A Tale of Toxin Promiscuity: The Versatile Pharmacological Effects of Hcr 1b-2 Sea Anemone Peptide on Voltage-Gated Ion Channels.

Sea anemones are a rich source of biologically active compounds. Among approximately 1100 species described so far, Heteractis crispa species, also known as sebae anemone, is native to the Indo-Pacific area. As part of its venom components, the Hcr 1b-2 peptide was first described as an ASIC1a and ASIC3 inhibitor. Using Xenopus laevis oocytes and the two-electrode voltage-clamp technique, in the present work we describe the remarkable lack of selectivity of this toxin. Besides the acid-sensing ion channels previously described, we identified 26 new targets of this peptide, comprising 14 voltage-gated potassium channels, 9 voltage-gated sodium channels, and 3 voltage-gated calcium channels. Among them, Hcr 1b-2 is the first sea anemone peptide described to interact with isoforms from the Kv7 family and T-type Cav channels. Taken together, the diversity of Hcr 1b-2 targets turns this toxin into an interesting tool to study different types of ion channels, as well as a prototype to develop new and more specific ion channel ligands.

Protective role of activating PPARγ in advanced glycation end products-induced impairment of coronary artery vasodilation via inhibiting p38 phosphorylation and reactive oxygen species production.

Advanced glycation end products (AGEs) can damage voltage-gated K+ (Kv) channels and attenuate coronary artery vasodilation, but the underlying mechanisms remain unclear. The aim of this study was to investigate the role and potential mechanism of PPARγ in AGEs-induced Kv 1 channels impairment. We used both primary rat coronary smooth muscle cells (CSMCs) in vitro and Zucker Diabetic Fatty (ZDF) rat model in vivo. Overexpression of the Pparg gene by lentivirus vector (LV-Pparg) was used to transfect CSMCs for upregulation PPARγ. Kv 1.2 and Kv 1.5 currents were measured by patch clamp. The vascular tone of coronary artery was evaluated by isometric force measurements. The proteins expression of Kv1.2 and Kv1.5 channel were detected by western blot. PPARγ was detected by immunofluorescence and western blot. Oxidative stress markers including superoxide dismutase (SOD), glutathione peroxidase (GPx) and malondialdehyde (MDA) were detected by enzyme linked immunosorbent assay (ELISA). The phosphorylation of p38 mitogen-activated protein kinase (MAPK) and total p38 expression were detected by western blot. The intracellular ROS levels were measured by the fluorescent dye 2',7'- dichlorofluorescein diacetate (DCFDA) and a cellular ROS assay kit. We found that activating PPARγ via LV-Pparg (100 MOI, 5 × 108 TU/mL) prevented AGEs (100 µg/mL) -mediated impairment of Kv 1.2 and Kv 1.5 channels activity and improved the reduction of Kv 1.2 and Kv 1.5 protein expression in CSMCs. Isometric force measurements showed that activating PPARγ by pioglitazone (10 mg/kg/d, intragastric administration) improved the impairment of coronary artery vasodilation, and western blot analysis showed that activating PPARγ increased the Kv 1.2 and Kv 1.5 protein expression, while inhibiting PPARγ by GW9662 (10 mg/kg/d, intraperitoneal injection) attenuated these effects in ZDF rats. Furthermore, LV-Pparg overexpression PPARγ attenuated NADPH oxidase activity, which was shown as the reduction of the NOX2 and p22phox expression by western blot analysis, decreased the MDA production and increased the SOD and GPx activities by ELISA, finally led to reduce AGEs-mediated ROS production. Moreover, activating PPARγ by LV-Pparg inhibited AGEs-induced phosphorylation of p38 MAPK, by which could reduce AGEs-mediated NOX2, p22phox expression and ROS production, while CSMCs treatment with SB203580 (10 µmol/L), a p38 MAPK inhibitor, attenuated these effects. Activating PPARγ plays a protective role in AGEs-induced impairment of coronary artery vasodilation by inhibiting p38 phosphorylation to attenuate NOX2 and p22phox expression and further decrease oxidative stress induced by ROS overproduction.

Ventricular voltage-gated ion channels: Detection, characteristics, mechanisms, and drug safety evaluation.

Cardiac voltage-gated ion channels (VGICs) play critical roles in mediating cardiac electrophysiological signals, such as action potentials, to maintain normal heart excitability and contraction. Inherited or acquired alterations in the structure, expression, or function of VGICs, as well as VGIC-related side effects of pharmaceutical drug delivery can result in abnormal cellular electrophysiological processes that induce life-threatening cardiac arrhythmias or even sudden cardiac death. Hence, to reduce possible heart-related risks, VGICs must be acknowledged as important targets in drug discovery and safety studies related to cardiac disease. In this review, we first summarize the development and application of electrophysiological techniques that are employed in cardiac VGIC studies alone or in combination with other techniques such as cryoelectron microscopy, optical imaging and optogenetics. Subsequently, we describe the characteristics, structure, mechanisms, and functions of various well-studied VGICs in ventricular myocytes and analyze their roles in and contributions to both physiological cardiac excitability and inherited cardiac diseases. Finally, we address the implications of the structure and function of ventricular VGICs for drug safety evaluation. In summary, multidisciplinary studies on VGICs help researchers discover potential targets of VGICs and novel VGICs in heart, enrich their knowledge of the properties and functions, determine the operation mechanisms of pathological VGICs, and introduce groundbreaking trends in drug therapy strategies, and drug safety evaluation.

Arecoline hydrobromide enhances jejunum smooth muscle contractility via voltage-dependent potassium channels in W/Wv mice.

Gastrointestinal motility was disturbed in W/Wv, which were lacking of interstitial cells of Cajal (ICC). In this study, we have investigated the role of arecoline hydrobromide (AH) on smooth muscle motility in the jejunum of W/Wv and wild-type (WT) mice. The jejunum tension was recorded by an isometric force transducer. Intracellular recording was used to identify whether AH affects slow wave and resting membrane potential (RMP) in vitro. The whole-cell patch clamp technique was used to explore the effects of AH on voltage-dependent potassium channels for jejunum smooth muscle cells. AH enhanced W/Wv and WT jejunum contractility in a dose-dependent manner. Atropine and nicardipine completely blocked the excitatory effect of AH in both W/Wv and WT. TEA did not reduce the effect of AH in WT, but was sufficient to block the excitatory effect of AH in W/Wv. AH significantly depolarized the RMP of jejunum cells in W/Wv and WT. After pretreatment with TEA, the RMP of jejunum cells indicated depolarization in W/Wv and WT, but subsequently perfused AH had no additional effect on RMP. AH inhibited the voltage-dependent K+ currents of acutely isolated mouse jejunum smooth muscle cells. Our study demonstrate that AH enhances the contraction activity of jejunum smooth muscle, an effect which is mediated by voltage-dependent potassium channels that acts to enhance the excitability of jejunum smooth muscle cells in mice.

Effects of antipsychotic drugs and potassium channel modulators on spectral properties of local field potentials in mouse hippocampus and pre-frontal cortex.

Local field potentials (LFPs) recorded intracranially display a range of location-specific oscillatory spectra which have been related to cognitive processes. Although the mechanisms producing LFPs are not completely understood, it is likely that voltage-gated ion channels which produce action potentials and patterned discharges play a significant role. It is also known that antipsychotic drugs (APDs) affect LFP spectra and a direct inhibitory effect on voltage-gated potassium channels has been reported. Additionally, voltage-gated potassium channels have been implicated in the pathophysiology of schizophrenia, a disorder for which APDs are primary therapies. In this study we sought to: i) better characterise the effects of two APDs on LFPs spectra and connectivity measures and ii) examine the effects of potassium channel modulators on LFPs and potential overlap of effects with APDs. Intracranial electrodes were implanted in hippocampus (HIP) and pre-frontal cortex (PFC) of C57BL/6J mice; power spectra, coherence and phase-amplitude cross-frequency coupling were measured. Drugs tested were APDs haloperidol and clozapine as well as voltage-gated potassium channel modulators (KVMs) 4-aminopyridine (4-AP), tetraethylammonium, retigabine and E-4031. Both APDs and KVMs significantly reduced gamma power except 4-AP, which conversely increased gamma power. Clozapine and retigabine additionally reduced gamma coherence between HIP and PFC, while 4-AP demonstrated the opposite effect. Phase-amplitude coupling between theta and gamma oscillations in HIP was significantly reduced by the administration of haloperidol and retigabine. These results provide previously undescribed effects of APDs on LFP properties and demonstrate novel modulation of LFP characteristics by KVMs that intriguingly overlap with the APD effects.

Antisense Oligonucleotide Therapy Targeted Against ATXN3 Improves Potassium Channel-Mediated Purkinje Neuron Dysfunction in Spinocerebellar Ataxia Type 3.

Spinocerebellar ataxia type 3 (SCA3) is the second-most common CAG repeat disease, caused by a glutamine-encoding expansion in the ATXN3 protein. SCA3 is characterized by spinocerebellar degeneration leading to progressive motor incoordination and early death. Previous studies suggest that potassium channel dysfunction underlies early abnormalities in cerebellar cortical Purkinje neuron firing in SCA3. However, cerebellar cortical degeneration is often modest both in the human disease and mouse models of SCA3, raising uncertainty about the role of cerebellar dysfunction in SCA3. Here, we address this question by investigating Purkinje neuron excitability in SCA3. In early-stage SCA3 mice, we confirm a previously identified increase in excitability of cerebellar Purkinje neurons and associate this excitability with reduced transcripts of two voltage-gated potassium (KV) channels, Kcna6 and Kcnc3, as well as motor impairment. Intracerebroventricular delivery of antisense oligonucleotides (ASO) to reduce mutant ATXN3 restores normal excitability to SCA3 Purkinje neurons and rescues transcript levels of Kcna6 and Kcnc3. Interestingly, while an even broader range of KV channel transcripts shows reduced levels in late-stage SCA3 mice, cerebellar Purkinje neuron physiology was not further altered despite continued worsening of motor impairment. These results suggest the progressive motor phenotype observed in SCA3 may not reflect ongoing changes in the cerebellar cortex but instead dysfunction of other neuronal structures within and beyond the cerebellum. Nevertheless, the early rescue of both KV channel expression and neuronal excitability by ASO treatment suggests that cerebellar cortical dysfunction contributes meaningfully to motor dysfunction in SCA3.

The effects of gastrin-releasing peptide on the voltage-gated channels in rat hippocampal neurons.

Gastrin-releasing peptide (GRP) has been implicated in several aspects of physiology and behavior including digestion, cancer, lung development, and memory process. Increasing evidence in rodents shows that GRP may contribute to hippocampal circuit function. Though the central role of GRP in the brain has been established, the cellular and molecular mechanisms of its actions have not been well defined. Thus in this study, we verified the expression of GRPR in the rat hippocampal CA1 region. Then we examined the mechanisms closely related to neuronal excitability, the effects of GRP on voltage-gated ion channels in CA1 neurons using patch-clamp. The results showed that GRP could decrease voltage-gated sodium currents mainly by affecting the kinetics of recovery from the inactivated state. However, GRP enhanced both kinds of voltage-gated potassium channels, the A-type channels were more sensitive to GRP than K-type channels. In conclusion, we found that GRP could alter the voltage-gated Na+ and K+ ion channel characteristics which might be the ionic mechanisms of the physiological function of GRP in the brain.

Anticancer potential of new 3-nitroaryl-6-(N-methyl)piperazin-1,2,4-triazolo[3,4-a]phthalazines targeting voltage-gated K+ channel: Copper-catalyzed one-pot synthesis from 4-chloro-1-phthalazinyl-arylhydrazones.

A series of six 3-aryl-6-(N-methylpiperazin)-1,2,4-triazolo[3,4-a]phthalazines were prepared through a facile and efficient one-pot copper-catalyzed procedure from 4-chloro-1-phthalazinyl-arylhydrazones with relatively good yields (62-83%). The one-pot copper-catalytic procedure consists of two simultaneous reactions: (i) a direct intramolecular dehydrogentaive cyclization between ylidenic carbon and adjacent pyrazine nitrogen to form 1,2,4-triazolo ring and, (ii) a direct N-amination on carbon-chlorine bond. Then, an in vitro anticancer evaluation was performed for the synthesized compounds against five selected human cancer cells (A549, MCF-7, SKBr3, PC-3 and HeLa). The nitro-derivatives were significantly more active against cancer strains than against the rest of tested compounds. Specifically, compound 8d was identified as the most promising anticancer agent with significant biological responses and low relative toxicities on human dermis fibroblast. The cytotoxic effect of compound 8d was more significant on PC3, MCF-7 and SKBr3 cancer cells with low-micromolar IC50 value ranging from 0.11 to 0.59 µM, superior to Adriamycin drug. Mechanistic experimental and theoretical studies demonstrated that compounds 8d act as a K+ channel inhibitor in cancer models. Further molecular docking studies suggest that the EGFR Tyrosine Kinase enzyme may be a potential target for the most active 3-aryl-6-(N-methylpiperazin)-1,2,4-triazolo[3,4-a]phthalazines.

Signaling pathways targeting mitochondrial potassium channels.

In this review, we describe key signaling pathways regulating potassium channels present in the inner mitochondrial membrane. The signaling cascades covered here include phosphorylation, redox reactions, modulation by calcium ions and nucleotides. The following types of potassium channels have been identified in the inner mitochondrial membrane of various tissues: ATP-sensitive, Ca2+-activated, voltage-gated and two-pore domain potassium channels. The direct roles of these channels involve regulation of mitochondrial respiration, membrane potential and synthesis of reactive oxygen species (ROS). Changes in channel activity lead to diverse pro-life and pro-death responses in different cell types. Hence, characterizing the signaling pathways regulating mitochondrial potassium channels will facilitate understanding the physiological role of these proteins. Additionally, we describe in this paper certain regulatory mechanisms, which are unique to mitochondrial potassium channels.

Polyunsaturated fatty acid analogues differentially affect cardiac NaV, CaV, and KV channels through unique mechanisms.

The cardiac ventricular action potential depends on several voltage-gated ion channels, including NaV, CaV, and KV channels. Mutations in these channels can cause Long QT Syndrome (LQTS) which increases the risk for ventricular fibrillation and sudden cardiac death. Polyunsaturated fatty acids (PUFAs) have emerged as potential therapeutics for LQTS because they are modulators of voltage-gated ion channels. Here we demonstrate that PUFA analogues vary in their selectivity for human voltage-gated ion channels involved in the ventricular action potential. The effects of specific PUFA analogues range from selective for a specific ion channel to broadly modulating cardiac ion channels from all three families (NaV, CaV, and KV). In addition, a PUFA analogue selective for the cardiac IKs channel (Kv7.1/KCNE1) is effective in shortening the cardiac action potential in human-induced pluripotent stem cell-derived cardiomyocytes. Our data suggest that PUFA analogues could potentially be developed as therapeutics for LQTS and cardiac arrhythmia.

Efficacy of retigabine in ameliorating the brain insult following sarin exposure in the rat.

BACKGROUND: Sarin is an irreversible organophosphate cholinesterase inhibitor. Following toxic signs, an extensive long-term brain damage is often reported. Thus, we evaluated the efficacy of a novel anticonvulsant drug retigabine, a modulator of neuronal voltage gated K+ channels, as a neuroprotective agent following sarin exposure. METHODS: Rats were exposed to 1 LD50 or 1.2 LD50 sarin and treated at onset of convulsions with retigabine (5 mg/kg, i.p.) alone or in combination with 5 mg/kg atropine and 7.5 mg/kg TMB-4 (TA) respectively. Brain biochemical and immunohistopathological analyses were processed 24 h and 1 week following 1 LD50 sarin exposure and at 4 weeks following exposure to 1.2 LD50 sarin. EEG activity in freely moving rats was also monitored by telemetry during the first week following exposure to 1.2 LD50 and behavior in the Open Field was evaluated 3 weeks post exposure. RESULTS: Treatment with retigabine following 1 LD50 sarin exposure or in combination with TA following 1.2 LD50 exposure significantly reduced mortality rate compared to the non-treated groups. In both experiments, the retigabine treatment significantly reduced gliosis, astrocytosis and brain damage as measured by translocator protein (TSPO). Following sarin exposure the combined treatment (retigabine+ TA) significantly minimized epileptiform seizure activity. Finally, in the Open Field behavioral test the non-treated sarin group showed an increased mobility which was reversed by the combined treatment. CONCLUSIONS: The M current modulator retigabine has been shown to be an effective adjunct therapy following OP induced convulsion, minimizing epileptiform seizure activity and attenuating the ensuing brain damage.

Protriptyline, a tricyclic antidepressant, inhibits voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells.

In this study, we explore the inhibitory effects of protriptyline, a tricyclic antidepressant drug, on voltage-dependent K+ (Kv) channels of rabbit coronary arterial smooth muscle cells using a whole-cell patch clamp technique. Protriptyline inhibited the vascular Kv current in a concentration-dependent manner, with an IC50 value of 5.05 ± 0.97 µM and a Hill coefficient of 0.73 ± 0.04. Protriptyline did not affect the steady-state activation kinetics. However, the drug shifted the steady-state inactivation curve to the left, suggesting that protriptyline inhibited the Kv channels by changing their voltage sensitivity. Application of 20 repetitive train pulses (1 or 2 Hz) progressively increased the protriptyline-induced inhibition of the Kv current, suggesting that protriptyline inhibited Kv channels in a use (state)-dependent manner. The extent of Kv current inhibition by protriptyline was similar during the first, second, and third step pulses. These results suggest that protriptyline-induced inhibition of the Kv current mainly occurs principally in the closed state. The increase in the inactivation recovery time constant in the presence of protriptyline also supported use (state)-dependent inhibition of Kv channels by the drug. In the presence of the Kv1.5 inhibitor, protriptyline did not induce further inhibition of the Kv channels. However, pretreatment with a Kv2.1 or Kv7 inhibitor induced further inhibition of Kv current to a similar extent to that observed with protriptyline alone. Thus, we conclude that protriptyline inhibits the vascular Kv channels in a concentration- and use-dependent manner by changing their gating properties. Furthermore, protriptyline-induced inhibition of Kv channels mainly involves the Kv1.5.

Trans-toxin ion-sensitivity of charybdotoxin-blocked potassium-channels reveals unbinding transitional states.

In silico and in vitro studies have made progress in understanding protein-protein complex formation; however, the molecular mechanisms for their dissociation are unclear. Protein-protein complexes, lasting from microseconds to years, often involve induced-fit, challenging computational or kinetic analysis. Charybdotoxin (CTX), a peptide from the Leiurus scorpion venom, blocks voltage-gated K+-channels in a unique example of binding/unbinding simplicity. CTX plugs the external mouth of K+-channels pore, stopping K+-ion conduction, without inducing conformational changes. Conflicting with a tight binding, we show that external permeant ions enhance CTX-dissociation, implying a path connecting the pore, in the toxin-bound channel, with the external solution. This sensitivity is explained if CTX wobbles between several bound conformations, producing transient events that restore the electrical and ionic trans-pore gradients. Wobbling may originate from a network of contacts in the interaction interface that are in dynamic stochastic equilibria. These partially-bound intermediates could lead to distinct, and potentially manipulable, dissociation pathways.

PACAP stimulates insulin secretion by PAC1 receptor and ion channels in ß-cells.

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) plays a crucial role in the endocrine system. The present study aimed to investigate the effect of PACAP38 on insulin secretion and the underlying mechanism in rat pancreatic ß-cells. The insulin secretion results showed that PACAP38 stimulated insulin secretion in a glucose- and dose-dependent manner. The insulinotropic effect was mediated by PAC1 receptor, but not by VPAC1 and VPAC2 receptors. Inhibition of adenylyl cyclase and protein kinase A suppressed PACAP38-augmented insulin secretion. Glucose-regulated insulin secretion is dependent on a series of electrophysiological activities. Current-clamp technology suggested that PACAP38 prolonged action potential duration. Voltage-clamp recordings revealed that PACAP38 blocked voltage-dependent potassium currents, and this effect was reversed by inhibition of PAC1 receptor, adenylyl cyclase, or protein kinase A. Activation of Ca2+ channels by PACAP38 was also observed, which could be antagonized by the PAC1 receptor antagonist. In addition, calcium-imaging analysis indicated that PACAP38 increased intracellular Ca2+ concentration, which was decreased by PAC1 receptor antagonist. These findings demonstrate that PACAP38 stimulates glucose-induced insulin secretion mainly by acting on PAC1 receptor, inhibiting voltage-dependent potassium channels, activating Ca2+ channels and increasing intracellular Ca2+ concentration. Further, PACAP blocks voltage-dependent potassium currents via the adenylyl cyclase/protein kinase A signaling pathway.

Luteolin-induced coronary arterial relaxation involves activation of the myocyte voltage-gated K+ channels and inward rectifier K+ channels.

AIMS: Luteolin has been shown to be beneficial to cardiovascular tissues and organs. We aimed to study its vasospasmolytic effects against various vasoconstrictors in the isolated rat coronary arteries (RCAs) and its electrophysiological effects on K+ currents via voltage-gated potassium (Kv) channels and inward rectifier potassium (Kir) channels in freshly isolated rat coronary arterial smooth muscle cells (RCASMCs). MAIN METHODS: The vascular tone of the endothelium-denuded RCAs was recorded by a wire myograph. Kv currents and Kir currents in RCASMCs were assessed using whole-cell patch clamp. KEY FINDINGS: Preincubation with luteolin depressed the contractions elicited by KCl, thromboxane A2 analog U46619, vasopressin, Kir blocker BaCl2, Kv blocker 4-aminopyridine and elevation of extracellular calcium ([Ca2+]o) in high K+ depolarizing solution. Instant application of luteolin produced concentration-dependent relaxations in the endothelium-denuded RCAs precontracted with KCl or U46619. Both 4-aminopyridine and BaCl2 attenuated luteolin-induced relaxation in U46619-precontracted RCAs, while neither nitric oxide synthetase inhibitor NG-nitro-L-arginine methyl ester nor cyclooxygenase inhibitor indomethacin affected the relaxation. Luteolin augmented both Kv currents and Kir currents in RCASMCs and the augmentations were antagonized by 4-aminopyridine and BaCl2, respectively. SIGNIFICANCE: The present results demonstrated that luteolin antagonizes various vasoconstrictors in RCAs and augments both Kv currents and Kir currents in RCASMCs, suggesting that the direct action of luteolin on Kv channels and Kir channels is contributory to its vasospasmolytic effect. These findings indicate that luteolin may be a promising food additive with the aim of preventing coronary arterial spasm.

Treating TB human neuroectodermal cell line with retinoic acid induces the appearance of neuron-like voltage-gated ionic currents.

TB is a cell line derived from the cerebrospinal fluid sample of a patient with primary leptomeningeal melanomatosis. Our previous immunological and ultrastructural analysis revealed that TB cells differentiate towards a neuronal phenotype when grown in vitro up to 7 days in presence of 10 µM all-trans retinoic acid (RA). Recently, we reported that TB cells are sensitive to the cytotoxic effects of ß-amyloid peptides, activating the cytosolic phospholipase A2. To date, it is not known if RA, in addition to inducing morphological changes, also causes functional modification in TB cells, by regulating voltage-gated ionic currents. To this purpose, we performed electrophysiological characterization of undifferentiated (TB) and differentiated (RA-TB) cells by means of whole-cell patch clamp recordings. Upon depolarizing stimuli, both groups displayed voltage-gated K+ outward currents of similar amplitude. By contrast, the low amplitude voltage-gated Na+ currents recorded in undifferentiated TB cells were largely up-regulated by RA exposure. This current was strongly reduced by TTX and lidocaine and completely abolished by removal of extracellular sodium. Furthermore, treatment with RA caused the appearance of a late-onset inward current carried by Ca2+ ions in a subpopulation of TB cells. This current was not affected by removal of extracellular Na+ and was completely blocked by Cd2+, a broad-spectrum blocker of Ca2+ currents. Altogether, our results indicate that RA-differentiation of TB cells induces functional changes by augmenting the amplitude of voltage-gated sodium current and by inducing, in a subpopulation of treated cells, the appearance of a voltage-gated calcium current.

Sulfonyl(thio)urea derivative induction of insulin secretion is mediated by potassium, calcium, and sodium channel signal transduction.

AIM: To investigate the mechanism of action of sulfonyl(thio)urea derivative (SD) on glycemia and on insulin secretion in pancreatic islets. METHODS: Wistar rats were divided into hyperglycemic control group, rats received 4 g/kg body weight glucose plus sitagliptin 10 mg/kg (p.o.); hyperglycemic plus SD 10 mg/kg (p.o.); hyperglycemic plus SD plus sitagliptin. Blood was collected before glucose overloading (zero time), and at 15, 30, 60, and 180 min after glucose, from the afore mentioned groups for glycemia and glucagon-like peptide 1 (GLP-1) measurements and intestinal disaccharidases activity. Pancreatic islets were isolated for the calcium influx and insulin secretion in in vitro studies. RESULTS: SD reduced glycemia and increased GLP-1 secretion, while inhibited sucrase and lactase activity. This SD (1.0 and 10.0 µM) stimulated calcium influx in a similar percentile to that of glibenclamide, and in a nonsynergic manner. In addition, the trigger effect of SD on calcium influx was through the K+ -ATP-dependent channels, and partially by activating voltage-dependent K + channels and voltage-dependent calcium channels. Furthermore, SD-stimulated Na + and Ca 2+ entry, induced by the transient receptor potential ankyrin 1 and by modulation of Na + /Ca 2+ exchange. The activation of these pathways by SD culminated in in vitro insulin secretion, reinforcing the critical role of K + -ATP channels in the secretagogue effect of SD. CONCLUSIONS: SD diminish glycemia by inducing GLP-1 secretion and inhibiting disaccharidases. To our knowledge, this is the first report of an insulin secretagogue effect of SD that is mediated by potassium and calcium, as well as sodium, signal transduction.

Effect of Titanium Particles on the Voltage-Gated Potassium Channel Currents in Trigeminal Root Ganglion Neurons.

PURPOSE: Titanium (Ti) is the key material used in dental implants because of its excellent biocompatibility. But wear and corrosion Ti particles had been widely reported to induce inflammation and promote bone absorption. However, little information is known about the damage of Ti particles on neurons. MATERIALS AND METHODS: Trigeminal root ganglion (TRG) neurons were exposed to Ti particles (<5 µm). The electrophysiological properties of 2 main subtypes of voltage-gated potassium channels (VGPCs) (KA and KV) were examined by whole-cell patch-clamp techniques. RESULT: With the presence of 0.25 mg/mL Ti particles, amplitudes of IK, A and IK, V were both obviously inhibited. For IK, A, the activation V1/2 shifted to the depolarizing direction with an increased k value, whereas the inactivation V1/2 showed obvious hyperdepolarizing shifts. For IK, V, 0.5 mg/mL Ti particles produced a depolarizing shift of activation V1/2 with a slower activation rate. No significant changes of its inactivation kinetics were found. CONCLUSION: Titanium (Ti) particles might alter the electrophysiological properties of VGPCs on TRG neurons, which are likely to further influence the excitability of neurons.

A SGLT2 inhibitor dapagliflozin suppresses prolonged ventricular-repolarization through augmentation of mitochondrial function in insulin-resistant metabolic syndrome rats.

BACKGROUND: Metabolic syndrome (MetS) is a prevalent risk factor for cardiac dysfunction. Although SGLT2-inhibitors have important cardioprotective effects in hyperglycemia, their underlying mechanisms are complex and not completely understood. Therefore, we examined mechanisms of a SGLT2-inhibitor dapagliflozin (DAPA)-related cardioprotection in overweight insulin-resistant MetS-rats comparison with insulin (INSU), behind its glucose-lowering effect. METHODS: A 28-week high-carbohydrate diet-induced MetS-rats received DAPA (5 mg/kg), INSU (0.15 mg/kg) or vehicle for 2 weeks. To validate MetS-induction, we monitored all animals weekly by measuring body weight, blood glucose and HOMO-IR index, electrocardiograms, heart rate, systolic and diastolic pressures. RESULTS: DAPA-treatment of MetS-rats significantly augmented the increased blood pressure, prolonged Q-R interval, and low heart rate with depressed left ventricular function and relaxation of the aorta. Prolonged-action potentials were preserved with DAPA-treatment, more prominently than INSU-treatment, at most, through the augmentation in depressed voltage-gated K+-channel currents. DAPA, more prominently than INSU-treatment, preserved the depolarized mitochondrial membrane potential, and altered mitochondrial protein levels such as Mfn-1, Mfn-2, and Fis-1 as well as provided significant augmentation in cytosolic Ca2+-homeostasis. Furthermore, DAPA also induced significant augmentation in voltage-gated Na+-currents and intracellular pH, and the cellular levels of increased oxidative stress, protein-thiol oxidation and ADP/ATP ratio in cardiomyocytes from MetS rats. Moreover, DAPA-treatment normalized the increases in the mRNA level of SGLT2 in MetS-rat heart. CONCLUSIONS: Overall, our data provided a new insight into DAPA-associated cardioprotection in MetS rats, including suppression of prolonged ventricular-repolarization through augmentation of mitochondrial function and oxidative stress followed by improvement of fusion-fission proteins, out of its glucose-lowering effect.