Pharmacological regulators of intracellular calcium release channels

Intracellular Ca(2+) release channels, such as inositol 1,4,5-trisphosphate receptors (IP(3)Rs) and ryanodine receptors (RyRs), facilitate the release of Ca(2+) from intracellular storage organelles in response to extracellular and intracellular stimuli. Consequently, these large, tetrameric proteins play a central role in Ca(2+) signalling and Ca(2+) homeostasis in virtually all cells. Recent data suggests that intracellular Ca(2+) release channels may also have an important pathophysiological function in certain disease states, including cardiac arrhythmias and heart failure. As a result, there has been much interest in the identification and characterization of novel, selective regulators of these channels. In this article, we review the wide array of pharmacological agents that interact directly with intracellular Ca(2+) release channels and describe the mechanisms underlying their ability to modify channel function.     

The omega-atracotoxins: selective blockers of insect M-LVA and HVA calcium channels

The omega-atracotoxins (omega-ACTX) are a family of arthropod-selective peptide neurotoxins from Australian funnel-web spider venoms (Hexathelidae: Atracinae) that are candidates for development as biopesticides. We isolated a 37-residue insect-selective neurotoxin, omega-ACTX-Ar1a, from the venom of the Sydney funnel-web spider Atrax robustus, with high homology to several previously characterized members of the omega-ACTX-1 family. The peptide induced potent excitatory symptoms, followed by flaccid paralysis leading to death, in acute toxicity tests in house crickets. Using isolated smooth and skeletal nerve-muscle preparations, the toxin was shown to lack overt vertebrate toxicity at concentrations up to 1 microM. To further characterize the target of the omega-ACTXs, voltage-clamp analysis using the whole-cell patch-clamp technique was undertaken using cockroach dorsal unpaired median neurons. It is shown here for the first time that omega-ACTX-Ar1a, and its homolog omega-ACTX-Hv1a from Hadronyche versuta, reversibly block both mid-low- (M-LVA) and high-voltage-activated (HVA) insect calcium channel (Ca(v)) currents. This block occurred in the absence of alterations in the voltage-dependence of Ca(v) channel activation, and was voltage-independent, suggesting that omega-ACTX-1 family toxins are pore blockers rather than gating modifiers. At a concentration of 1 microM omega-ACTX-Ar1a failed to significantly affect global K(v) channel currents. However, 1 microM omega-ACTX-Ar1a caused a modest 18% block of insect Na(v) channel currents, similar to the minor block of Na(v) channels reported for other insect Ca(v) channel blockers such as omega-agatoxin IVA. These findings validate both M-LVA and HVA Ca(v) channels as potential targets for insecticides.     

Effects of calcium entry blockers not connected with calcium channels inhibition

We tested the effects of calcium entry blockers (verapamil, nifedipine, diltiazem) in comparison with nitroglycerin to investigate their inhibitory mechanisms of smooth muscle contraction not connected with the calcium blockade through calcium channels. The experiments were carried out by using isolated rabbit aorta strips in both normal and Ca2+-free solutions. Our results indicate that nitroglycerin and calcium entry blockers act on different calcium pools; the differences observed among calcium entry blockers suggest that other factors, different from calcium channels blockade (alpha-adrenergic block, unspecific effects), may be responsible for their inhibitory effect when high doses are employed.     

Substituted 1,2,3,4-tetrahydroaminonaphthols: antihypertensive agents, calcium channel blockers, and adrenergic receptor blockers with catecholamine-depleting effects

Substituted 1,2,3,4-tetrahydroaminonaphthols were found to be calcium channel blockers with antihypertensive properties. These compounds also possessed adrenergic beta-receptor blocking activity. From the structure-activity studies, no clear correlation emerged between the in vitro calcium channel blocking activity and the acute anti-hypertensive activity in cannulated spontaneously hypertensive rats. Extensive pharmacological testing of selected compounds indicated that aminonaphthols are antihypertensive agents with many pharmacological properties. The relative contribution of various pharmacological actions toward the observed antihypertensive activity is unclear. Since the clinically useful calcium channel blocker verapamil is structurally related to these compounds, one of the aminonaphthols, trans-3-[(3,3-diphenylpropyl)amino]-1,2,3,4-tetrahydro-6,7 -dimethoxy-2-naphthalenol (12), was compared with verapamil for calcium channel blocking activity, adrenergic blocking activity, and catecholamine-depleting activity. Both compounds were found to be equipotent in these test systems.     

Neuroprotection against ischemic/hypoxic brain damage: blockers of ionotropic glutamate receptor and voltage sensitive calcium channels

The growing number of cellular and molecular pathways believed to be involved in mechanisms of ischemic cell death in the brain has spurred a similar growth in the number of potential neuroprotective modalities, the majority of which are pharmacological in nature. Preventing or minimizing the first few steps in the cascade of events leading to ischemic cell death would have a more profound effect on the postischemic outcome than intervention at later steps in that cascade. This logic is, of course, at the heart of the urgency in providing the stroke or cardiac arrest patient with the earliest possible neuroprotective treatment. For the purpose of assessing potential neuroprotective modalities, the use of a well-established cerebral hypoxic/ischemic model system is a prerequisite. In our studies, we have used two major approaches, in vitro and in vivo. We evaluated both agonists and antagonists of ionotropic glutamate receptor channels (IGRC) and their effects in exacerbating and attenuating, respectively, the posthypoxic/ischemic outcome. Other drugs were tested for their ability to block the L-type voltage-sensitive calcium channels (VSCC), which are responsible for calcium influx and overload upon hypoxia/ischemia. These two membrane protein entities, the IGRC and the VSCC, are believed to be involved in the early stages of the cellular cascade that leads to the demise of neurons posthypoxia/ischemia. Some of the drugs were also tested for possible interaction with each other searching for possible synergy. These and other published studies in the field are reviewed here.     

Presynaptic alpha 2-adrenoceptor and kappa-opiate receptor occupancy promotes closure of neuronal (N-type) calcium channels

Synaptosomes prepared from rat cerebral cortex by homogenization in isotonic sucrose and centrifugation on four-step discontinuous percoll density gradients were loaded with the fluorescent indicator fura-2 to allow measurement of intrasynaptosomal free calcium concentrations [( Ca2+]i). Incubation of fura-2 loaded synaptosomes with either the kappa-opiate agonist U-50,488H (0.1-100 microM) or the alpha 2-adrenoceptor agonist clonidine (0.1-100 microM), resulted in a dose-dependent reduction in [Ca2+]i and these changes were completely antagonised by prior inclusion of naloxone (20 microM) or idazoxan (RX781094) (2 microM) respectively. When the 1,4-dihydropyridine Ca2+-channel blocker nifedipine (1 microM) was incubated with synaptosomes for 1 min, there was a 17.0% decrease in [Ca2+]i and when it was combined with either U-50,488H (1 microM) or clonidine (1 microM) there was a reduction in [Ca2+]i of 35.0 and 48.1% respectively i.e. the effects were additive. The increases in the depression of [Ca2+]i produced by these drug combinations were antagonised by the inclusion of naloxone (20 microM) or idazoxan (2 microM) which resulted in decreases in free [Ca2+]i of 26.5 and 14.1% respectively. These data indicate that the effects of clonidine and U-50,488H are not mediated by L-type Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Parkinsonian syndrome and calcium channel blockers

(1) Parkinsonian syndromes have occasionally been attributed to diltiazem, a calcium channel blocker, sometimes with a positive rechallenge. (2) Verapamil and amlodipine have also been implicated. (3) Prescribers should be aware of the possible role of these drugs in patients presenting with a parkinsonian syndrome.     

Toxin modulators and blockers of hERG K channels

The K⁺ channel encoded by the Ether-á-go-go-Related Gene (ERG) is expressed in different tissues of different animal species. There are at least three subtypes of this channel, being the sub-type 1 (ERG1) crucial in the repolarization phase of the cardiac action potential. Mutations in this gene can affect the properties of the channel producing the type II long QT syndrome (LQTS2) and many drugs are also known to affect this channel with a similar side effect. Various scorpion, spider and sea anemone toxins affect the ERG currents by blocking the ion-conducting pore from the external side or by modulating channel gating through binding to the voltage-sensor domain. By doing so, these toxins become very useful tools for better understanding the structural and functional characteristics of these ion channels. This review discusses the interaction between the ERG channels and the peptides isolated from venoms of these animals. Special emphasis is placed on scorpion toxins, although the effects of several spider venom toxins and anemone toxins will be also revised.     

Interactions between analgesics and calcium channel blockers

1. The findings, derived from different experimental models, examined in this review, provide evidence that the calcium channel blockers and related drugs possess analgesic effects. 2. The antinociceptive action that some analgesic drugs exhibit may be related to calcium channel blockade. 3. Evidence from a variety of biochemical and pharmacological experimental approaches, support the existence of an interelation between the calcium modulators and the opioid drugs. 4. This idea agrees with the novel neuropharmacological hypothesis that a common very high affinity binding site for multiple neurotransmitters could exist, as has been proposed by Pasternak and Wood (1986). 5. This hypothesis could be extended to the neuromodulators or other neuromediators.     

Selective modulation by membrane potential of the interaction of some calcium entry blockers with calcium channels in rat mesenteric artery.

1. The effects of flunarizine, (+)-PN 200-110 and nifedipine on [3H]-(+)-PN 200-110 specific binding were investigated in intact rat mesenteric arteries bathed in physiological solution or in KCl-depolarizing solution, and in a membrane fraction from rat mesenteric arteries. 2. Unlabelled dihydropyridines, (+)-PN 200-110 and nifedipine, inhibited [3H]-(+)-PN 200-110 specific binding concentration-dependently in polarized as well as in depolarized intact arteries. The Ki value of (+)-PN 200-110 was decreased in arteries bathed in KCl-depolarizing solution compared to arteries bathed in physiological solution, while the Ki value of nifedipine was not significantly changed. Ki values measured in depolarized arteries were close to the IC50 values (concentrations inhibiting by 50% the KCl-contraction of rat mesenteric artery). 3. Flunarizine (10(-6) M) was unable to displace the specific binding of [3H]-(+)-PN 200-110 in intact arteries bathed in physiological solution. At 10(-7) M-10(-6) M, it inhibited the binding in depolarized arteries, suggesting that prolonged depolarization is required for the interaction of flunarizine with the dihydropyridine receptor. 4. In a membrane fraction isolated from rat mesenteric arteries, (+)-PN 200-110, nifedipine and flunarizine were all able to displace completely the specific binding of [3H]-(+)-PN 200-110. Displacement curves were parallel and Hill coefficients were close to unity. Ki values were close to the values obtained in depolarized intact arteries.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation of prostacyclin biosynthesis by calcium entry blockers and extracellular calcium

The influence of variations in the availability of extracellular Ca2+ and of Ca2+-entry blockers on prostacyclin production by mesothelial cells in culture was studied. The Ca2+-entry blockers nifedipine and verapamil suppressed the basal, as well as the thrombin-, bradykinin-, and ionophore A23187-stimulated biosynthesis by about 50-60%, but high concentrations were required and the inhibition was never complete. Basal prostacyclin formation was unaffected by a Ca2+-poor buffer, but showed 50% reduction in the Ca2+-free buffer. Although the thrombin-stimulated prostacyclin formation was not significantly influenced by a Ca2+-poor or a Ca2+-free buffer, prostacyclin release stimulated by A23187 and bradykinin was diminished in the presence of these modified incubation media; the reduction of bradykinin stimulated biosynthesis was rather small (30%). These results suggest that the Ca2+ from intracellular stores is sufficient for half maximal stimulation of the phospholipases involved in the biosynthetic pathway of prostacyclin and that--depending on the nature of the stimulus--different phospholipases are activated with varying requirements for free Ca2+.     

几种钾通道阻滞剂的心脏效应

心脏Ito、IK1和IK 通道的分布具有明显的种属差异和组织差异 ,IK 通道至少存在有 3种亚型 ,即IKur、IKr和IKs。Ito、IK1和IK 通道阻滞剂多具有正性肌力作用和负性频率作用。与Ⅰ类抗心律失常药不同 ,IKr通道阻滞剂对迷走神经兴奋所致的心脏电生理效应具有显著的选择性抑制或增强作用。Ito抑制剂选择性地延长心房肌动作电位时程 ;IK 抑制剂延长哺乳动物心房和心室肌有效不应期的作用不同。逆向使用依赖性是目前临床应用的Ⅲ类抗心律失常药物 (主要作用于IKr)主要缺点 ,它可能与心率加快时IKs的累积有关。并用IKs阻滞剂或并用 β受体拮抗剂可能提高Ⅲ类抗心律失常药的疗效 ,避免尖端扭转型室性心动过速的发生。     

钙通道阻滞剂的临床应用

钙通道阻滞剂(CCB)能阻滞钙离子进入细胞内,引起各组织系统的平滑肌松弛,减轻缺血性细胞中毒。CCB在结构上主要分四类:二氢吡啶类、硫氮(艹卓)酮类、异搏定类、氟桂嗪类。 二氢吡啶类常用的有非洛地平(波依定)、尼索地平、硝苯地平、尼莫地平、氨氯地平(络活喜)、尼群地平等。硫氮(艹卓)酮类有地尔硫(艹卓)(恬尔心)等。维拉帕米类有维拉帕米(异搏定)等。氟桂嗪类有氟桂利嗪(西比灵)等。 本文对CCB的应用作一概述。 1 治疗心肌缺血和心绞痛 短期急性心肌缺血会导致代谢紊乱,包括ATP耗竭,细胞外钾离子积聚,细胞内钠离子积聚而失去细胞内对钙离子转运的正常机制,结果增加了细胞内钙离子的水平,厌氧代谢变成需氧代谢,糖原贮存被耗竭,伴酸中毒和线粒体