioning pulse on the activation kinetics of the skeletal muscle DHP-sensitive calcium channels reconstituted into lipid bilayer membranes. Activation of the incorporated calcium channel was imposed by depolarizing test pulses from a holding potential of -80 mV. The gating kinetics of the channel was studied with ensemble averages of repeated episodes. Based on a first latency analysis, two distinct classes of channel openings occurred after depolarization: most had delayed latencies, distributed with a mode of 70 ms (slow gating); a small number of openings had short first latencies, < 12 ms (fast gating). A depolarizing conditioning pulse to +20 mV placed 200 ms before the test pulse (-10 mV), led to a significant increase in the activation rate of the ensemble averaged-current; the time constant of activation went from tau m = 110 ms (reference) to tau m = 45 ms after conditioning. This enhanced activation by the conditioning pulse was due to the increase in frequency of fast open events, which was a steep function of the intermediate voltage and the interval between the conditioning pulse and the test pulse. Additional analysis demonstrated that fast gating is the property of the same individual channels that normally gate slowly and that the channels adopt this property after a sojourn in the open state. The rapid secondary activation seen after depolarizing prepulses is not compatible with a linear activation model for the calcium channel, but is highly consistent with a cyclical model. A six- state cyclical model is proposed for the DHP-sensitive Ca channel, which pictures the normal pathway of activation of the calcium channel as two voltage-dependent steps in sequence, plus a voltage-independent step which is rate limiting. The model reproduced well the fast and slow gating models of the calcium channel, and the effects of conditioning pulses. It is possible that the voltage-sensitive gating transitions of the DHP receptor, which occur early in the calcium channel activation sequence, could underlie the role of the voltage sensor and yield the rapid excitation-contraction coupling in skeletal muscle, through either electrostatic or allosteric linkage to the ryanodine receptors/calcium release channels.