Anti-Bronchospasmodic Effect of JME-173, a Novel Mexiletine Analog Endowed With Highly Attenuated Anesthetic Activity.

Local anesthetics (LAs), such as lidocaine and mexiletine, inhibit bronchoconstriction in asthmatics, but adverse effects limit their use for this specific clinical application. In this study, we describe the anti-spasmodic properties of the mexiletine analog 2-(2-aminopropoxy)-3,5-dimethyl, 4-Br-benzene (JME-173), which was synthesized and screened for inducing reduced activity on Na+ channels. The effectiveness of JME-173 was assessed using rat tracheal rings, a GH3 cell line and mouse cardiomyocytes to access changes in smooth muscle contraction, and Na+, and Ca++ionic currents, respectively. Bronchospasm and airway hyper-reactivity (AHR) were studied using whole-body barometric plethysmography in A/J mice. We observed that the potency of JME-173 was 653-fold lower than mexiletine in inhibiting Na+ currents, but 12-fold higher in inhibiting L-type Ca++ currents. JME-173 was also more potent than mexiletine in inhibiting tracheal contraction by carbachol, allergen, extracellular Ca++, or sodium orthovanadate provocations. The effect of JME-173 on carbachol-induced tracheal contraction remained unaltered under conditions of de-epithelized rings, ß2-receptor blockade or adenylate cyclase inhibition. When orally administered, JME-173 and theophylline inhibited methacholine-induced bronchospasm at time points of 1 and 3 h post-treatment, while only JME-173 remained active for at least 6 h. In addition, JME-173 also inhibited AHR in a mouse model of lipopolysaccharide (LPS)-induced lung inflammation. Thus, the mexiletine analog JME-173 shows highly attenuated activity on Na+ channels and optimized anti-spasmodic properties, in a mechanism that is at least in part mediated by regulation of Ca++ inflow toward the cytosol. Thus, JME-173 is a promising alternative for the treatment of clinical conditions marked by life-threatening bronchoconstriction.

Hydroalcoholic extract from Nerium oleander L. (Apocynaceae) elicits arrhythmogenic activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Nerium oleander L. (OLE) has been used medicinally and is reported to possess a wide range of pharmacological activities. OLE effects are caused by different cardiac glycosides (CG), primarily oleandrin, found within the plant. CG can potentially impair sodium-potassium ATPase (NKA) pump activity and cause positive inotropic effects on the heart. AIM OF THE STUDY: The aim of this study was to investigate the potential arrhythmogenic effects of hydroalcoholic extracts from N. oleander (OLE). MATERIALS AND METHODS: OLE hydroalcoholic extracts were obtained from N. oleander leaves and analyzed by HPLC. In vivo experiments with guinea pigs consisted if oral administration of water, 150mg/kg and 300mg/kg OLE extract. Clinical signs and ECG analysis were evaluated. Sample tissues from the heart were processed for histopathological and ultra-structural analysis. Autonomic effects were assessed through pharmacological blockade and ECG monitoring. In vitro experiments were conducted with isolated ventricular myocytes from adult mice. The effects of OLE extract on cardiac excitability, Na+/K+ pump current and global Ca2+ transients were evaluated. RESULTS: Our results demonstrated that OLE hydroalcoholic extract elicited severe cardiac arrhythmias that can lead to death with minimal tissue damage. In vitro experiments suggest that OLE causes electromechanical disturbances in the heart due to inhibition of Na+/K+ pump, mitochondrial swelling, and modulation of the sarco(endo)plasmic Ca2+ ATPase without interfering with the autonomic nervous system. Thus, arrhythmias and electrical conduction disturbances promoted by OLE are mainly associated with impaired cardiomyocyte dysfunction, rather than anatomical tissue remodeling and/or autonomic modulation. CONCLUSION: Our data revealed the potential cardiotoxicity and positive inotropic effect of OLE and its important role in modulation of electrophysiology in cardiomyocytes.

New insights into the elucidation of angiotensin-(1-7) in vivo antiarrhythmic effects and its related cellular mechanisms.

NEW FINDINGS: What is the central question of this study? Recently, there have been many studies exploring the biological effects of angiotensin-(1-7), which has been proved to have cardioprotective actions. However, the effects of this peptide on cardiac arrhythmias in vivo and details regarding its mechanism of action are still undetermined. What is the main finding and its importance? We investigated protective effects of angiotensin-(1-7) on cardiac arrhythmias in vivo, which were not properly explored in terms of cellular mechanisms. To verify effects of angiotensin-(1-7), we used different but complementary experimental approaches. Our data provide new evidence on the cellular mechanism and an in vivo demonstration of the acute antiarrhythmic effect of angiotensin-(1-7). Angiotensin-(1-7) [Ang-(1-7)] has been proved to have cardioprotective effects. However, the effects of this peptide on cardiac arrhythmias in vivo and details regarding its mechanism of action are still undetermined. The aim of this study was to investigate the protective effects of Ang-(1-7) against cardiac arrhythmias, its in vivo effects and cellular mechanism of action. We analysed the ECG upon inducement of arrhythmias in vivo in rats using a combination of halothane and adrenaline. To analyse the effects of Ang-(1-7) on cells, fresh mouse ventricular cardiomyocytes were isolated. The cardiomyocytes were superfused with a solution containing halothane and isoprenaline as a model to induce arrhythmias and used in three different approaches, namely a contractility assay, patch-clamp technique and confocal microscopy. The in vivo ECG showed that the injection of Ang-(1-7) (4 nm i.v.) significantly reduced cardiac arrhythmias [before, 49 ± 43 arrhythmic events versus after Ang-(1-7), 16 ± 14 arrhythmic events]. This effect was blocked by injection of A-779 and l-NAME, without changes in haemodynamic parameters. In addition, contractility experiments showed that Ang-(1-7) significantly decreased the number of arrhythmic events without changing the fractional shortening. This protection was associated with a reduction of the action potential repolarization and membrane hyperpolarization. Moreover, Ang-(1-7) decreased the number of calcium waves without any changes in the amplitude of the calcium transient, despite a significant reduction in the decay rate. Our data provide new evidence on the cellular mechanism together with an in vivo demonstration of the antiarrhythmic effects of Ang-(1-7).

Carvacrol modulates voltage-gated sodium channels kinetics in dorsal root ganglia.

Recent studies have shown that many of plant-derived compounds interact with specific ion channels and thereby modulate many sensing mechanisms, such as nociception. The monoterpenoid carvacrol (5-isopropyl-2-methylphenol) has an anti-nociceptive effect related to a reduction in neuronal excitability and voltage-gated Na(+) channels (NaV) inhibition in peripheral neurons. However, the detailed mechanisms of carvacrol-induced inhibition of neuronal NaV remain elusive. This study explores the interaction between carvacrol and NaV in isolated dorsal root ganglia neurons. Carvacrol reduced the total voltage-gated Na(+) current and tetrodotoxin-resistant (TTX-R) Na(+) current component in a concentration-dependent manner. Carvacrol accelerates current inactivation and induced a negative-shift in voltage-dependence of steady-state fast inactivation in total and TTX-R Na(+) current. Furthermore, carvacrol slowed the recovery from inactivation. Carvacrol provoked a leftward shift in both the voltage-dependence of steady-state inactivation and activation of the TTX-R Na(+) current component. In addition, carvacrol-induced inhibition of TTX-R Na(+) current was enhanced by an increase in stimulation frequency and when neurons were pre-conditioned with long depolarization pulse (5s at -50 mV). Taken all results together, we herein demonstrated that carvacrol affects NaV gating properties. The present findings would help to explain the mechanisms underlying the analgesic activity of carvacrol.

Investigation of terpinen-4-ol effects on vascular smooth muscle relaxation.

AIMS: This study investigated the mechanisms underlying the vascular effects of terpinen-4-ol in isolated rat aortic ring preparations. MAIN METHODS: The thoracic aortae of healthy rats were submitted to isometric tension recording. Membrane resting potential and input membrane resistance were measured by conventional microelectrode technique. KEY FINDINGS: Terpinen-4-ol reversibly relaxed endothelium-containing preparations pre-contracted with high K(+) and phenylephrine with IC50 values of 421.43 µM and 802.50 µM, respectively. These effects were significantly reduced by vascular endothelium removal. In Ca(2+)-free and high K(+) (80 mM) medium, the contractions produced by Ba(2+) were reduced by terpinen-4-ol (100-1000 µM) in a concentration-dependent manner. In aortic rings maintained under Ca(2+)-free conditions, terpinen-4-ol significantly reduced the contractions induced by either phenylephrine (1 µM) or phorbol 12,13-dibutyrate (1 µM). Terpinen-4-ol (10-1000 µM) also relaxed the contractions evoked by BAYK-8644 (3 µM) with an IC50 of 454.23 µM. Neither membrane resting potential nor input resistance of smooth muscle cells was altered by terpinen-4-ol exposure. SIGNIFICANCE: The present results suggest that terpinen-4-ol induced vascular smooth muscle relaxation that was preferentially due to the inhibition of electromechanical pathways related to calcium influx through voltage-operated calcium channels.

n5-STZ Diabetic Model Develops Alterations in Sciatic Nerve and Dorsal Root Ganglia Neurons of Wistar Rats.

One experimental model of diabetes mellitus (DM) similar to type II DM, called n5-STZ, is obtained by a single injection (via i.p.) of streptozotocin (STZ) in the 5th day of life of newborn rats. The present investigation aimed to characterize alterations in excitability of rat peripheral neurons in n5-STZ model. n5-STZ DM was induced, and electrophysiological evaluation was done at 12th week of rat life. Rats developed glucose intolerance, sensory alteration, and hyperglycemia or near-normoglycemia (21.2 ± 1.6 and 7.4 ± 0.4 mmol/L). In near-normoglycemia group the significant electrophysiological alteration observed was decreased in amplitude of 2nd wave (2nd component, conduction velocity: 48.8 m/s) of compound action potential (CAP) of sciatic nerve. For hyperglycemic rats, decreased excitability, amplitude, and conduction velocity of 2nd CAP component of sciatic nerve were found; a depolarization of resting potential (4-5 mV) and reduction in maximum ascendant and descendant inclinations of action potential were found in DRG neurons but no alteration on Na(+) current (INa(+) ). Thus, n5-STZ rats develop alterations in excitability which were related to glycemic levels but were not likely attributable to changes on INa(+) . Our data confirm that n5-STZ model is a useful model to study type II DM.

Carvacrol decreases neuronal excitability by inhibition of voltage-gated sodium channels.

The monoterpenoid carvacrol (1) is present in many essential oils of plants and has attracted attention because of its beneficial biological activities, especially analgesic activity. However, the mechanism of action of 1 remains unknown. The present study aimed to explore the mechanisms whereby 1 produces its effects on the peripheral nervous system. Carvacrol reversibly blocked the excitability of the rat sciatic nerve in a concentration-dependent manner with an IC(50) value of 0.50 ± 0.04 mM. At 0.6 mM, 1 increased the rheobase from 3.30 ± 0.06 V to 4.16 ± 0.14 V and the chronaxy from 59.6 ± 1.22 µs to 75.0 ± 1.82 µs. Also, 1 blocked the generation of action potentials (IC(50) 0.36 ± 0.14 mM) of the intact dorsal root ganglion (DRG) neurons without altering the resting potential and input resistance. Carvacrol reduced the voltage-gated sodium current of dissociated DRG neurons (IC(50) 0.37 ± 0.05 mM). In this study it has been demonstrated that 1 blocks neuronal excitability by a direct inhibition of the voltage-gated sodium current, which suggests that this compound acts as a local anesthetic. The present findings add valuable information to help understand the mechanisms implicated in the analgesic activity of carvacrol.

Linalool blocks excitability in peripheral nerves and voltage-dependent Na+ current in dissociated dorsal root ganglia neurons.

Linalool is a terpene that occurs as a major constituent of essential oils of many plants of widespread distribution. It possesses several biological and pharmacological activities, including depressant effects on the central nervous system and olfactory receptors. The present study investigated whether linalool affects the excitability of peripheral components of the somatic sensory system. We used sciatic nerve and preparations of intact and dissociated neurons of dorsal root ganglion for extracellular, intracellular and patch-clamp recordings. Linalool concentration-dependently (0.3-2.0mM) and reversibly blocked the excitability of the sciatic nerve. It inhibited peak-to-peak amplitude of the compound action potential (IC(50) was 0.78+/-0.04 mM). At 0.8mM, it reversibly increased rheobase and chronaxy (from 3.2+/-0.1 V and 52.4+/-4.1 micros to 4.2+/-0.3 V and 71.2+/-5.5 micros (n=5), respectively) and inhibited with greater pharmacological potency the amplitude of the compound action potential components corresponding to axons with slower velocity of conduction. In a similar concentration range (0.1-6mM), linalool concentration-dependently and reversibly blocked the generation of action potentials of intact dorsal root ganglion neurons without alteration of resting membrane potential and input resistance, and inhibited the voltage-gated Na(+) current of dissociated dorsal root ganglion neurons. In conclusion, we demonstrated that linalool acts on the somatic sensory system with local anesthetic properties, since it blocked the action potential by acting on voltage-dependent Na(+) channels. This finding is important in showing the potential usefulness of linalool as a pharmacotherapeutic agent.