Synergy between intrathecal omega-conotoxin CVID and dexmedetomidine to attenuate mechanical hypersensitivity in the rat

The analgesic effects of intrathecal (i.t.) omega-conotoxin CVID, an N-type Ca2+ channel antagonist, and the alpha2-adrenoceptor agonist, dexmedetomidine, were tested alone and in combination following unilateral ligation of L (lumbar) 5/6 spinal nerves in rats. Mechanical allodynia was observed prior to insertion of an i.t. catheter. Effects and interactions of omega-conotoxin CVID (0.01-10 microg/kg) and dexmedetomidine (0.1-10 microg/kg) were tested on allodynic and tail flick (thermal stimulus) responses. Only dexmedetomidine increased the latency of the tail flick response. Both dexmedetomidine and omega-conotoxin CVID completely inhibited allodynia (ED50 0.78+/-0.02 and 0.35+/-0.08 microg/kg, respectively; n=63, 41). Dexmedetomidine and omega-conotoxin CVID combined in dose ratios 0.7 and 1.3 (adjusted for ED50) were synergistic in decreasing mechanical hypersensitivity; interaction index (gamma) 0.39 (confidence interval [CI] 0.33, 0.46) and 0.3 (CI 0.23, 0.38). Despite the necessity for i.t. administration, these data suggest that the synergistic combination confers enhanced potency (lower doses) of both drugs that may avoid clinical toxicity of single drug therapy.     

Anti-nociceptive synergism of morphine and gabapentin in neuropathic pain induced by chronic constriction injury

In order to detect an anti-nociceptive interaction between morphine and gabapentin, the anti-allodynic and anti-hyperalgesic effects of these drugs, administered either separately or in combination, were determined using the von Frey and acetone tests in a rat model of neuropathic pain (Bennett model). Morphine and gabapentin individually induced moderate attenuation of mechanical hyperalgesia, whereas the morphine and gabapentin combination completely decreased hyperalgesia. Morphine showed its maximal effect at 30 min post-injection in the acetone test; however, this effect gradually returned to the baseline value. Gabapentin did not produce an anti-allodynic effect, whereas the morphine and gabapentin combination completely decreased allodynia behavior at 30 min post-injection, an effect that persisted until 120 min. The area under the curve (AUC) of the anti-allodynic or anti-hyperalgesic effects produced by the combinations were significantly greater than the theoretical sum of effects produced by each drug alone or similar to the theoretical sum. The analysis of the effect, expressed as the AUC of the time course, supports the hypothesis that the combination of these drugs is useful in neuropathic pain therapy.     

鞘内联合应用吗啡和氯胺酮对神经痛大鼠脊髓P物质表达的影响

目的研究鞘内联合应用吗啡和氯胺酮对神经痛大鼠脊髓P物质表达的影响。方法12只Wistar大鼠,体质量220~260 g,制备坐骨神经结扎模型并鞘内置管,随机分为4组(n=3):B组为空白对照组;C组鞘内注射生理盐水10μL;M组鞘内注射吗啡20μg;KM组鞘内注射吗啡10μg和氯胺酮25μg。坐骨神经结扎术后第4天开始鞘内给药,1次/d,连续7 d。用药7 d后处死大鼠,取腰段脊髓,免疫组化法测定脊髓P物质的含量。结果C组P物质表达明显低于B组(P<0.05)和KM组(P<0.01);KM组P物质表达明显高于M组(P<0.05)。结论鞘内联合应用吗啡和氯胺酮的抗伤害性作用,可能与其增加脊髓背角P物质的含量有关;脊髓背角P物质含量减少,可能介导慢性坐骨神经损伤后的痛敏持续状态。     

Minocycline and pentoxifylline attenuate allodynia and hyperalgesia and potentiate the effects of morphine in rat and mouse models of neuropathic pain

Recent research has shown that microglial cells which are strongly activated in neuropathy can influence development of allodynia and hyperalgesia. Here we demonstrated that preemptive and repeated i.p., administration (16 h and 1 h before injury and then after nerve ligation twice daily for 7 days) of minocycline (15; 30; 50 mg/kg), a potent inhibitor of microglial activation, significantly attenuated the allodynia (von Frey test) and hyperalgesia (cold plate test) measured on day 3, 5, 7 after chronic constriction injury (CCI) in rats. Moreover, the 40% improvement of motor function was observed. In mice, i.p., administration of minocycline (30 mg/kg) or pentoxifylline (20 mg/kg) according to the same schedule also significantly decreased allodynia and hyperalgesia on day 7 after CCI. Antiallodynic and antihyperalgesic effect of morphine (10 mg/kg; i.p.) was significantly potentiated in groups preemptively and repeatedly injected with minocycline (von Frey test, 18 g versus 22 g; cold plate test, 13 s versus 20 s in rats and 1.2 g versus 2.2 g; 7.5 s versus 10 s in mice; respectively) or pentoxifylline (1.3 g versus 3 g; 7.6 s versus 15 s in mice; respectively). Antiallodynic and antihyperalgesic effect of morphine (30 μg; i.t.) given by lumbar puncture in mice was also significantly potentiated in minocycline-treated group (1.2 g versus 2.2 g; 7.5 s versus 11 s; respectively). These findings indicate that preemptive and repeated administration of glial inhibitors suppresses development of allodynia and hyperalgesia and potentiates effects of morphine in rat and mouse models of neuropathic pain.     

Post-injury repeated administrations of minocycline improve the antinociceptive effect of morphine in chronic constriction injury model of neuropathic pain in rat

It is confirmed that pharmacological attenuation of glial cells can alleviate neuropathic pain by lowering proinflammatory cytokine expression. The present study tries to confirm that post-injury administration of glia inhibitor, minocycline, can attenuate the neuropathic pain symptoms and improves the efficacy of morphine anti-nociception in chronic constriction injury (CCI). Male Wistar rats (230-270 g) underwent surgery for induction CCI model of neuropathy. For assessment of the thermal hyperalgesia and mechanical allodynia after CCI induction, morphine (2.5, 5, 7.5, 10 and 15 mg/kg; s.c.) and saline were administered on post-operative days (PODs) 0, 6 and 14. Hargreaves and Von-Frey tests were performed before and 30 min after morphine administration, respectively. The results showed significant decrease in antinociceptive effect of morphine on POD 6 compared to POD 0 only at the dose of 5 mg/kg. On the other hand, on POD 14 the antinociceptive effect of morphine (5, 7.5, 10 and 15 mg/kg) significantly decreased in comparison with POD 0. In another set of experiments, animals received minocycline (10, 20 and 40 mg/kg; i.p.) for eight days from POD 6 to 13 and then the antinociceptive effect of single dose of morphine 5 mg/kg was tested on POD 14. Behavioral tests showed that minocycline (40 mg/kg) could effectively attenuate the thermal hyperalgesia and mechanical allodynia on POD 13. Moreover, minocycline (40, 20 mg/kg) improved the anti-hyperalgesic, and minocycline (40 mg/kg) improved the anti-allodynic effects of morphine 5 mg/kg on POD 14. It seems that the reduction of antinociceptive effect of morphine after CCI may be mediated through glia activation. Modulation of glial activity by minocycline can attenuate CCI-induced neuropathic pain. It is also shown that repeated post-injury administration of minocycline improves the antinociceptive effect of morphine in neuropathic pain.     

Potentiation of morphine antiallodynic efficacy by ACPT-III, a Group III metabotropic glutamate receptor agonist, in rat spinal nerve ligation-induced neuropathic pain

Despite the importance of spinal metabotropic glutamate receptors (mGluRs) and opioid receptors in nociceptive processing, the roles of these receptors in the modulation of neuropathic pain at the spinal level have not been thoroughly investigated. The purpose of this study was to investigate the effects of spinal mGluR agents and opioids (morphine) on neuropathic pain. Male Sprague-Dawley rats underwent L5 and L6 spinal nerve ligation to induce neuropathic pain and intrathecal catheterization for drug administration. A paw-withdrawal threshold to mechanical stimulus was measured using the “up and down” method. When administered intrathecally, neither Group I mGluR antagonists nor Group II or III agonists modified the withdrawal threshold after spinal nerve ligation. Intrathecal administration of morphine dose-dependently increased the withdrawal threshold. Whereas ACPT-III, a Group III mGluR agonist, enhanced the antiallodynic action of morphine, other mGluR agents did not. Collectively, mGluRs may not directly modulate the processing of spinal nerve ligation-induced neuropathic pain at the spinal level. However, Group III mGluR agonists in the spinal cord may indirectly contribute to the potentiation of morphine antiallodynia, indicating that these agonists might be used as adjuvants for spinal morphine.     

Calcium channel dysfunction in inferior colliculus neurons of the genetically epilepsy-prone rat

Voltage-gated calcium (Ca²⁺) channels are thought to play an important role in epileptogenesis and seizure generation. Here, using the whole cell configuration of patch-clamp techniques, we report on the modifications of biophysical and pharmacological properties of high threshold voltage-activated Ca²⁺ channel currents in inferior colliculus (IC) neurons of the genetically epilepsy-prone rats (GEPR-3s). Ca²⁺ channel currents were measured by depolarizing pulses from a holding potential of −80 mV using barium (Ba²⁺) as the charge carrier. We found that the current density of high threshold voltage-activated Ca²⁺ channels was significantly larger in IC neurons of seizure-naive GEPR-3s compared to control Sprague-Dawley rats, and that seizure episodes further enhanced the current density in the GEPR-3s. The increased current density was reflected by both a −20 mV shifts in channel activation and a 25% increase in the non-inactivating fraction of channels in seizure-naive GEPR-3s. Such changes were reduced by seizure episodes in the GEPR-3s. Pharmacological analysis of the current density suggests that upregulation of L-, N- and R-type of Ca²⁺ channels may contribute to IC neuronal hyperexcitability that leads to seizure susceptibility in the GEPR-3s.     

Bile Acid Inhibition of N-type Calcium Channel Currents from Sympathetic Ganglion Neurons

Under some pathological conditions as bile flow obstruction or liver diseases with the enterohepatic circulation being disrupted, regurgitation of bile acids into the systemic circulation occurs and the plasma level of bile acids increases. Bile acids in circulation may affect the nervous system. We examined this possibility by studying the effects of bile acids on gating of neuronal (N)-type Ca(2+) channel that is essential for neurotransmitter release at synapses of the peripheral and central nervous system. N-type Ca(2+) channel currents were recorded from bullfrog sympathetic neuron under a cell-attached mode using 100 mM Ba(2+) as a charge carrier. Cholic acid (CA, 10(-6) M) that is relatively hydrophilic thus less cytotoxic was included in the pipette solution. CA suppressed the open probability of N-type Ca(2+) channel, which appeared to be due to an increase in null (no activity) sweeps. For example, the proportion of null sweep in the presence of CA was ~40% at +40 mV as compared with ~8% in the control recorded without CA. Other single channel properties including slope conductance, single channel current amplitude, open and shut times were not significantly affected by CA being present. The results suggest that CA could modulate N-type Ca(2+) channel gating at a concentration as low as 10(-6) M. Bile acids have been shown to activate nonselective cation conductance and depolarize the cell membrane. Under pathological conditions with increased circulating bile acids, CA suppression of N-type Ca(2+) channel function may be beneficial against overexcitation of the synapses.     

Gabapentin inhibits presynaptic Ca(2+) influx and synaptic transmission in rat hippocampus and neocortex

Gabapentin is a widely used drug with anticonvulsant, antinociceptive and anxiolytic properties. Although it has been previously shown that Gabapentin binds with high affinity to the alpha(2)delta subunit of voltage-operated Ca(2+) channels (VOCC), little is known about the functional consequences of this interaction. Here, we investigated the effect of Gabapentin on VOCCs and synaptic transmission in rat hippocampus and neocortex using whole-cell patch clamp and confocal imaging techniques. Gabapentin (100-300 microM) did not affect the peak amplitude or voltage-dependency of VOCC currents recorded from either dissociated or in situ neocortical and hippocampal pyramidal cells. In contrast, Gabapentin inhibited K(+)-evoked increases in [Ca(2+)] in a subset of synaptosomes isolated from rat hippocampus and neocortex in a dose-dependent manner, with an apparent half-maximal inhibitory effect at approximately 100 nM. In hippocampal slices, Gabapentin (300 microM) inhibited the amplitude of evoked excitatory- and inhibitory postsynaptic currents recorded from CA1 pyramidal cells by 30-40%. Taken together, the results suggest that Gabapentin selectively inhibits Ca(2+) influx by inhibiting VOCCs in a subset of excitatory and inhibitory presynaptic terminals, thereby attenuating synaptic transmission.     

Effects of the endogenous cannabinoid anandamide on voltage-dependent sodium and calcium channels in rat ventricular myocytes

BACKGROUND AND PURPOSE

The endocannabinoid anandamide (N-arachidonoyl ethanolamide; AEA) exerts negative inotropic and antiarrhythmic effects in ventricular myocytes.

EXPERIMENTAL APPROACH

Whole-cell patch-clamp technique and radioligand-binding methods were used to analyse the effects of anandamide in rat ventricular myocytes.

KEY RESULTS

In the presence of 1–10 μM AEA, suppression of both Na⁺ and L-type Ca²⁺ channels was observed. Inhibition of Na⁺ channels was voltage and Pertussis toxin (PTX) – independent. Radioligand-binding studies indicated that specific binding of [³H] batrachotoxin (BTX) to ventricular muscle membranes was also inhibited significantly by 10 μM metAEA, a non-metabolized AEA analogue, with a marked decrease in B_max values but no change in K_d. Further studies on L-type Ca²⁺ channels indicated that AEA potently inhibited these channels (IC₅₀ 0.1 μM) in a voltage- and PTX-independent manner. AEA inhibited maximal amplitudes without affecting the kinetics of Ba²⁺ currents. MetAEA also inhibited Na⁺ and L-type Ca²⁺ currents. Radioligand studies indicated that specific binding of [³H]isradipine, was inhibited significantly by metAEA. (10 μM), changing B_max but not K_d.

CONCLUSION AND IMPLICATIONS

Results indicate that AEA inhibited the function of voltage-dependent Na⁺ and L-type Ca²⁺ channels in rat ventricular myocytes, independent of CB₁ and CB₂ receptor activation.     

Ambroxol, a Nav1.8-preferring Na(+) channel blocker, effectively suppresses pain symptoms in animal models of chronic, neuropathic and inflammatory pain

Neuropathic pain affects many patients, and treatment today is far from being perfect. Nav1.8 Na⁺ channels, which are expressed by small fibre sensory neurons, are promising targets for novel analgesics. Na⁺ channel blockers used today, however, show only limited selectivity for this channel subtype, and can cause dose-limiting side effects. Recently, the secretolytic ambroxol was found to preferentially inhibit Nav1.8 channels. We used this compound as a tool to investigate whether a Nav1.8-preferring blocker can suppress symptoms of chronic, neuropathic and inflammatory pain in animal models. The drug was tested in the formalin paw model, two models of mononeuropathy, and a model of monoarthritis in rats. Ambroxol's effects were compared with those of gabapentin. Ambroxol at a dose of 1 g/kg had to be administered to rats to achieve the plasma levels that are reached in clinical use (for the treatment of infant and acute respiratory distress syndrome). Ambroxol (1 g/kg) was only weakly effective in models for acute pain, but effectively reduced pain symptoms in all other models; in some cases it completely reversed pain behaviour. In most cases the effects were more pronounced than those of gabapentin (at 100 mg/kg). These data show that a Nav1.8-preferring Na⁺ channel blocker can effectively suppress pain symptoms in a variety of models for chronic, neuropathic and inflammatory pain at plasma levels, which can be achieved in the clinic.     

Novel sodium channel antagonists in the treatment of neuropathic pain

Introduction: Effective and safe drugs for the treatment of neuropathic pain are still an unmet clinical need. Neuropathic pain, caused by a lesion or disease that affects the somatosensory system, is a debilitating and hampering condition that has a great economic cost and, above all, a tremendous impact on the quality of life. Sodium channels are one of the major players in generating and propagating action potentials. They represent an appealing target for researchers involved in the development of new and safer drugs useful in the treatment of neuropathic pain. The actual goal for researchers is to target sodium channels selectively to stop the abnormal signaling that characterizes neuropathic pain while leaving normal somatosensory functions intact.

Areas covered: This review covers the most recent publications regarding sodium channel blockers and their development as new treatments for neuropathic pain. The main areas discussed are the natural sources of new blockers, such as venom extracts and the recent efforts from many pharmaceutical companies in the field.

Expert opinion: There have been serious efforts by both the pharmaceutical industry and academia to develop new and safer therapeutic options for neuropathic pain. A number of different strategies have been undertaken; the main efforts directed towards the identification of selective blockers starting from both natural products or screening chemical libraries. At this time, researchers have identified and characterized selective compounds against Na_V1.7 or Na_V1.8 voltage-gated sodium channels but only time will tell if they reach the market.     

T-type voltage-gated calcium channels as targets for the development of novel pain therapies

It is well recognized that voltage-gated calcium (Ca(2+)) channels modulate the function of peripheral and central pain pathways by influencing fast synaptic transmission and neuronal excitability. In the past, attention focused on the modulation of different subtypes of high-voltage-activated-type Ca(2+) channels; more recently, the function of low-voltage-activated or transient (T)-type Ca(2+) channels (T-channels) in nociception has been well documented. Currently, available pain therapies remain insufficient for certain forms of pain associated with chronic disorders (e.g. neuropathic pain) and often have serious side effects. Hence, the identification of selective and potent inhibitors and modulators of neuronal T-channels may help greatly in the development of safer, more effective pain therapies. Here, we summarize the available information implicating peripheral and central T-channels in nociception. We also discuss possible future developments aimed at selective modulation of function of these channels, which are highly expressed in nociceptors.     

Suppression of Peripheral Sympathetic Activity Underlies Protease-Activated Receptor 2-Mediated Hypotension

Protease-activated receptor (PAR)-2 is expressed in endothelial cells and vascular smooth muscle cells. It plays a crucial role in regulating blood pressure via the modulation of peripheral vascular tone. Although some reports have suggested involvement of a neurogenic mechanism in PAR-2-induced hypotension, the accurate mechanism remains to be elucidated. To examine this possibility, we investigated the effect of PAR-2 activation on smooth muscle contraction evoked by electrical field stimulation (EFS) in the superior mesenteric artery. In the present study, PAR-2 agonists suppressed neurogenic contractions evoked by EFS in endothelium-denuded superior mesenteric arterial strips but did not affect contraction elicited by the external application of noradrenaline (NA). However, thrombin, a potent PAR-1 agonist, had no effect on EFS-evoked contraction. Additionally, ω-conotoxin GVIA (CgTx), a selective N-type Ca²⁺ channel (I_Ca-N) blocker, significantly inhibited EFS-evoked contraction, and this blockade almost completely occluded the suppression of EFS-evoked contraction by PAR-2 agonists. Finally, PAR-2 agonists suppressed the EFS-evoked overflow of NA in endothelium-denuded rat superior mesenteric arterial strips and this suppression was nearly completely occluded by ω-CgTx. These results suggest that activation of PAR-2 may suppress peripheral sympathetic outflow by modulating activity of I_Ca-N which are located in peripheral sympathetic nerve terminals, which results in PAR-2-induced hypotension.     

KST5468, a new T-type calcium channel antagonist, has an antinociceptive effect on inflammatory and neuropathic pain models

The T-type Ca²⁺ channel is a low-voltage-activated Ca²⁺ channel related to nociceptive stimuli. Increases in Ca²⁺ due to calcium channel activation enhance pain sensitivity through both peripheral and central pain pathways. We have developed a novel compound, KST5468, which is a T-type calcium channel antagonist. The new synthetic compound may have an antinociceptive effect, and thus we evaluated KST5468 as a putative analgesic in a hot plate test, a formalin test, and two neuropathic pain models. KST5468 caused a significant increase in latency in the hot plate test at 30 min after a 10 mg/kg peritoneal injection of the compound. Interestingly, in the second phase of formalin test, KST5468 decreased pain behaviors in a dose-dependent manner. Moreover, in two neuropathic pain models induced by chronic constriction and spared nerve injury, KST5468 significantly increased the mechanical pain threshold. Using immunohistochemistry, expression of two well known pain-related molecular markers, c-Fos and calcitonin gene-related peptide (CGRP), and phosphorylated extracellular signal-related kinase (p-ERK) were found to be decreased in the laminae I-II layers of the ipsilateral L4-L5 spinal dorsal horn in KST5468 treated mice. Taken together, the results of this study suggest that KST5468 may be an effective antinociceptive agent for neuropathic pain.     

Electrophysiological effects of anandamide on rat myocardium

Background and purpose:

The endocannabinoid, anandamide, has anti-arrhythmic effects. The aim of the present study was to explore the electrophysiological effects of anandamide on rat myocardium.

Experimental approach:

Evoked action potentials (APs) were recorded using intracellular recording technique in rat cardiac papillary muscles. In addition, L-type Ca²⁺ current was measured and analysed using whole-cell patch-clamp recording technique in isolated rat cardiac ventricular myocytes.

Key results:

In cardiac papillary muscles, anandamide (1, 10, 100 nM) decreased AP duration in a concentration-dependent manner. Furthermore, 100 nM anandamide decreased AP amplitude, overshoot and V_max in partially depolarized papillary muscles. These effects were abolished by AM251 (100 nM), a selective antagonist for CB₁ receptors, but not AM630 (100 nM), a CB₂ receptor antagonist. Furthermore, an agonist of L-type Ca²⁺ channels, Bay K 8644 (0.5 µM), a K⁺ channel blocker tetraethylammonium chloride (20 mM) and the nitric oxide synthase inhibitor l-NAME (1 mM) had no effect on anandamide-induced decrease in AP duration. In isolated ventricular myocytes, anandamide (1, 10, 100 nM) decreased L-type Ca²⁺ current concentration-dependently, and shifted the current–voltage relationship curve of the Ca²⁺ current. Anandamide (100 nM) shifted the steady-state inactivation curve to the left and the recovery curve to the right. Blockade of CB₁ receptors with AM251 (100 nM), but not CB₂ receptors with AM630 (100 nM), eliminated the effect of anandamide on L-type Ca²⁺ currents.

Conclusions and implications:

These data suggest that anandamide suppressed AP and L-type Ca²⁺ current in cardiac myocytes through CB₁ receptors.     

Implication of the signal transduction pathways in the enhancement of noradrenaline turnover induced by morphine withdrawal in the heart

Our previous studies have shown an enhanced activity of the noradrenergic system in the heart in rats withdrawn from morphine. In the current study, we examined the role of protein kinase A, protein kinase C and Ca(2+) entry through L-type Ca(2+) channels in naloxone-precipitated increase turnover of noradrenaline in the right and left ventricle. Chronic pretreatment for 7 days with the selective protein kinase A inhibitor, HA-1004 (N-(2' guanidinoethyl)-5-isoquinolinesulfonamide) concomitantly with morphine significantly antagonized the increase in normetanephrine/noradrenaline ratio (an index of noradrenaline turnover) observed in morphine withdrawn rats. However, the infusion of calphostin C (2-(12-(2-(benzoyloxy)propyl)-3,10-dihydro-4,9-dihydroxy-2,6,7,11-tetramethoxy-3,10-dioxo-1-perylenyl)-1 methylethy carbonic acid 4-hydroxyphenyl ester, a selective protein kinase C inhibitor) did not modify the morphine withdrawal-induced increase in noradrenaline turnover. In addition, when the selective L-type Ca(2+) channel antagonist, nimodipine, was infused it diminished the increased in noradrenaline turnover observed after naloxone administration to morphine dependent rats. Taken together, these data might indicate that protein kinase A activity is necessary for the enhancement of noradrenaline turnover during morphine withdrawal and that an up-regulated Ca(2+) system might contribute to the increase of noradrenaline turnover. The present finding suggests that protein kinase A and Ca(2+) influx through L-type Ca(2+) channels might contribute to the activation of noradrenergic system in the heart observed during morphine withdrawal.     

Long-term blockade of L/N-type Ca2+ channels by cilnidipine ameliorates repolarization abnormality of the canine hypertrophied heart

Background and purpose:

The heart of the canine model of chronic atrioventricular block is known to have a ventricular electrical remodelling, which mimics the pathophysiology of long QT syndrome. Using this model, we explored a new pharmacological therapeutic strategy for the prevention of cardiac sudden death.

Experimental approach:

The L-type Ca²⁺ channel blocker amlodipine (2.5 mg·day⁻¹), L/N-type Ca²⁺ channel blocker cilnidipine (5 mg·day⁻¹), or the angiotensin II receptor blocker candesartan (12 mg·day⁻¹) was administered orally to the dogs with chronic atrioventricular block for 4 weeks. Electropharmacological assessments with the monophasic action potential (MAP) recordings and blood sample analyses were performed before and 4 weeks after the start of drug administration.

Key results:

Amlodipine and cilnidipine decreased the blood pressure, while candesartan hardly affected it. The QT interval, MAP duration and beat-to-beat variability of the ventricular repolarization period were shortened only in the cilnidipine group, but such effects were not observed in the amlodipine or candesartan group. Plasma concentrations of adrenaline, angiotensin II and aldosterone decreased in the cilnidipine group. In contrast, plasma concentrations of angiotensin II and aldosterone were elevated in the amlodipine group, whereas in the candesartan group an increase in plasma levels of angiotensin II and a decrease in noradrenaline and adrenaline concentrations were observed.

Conclusions and implications:

Long-term blockade of L/N-type Ca²⁺ channels ameliorated the ventricular electrical remodelling in the hypertrophied heart which causes the prolongation of the QT interval. This could provide a novel therapeutic strategy for the treatment of cardiovascular diseases.     

Mechanisms underlying the slow onset of action of a new dihydropyridine,NZ-105, on a cultured smooth muscle cell line

Summary The inhibitory effect of a new dihydropyridine derivative, (±)-2-[benzyl(phenyl)amino]ethyl-1,4-dihydro-2,6-dimethyl-5-(5,5-dimethyl-2-oxo-1,3,2-dioxaphosphorinan-2-yl)-4-(3-nitrophenyl)-3-pyridinecarboxylate hydrochloride (NZ-105), on whole cell Ca²⁺ current (I_Ca) in cultured vascular smooth muscle cells was investigated with the patch clamp technique. NZ-105 blocked I_Ca in a concentration-dependent manner when the command pulse ranged from +10 mV to –50 mV. The inhibitory effect of NZ-105 appeared at concentrations higher than 10 mol/l and it blocked I_Ca completely at a concentration of 1 nmol/l. The concentration which produced the half-maximal inhibitory effect was estimated to be around 20 mol/l. NZ-105 (500 pmol/l) completely blocked I_Ca elicited by depolarization to + 10 mV at a holding potential of –40 mV, whereas it blocked I_Ca by only 67% at a holding potential of –90 mV. NZ-105 (100 mol/l) shifted the steady-state inactivation curve by 40 mV to more negative potentials without affecting its slope factor. The blocking time constant of 500 mol/l NZ-105 was 57.6 + 9.9 s at a holding potential of –70 mV. These results indicate that NZ-105 has characteristics typical of dihydropyridines and binds to Ca²⁺ channels of vascular smooth muscle cells with a high affinity. They also suggested that the slow onset of its action is due to the slow binding of the drug to Ca²⁺ channels. Send offprint requests to S. Kokubun at the above address     

Effects of diltiazem,a Ca2+ channel blocker,on naloxone-precipitated changes in dopamine and its metabolites in the brains of opioid-dependent rats

The effects of diltiazem, an L-type Ca²⁺ channel blocker, on naloxone (an opioid receptor antagonist)-precipitated withdrawal signs and changes in extracellular levels of dopamine (DA) and its metabolites in various brain regions of morphine (a -opioid receptor agonist) or butorphanol (a// mixed opioid receptor agonist) dependent rats were investigated using high performance liquid chromatography fitted with an electrochemical detector (HPLC-ED). Rats were rendered opioid-dependent by continuous intracerebroventricular (ICV) infusion with morphine (26 nmol/µl per h) or butorphanol (26 nmol/µ1 per h) for 3 days. The expression of physical dependence produced by these opioids, as evaluated by naloxone (5 mg/kg, IP)-precipitated withdrawal signs, was reduced by concomitant infusion of diltiazem (10 and 100 nmol/µl per h). Under the same condition, naloxone decreased the levels of: DA in the cortex, striatum, and midbrain; 3,4-dihydroxyphenylacetic acid (DOPAC) in the cortex, striatum, limbic areas, and midbrain; and homovanilic acid (HVA) in the striatum, limbic areas, and midbrain regions. In animals rendered dependent on butorphanol, the results obtained were similar to those of morphine-dependent rats except for the changes in DOPAC levels. Furthermore, concomitant infusion of diltiazem and opioids blocked the decreases in levels of DA, DOPAC, and HVA in a dosedependent manner. These results suggest that the augmentation of intracellular Ca²⁺ mediated through L-type Ca²⁺ channels during continuous opioid infusion results in a decrease in extracellular levels of DA and its metabolites in some specific regions, which are intimately involved in the expression of withdrawal syndrome precipitated by naloxone.