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The correlation between long-QT and connexin 43 (Cx43) status and localization in elderly rats was determined to demonstrate a correlation between insulin resistance (I-R), ischemia-reperfusion, aging, and heart dysfunction. Male Wistar rats are grouped as 24-month-old rats (Aged-group), those with metabolic syndrome (8 months old; MetS-group), or controls (8 months old; Con-group). Both experimental groups have long-QT and low heart rate. Immunohistochemical imaging and quantification showed marked decreases in Cx43 staining of intercalated disc with less localizations in the Aged-group and MetS-group. The lateralization of Cx43 on longitudinal cell membrane was significantly high in the MetS-group than in the Con-group with no significant change in the Aged-group. Its significant cytoplasmic internalization was higher in the Aged-group than in the MetS-group. There were marked decreases in phospho-Cx43 (pCx43) staining of intercalated disc with less localizations in both groups than in the Con-group. Furthermore, lateralization of pCx43 was significantly low in the Aged-group and MetS-group, whereas there were no significant changes in the cytoplasmic internalization of both groups compared with the Con-group. Furthermore, the ratio of pCx43 to Cx43 was significantly small in both groups. We determined increases in RhoA and endothelin-1 in both groups, further supporting decreases in pCx43. Our data indicate the important role of I-R on long-QT in aging heart through alterations in both Cx43 protein level and localizations, leading to an abnormal spreading of ventricular repolarization in I-R heart

Metabolic syndrome (MetS) is associated with additional cardiovascular risk in mammalians while there are relationships between hyperglycemia-associated cardiovascular dysfunction and increased platelet P2Y12 receptor activation. Although P2Y12 receptor antagonist ticagrelor (Tica) plays roles in reduction of cardiovascular events, its beneficial mechanism remains poorly understood. Therefore, we aimed to clarify whether Tica can exert a direct protective effect in ventricular cardiomyocytes from high-carbohydrate diet-induced MetS rats, at least, through affecting sarcoplasmic reticulum (SR)-mitochondria (Mit) miscommunication. Tica treatment of MetS rats (150 mg/kg/day for 15 days) significantly reversed the altered parameters of action potentials by reversing sarcolemmal ionic currents carried by voltage-dependent Na+ and K+ channels, and Na+/Ca2+-exchanger in the cells, expressed P2Y12 receptors. The increased basal-cytosolic Ca2+ level and depressed SR Ca2+ load were also reversed in Tica-treated cells, at most, though recoveries in the phosphorylation levels of ryanodine receptors and phospholamban. Moreover, there were marked recoveries in Mit structure and function (including increases in both autophagosomes and fragmentations) together with recoveries in Mit proteins and the factors associated with Ca2+ transfer between SR-Mit. There were further significant recoveries in markers of both ER stress and oxidative stress. Taken into consideration the Tica-induced prevention of ER stress and mitochondrial dysfunction, our data provided an important document on the pleiotropic effects of Tica in the electrical activity of the cardiomyocytes from MetS rats. This protective effect seems through recoveries in SR-Mit miscommunication besides modulation of different sarcolemmal ion-channel activities, independent of P2Y12 receptor antagonism.

Purpose Metabolic syndrome (MetS) became a tremendous public health burden in the last decades. Store-operated calcium entry (SOCE) is a unique mechanism that causes a calcium influx, which is triggered by calcium store depletion. MetS-induced alterations in cardiac calcium signaling, especially in SOCE are still unclear. Therefore, we aim to examine the possible role of SOCE and its components (STIM1 and Orai1) in the MetS-induced cardiac remodeling. Methods We used male, adult (12 weeks) Wistar albino rats (n = 20). Animals were randomly divided into two groups which were: control (C) and MetS. We gave 33% sucrose solution to animals instead of water for 24 weeks to establish MetS model. In the end, papillary muscle function was evaluated, and various electrophysiological analyses were made in isolated cardiomyocytes. Additionally, STIM1 and Orai1 protein and mRNA expressions were analyzed. Results We observed a deterioration in contractility in MetS animals and demonstrated the contribution of SOCE by applying a SOCE inhibitor (BTP2). Calcium spark frequency was increased while its amplitude was decreasing in MetS hearts, which was reversed after SOCE inhibition. The amplitude of transient calcium changes in the MetS group was decreased, and it decreased further BTP2 application. Both protein and mRNA levels of STIM1 and Orai1 were increased significantly in MetS hearts. Conclusion Current data indicate the significant contribution of SOCE to cardiac calcium handling in the MetS model. We think MetS-induced SOCE activation is a compensation mechanism that is required for the continuum of proper cardiac functioning, although the activation can also cause cardiac hypertrophy.

Consumption of illicit pharmaceutical products containing sibutramine has been reported to cause cardiovascular toxicity problems. This study aimed to demonstrate the toxicity profile of sibutramine, and thereby provide important implications for the development of more effective strategies in both clinical approaches and drug design studies. Action potentials (APs) were determined from freshly isolated ventricular cardiomyocytes with whole-cell configuration of current clamp as online. The maximum amplitude of APs (MAPs), the resting membrane potential (RMP), and AP duration from the repolarization phases were calculated from original records. The voltage-dependent K+-channel currents (IK) were recorded in the presence of external Cd2+ and both inward and outward parts of the current were calculated, while their expression levels were determined with qPCR. The levels of intracellular free Ca2+ and H+ (pHi) as well as reactive oxygen species (ROS) were measured using either a ratiometric micro-spectrofluorometer or confocal microscope. The mechanical activity of isolated hearts was observed with Langendorff-perfusion system. Acute sibutramine applications (10–8–10−5 M) induced significant alterations in both MAPs and RMP as well as the repolarization phases of APs and IK in a concentration-dependent manner. Sibutramine (10 μM) induced Ca2+-release from the sarcoplasmic reticulum under either electrical or caffeine stimulation, whereas it depressed left ventricular developed pressure with a marked decrease in the end-diastolic pressure. pHi inhibition by sibutramine supports the observed negative alterations in contractility. Changes in mRNA levels of different IK subunits are consistent with the acute inhibition of the repolarizing IK, affecting AP parameters, and provoke the cardiotoxicity.

Aging in humans represents declining in cardio-protective systems, however its mechanisms are not known yet. We aimed to analyse how aging affects key mechanisms responsible for contractile dysfunction via comparing the improperly synchrony between electrical and mechanical activities in male aged-rats (24-month old) comparison to those of adult-rats (6-month old). We determined significantly increased systemic oxidative stress with decreased antioxidant capacity, clear insulin resistance and hypertrophy in aged-rats with normal fasting blood glucose. We also determined significantly high level of reactive oxygen species, ROS production in fluorescent dye chloromethyl-2′,7′-dichlorodihydrofluoroscein diacetate (DCFDA) loaded isolated cardiomyocytes from aged-rats, confirming the increased oxidative stress in these hearts. In situ electrocardiograms, ECGs presented significant prolongations in RR- and QT-intervals in the aged-rats. Invasive hemodynamic measurements demonstrated marked increases in the heart rate and mean arterial pressure and decreases in the ejection-fraction and preload-recruitable stroke-work, together with depressed contraction and relaxation activities in aortic rings. In light and electron microscopy examinations in aged-rats, significant increases in muscle fibre radius and amount of collagen fibres were detected in the heart as well as markedly flattened and partial local splitting in elastic lamellas in the aorta, besides irregularly clustered mitochondria and lysosomes around the myofilaments in cardiomyocytes. MitoTEMPO treatment of tissue samples and cardiomyocytes from aged-rats for 1-h induced significant structural improvements. In the second part of our study, we have shown that mitochondria-targeted antioxidant MitoTEMPO antagonized all alterations in the heart samples as well as penylephrine-induced contractile and acetylcholine-induced relaxation responses of aged-rat aortic rings. Overall, the present data strongly support the important role of mitochondrial oxidative stress in the development of aged-related insufficiencies and that antioxidant strategies specifically targeting this organelle could have therapeutic benefit in aging-associated complications.

Background Atrial fibrillation (AF) is a comorbidity associated with heart failure and catecholaminergic polymorphic ventricular tachycardia. Despite the Ca 2+ ‐dependent nature of both of these pathologies, AF often responds to Na + channel blockers. We investigated how targeting interdependent Na + /Ca 2+ dysregulation might prevent focal activity and control AF. Methods and Results We studied AF in 2 models of Ca 2+ ‐dependent disorders, a murine model of catecholaminergic polymorphic ventricular tachycardia and a canine model of chronic tachypacing‐induced heart failure. Imaging studies revealed close association of neuronal‐type Na + channels (nNa v ) with ryanodine receptors and Na + /Ca 2+ exchanger. Catecholamine stimulation induced cellular and in vivo atrial arrhythmias in wild‐type mice only during pharmacological augmentation of nNa v activity. In contrast, catecholamine stimulation alone was sufficient to elicit atrial arrhythmias in catecholaminergic polymorphic ventricular tachycardia mice and failing canine atria. Importantly, these were abolished by acute nNa v inhibition (tetrodotoxin or riluzole) implicating Na + /Ca 2+ dysregulation in AF. These findings were then tested in 2 nonrandomized retrospective cohorts: an amyotrophic lateral sclerosis clinic and an academic medical center. Riluzole‐treated patients adjusted for baseline characteristics evidenced significantly lower incidence of arrhythmias including new‐onset AF, supporting the preclinical results. Conclusions These data suggest that nNa V s mediate Na + ‐Ca 2+ crosstalk within nanodomains containing Ca 2+ release machinery and, thereby, contribute to AF triggers. Disruption of this mechanism by nNa v inhibition can effectively prevent AF arising from diverse causes.

The prevalence of death from cardiovascular disease is significantly higher in elderly populations; the underlying factors that contribute to the age‐associated decline in cardiac performance are poorly understood. Herein, we identify the involvement of sodium/glucose co‐transporter gene (SGLT2) in disrupted cellular Ca2+‐homeostasis, and mitochondrial dysfunction in age‐associated cardiac dysfunction. In contrast to younger rats (6‐month of age), older rats (24‐month of age) exhibited severe cardiac ultrastructural defects, including deformed, fragmented mitochondria with high electron densities. Cardiomyocytes isolated from aged rats demonstrated increased reactive oxygen species (ROS), loss of mitochondrial membrane potential and altered mitochondrial dynamics, compared with younger controls. Moreover, mitochondrial defects were accompanied by mitochondrial and cytosolic Ca2+ ([Ca2+]i) overload, indicative of disrupted cellular Ca2+‐homeostasis. Interestingly, increased [Ca2+]i coincided with decreased phosphorylation of phospholamban (PLB) and contractility. Aged‐cardiomyocytes also displayed high Na+/Ca2+‐exchanger (NCX) activity and blood glucose levels compared with young‐controls. Interestingly, the protein level of SGLT2 was dramatically increased in the aged cardiomyocytes. Moreover, SGLT2 inhibition was sufficient to restore age‐associated defects in [Ca2+]i‐homeostasis, PLB phosphorylation, NCX activity and mitochondrial Ca2+‐loading. Hence, the present data suggest that deregulated SGLT2 during ageing disrupts mitochondrial function and cardiac contractility through a mechanism that impinges upon [Ca2+]i‐homeostasis. Our studies support the notion that interventions that modulate SGLT2‐activity can provide benefits in maintaining [Ca2+]i and cardiac function with advanced age.

Aging is an important risk factor for cardiac dysfunction. Heart during aging exhibits a depressed mechanical activity, at least, through mitochondria-originated increases in ROS. Previously, we also have shown a close relationship between increased ROS and cellular intracellular free Zn2+ ([Zn2+]i) in cardiomyocytes under pathological conditions as well as the contribution of some re-expressed levels of Zn2+-transporters for redistribution of [Zn2+]i among suborganelles. Therefore, we first examined the cellular (total) [Zn2+] and then determined the protein expression levels of Zn2+-transporters in freshly isolated ventricular cardiomyocytes from 24-month rat heart compared to those of 6-month rats. The [Zn2+]i in the aged-cardiomyocytes was increased, at most, due to increased ZIP7 and ZnT8 with decreased levels of ZIP8 and ZnT7. To examine redistribution of the cellular [Zn2+]i among suborganelles, such as Sarco/endoplasmic reticulum, S(E)R, and mitochondria ([Zn2+]SER and [Zn2+]Mit), a cell model (with galactose) to mimic the aged-cell in rat ventricular cell line H9c2 was used and demonstrated that there were significant increases in [Zn2+]Mit with decreases in [Zn2+]SER. In addition, the re-distribution of these Zn2+-transporters were markedly changed in mitochondria (increases in ZnT7 and ZnT8 with no changes in ZIP7 and ZIP8) and S(E)R (increase in ZIP7 and decrease in ZnT7 with no changes in both ZIP8 and ZnT8) both of them isolated from freshly isolated ventricular cardiomyocytes from aged-rats. Furthermore, we demonstrated that cellular levels of ROS, both total and mitochondrial lysine acetylation (K-Acetylation), and protein-thiol oxidation were significantly high in aged-cardiomyocytes from 24-month old rats. Using a mitochondrial-targeting antioxidant, MitoTEMPO (1 &micro, M, 5-h incubation), we provided an important data associated with the role of mitochondrial-ROS production in the [Zn2+]i-dyshomeostasis of the ventricular cardiomyocytes from 24-month old rats. Overall, our present data, for the first time, demonstrated that a direct mitochondria-targeting antioxidant treatment can be a new therapeutic strategy during aging in the heart through a well-controlled [Zn2+] distribution among cytosol and suborganelles with altered expression levels of the Zn2+-transporters.

Sodium-glucose cotransporter 2 (SGLT2) inhibitors showed significant effects in patients with diabetes or metabolic syndrome (MetS) with high cardiovascular risk. Although the increased intracellular Zn

Role of β

Antiplatelet therapy has been widely used for management of patients with ischaemic heart diseases or thrombotic events. Experimental studies have shown that ticlopidine and clopidogrel decreased L-type Ca2+ currents (ICaL), altered action potential (AP) duration and thence exerted negative inotropic effects. In this study we tested if ticagrelor, a non-thienopyridine agent, has any influence on contractile and electrical properties of isolated ventricular myocytes. Cardiomyocytes were isolated from male rat hearts with an enzymatic dissociation procedure and left ventricular myocytes were used for experiments. The effects of ticagrelor (1 μM) on sarcomere shortening, ionic currents and action potentials were measured at 36 ± 1 °C. Ticagrelor significantly reduced ICaL density (~18 %, p

Due to the increasing use of wireless technology in developing countries, particularly mobile phones, the influence of electromagnetic fields (EMF) on biologic systems has become the subject of an intense debate. Therefore, in this study we investigated the effect of 2.1 GHz EMF on contractility and beta-adrenergic (β-AR) responsiveness of ventricular myocytes.Rats were randomized to the following groups: Sham rats (SHAM) and rats exposed to 2.1 GHz EMF for 2 h/day for 10 weeks (EM-10). Sarcomere shortening and Ca(2+) transients were recorded in isolated myocytes loaded with Fura2-AM and electrically stimulated at 1 Hz, while L-type Ca(2+) currents (I(CaL)) were measured using whole-cell patch clamping at 36 ± 1°C. Cardiac nitric oxide (NO) levels were measured in tissue samples using a colorimetric assay kit.Fractional shortening and amplitude of the matched Ca(2+) transients were not changed in EM-10 rats. Although the isoproterenol-induced (10(-6) M) I(CaL) response was reduced in rats exposed to EMF, basal I(CaL) density in myocytes was similar between the two groups (p0.01). Moreover, EMF exposure led to a significant increase in nitric oxide levels in rat heart (p0.02).Long-term exposure to 2.1 GHz EMF decreases β-AR responsiveness of ventricular myocytes through NO signaling.

Intracellular labile (free) Zn

Functional contribution of S(E)R-mitochondria coupling to normal cellular processes is crucial and any alteration in S(E)R-mitochondria axis may be responsible for the onset of diseases. Mitochondrial free Zn

Mechanical activity of the heart is adversely affected in metabolic syndrome (MetS) characterized by increased body mass and marked insulin resistance. Herein, we examined the effects of high carbohydrate intake on cardiac function abnormalities by evaluating in situ heart work, heart rate, and electrocardiograms (ECGs) in rats. MetS was induced in male Wistar rats by adding 32% sucrose to drinking water for 22–24 weeks and was confirmed by insulin resistance, increased body weight, increased blood glucose and serum insulin, and increased systolic and diastolic blood pressures in addition to significant loss of left ventricular integrity and increased connective tissue around myofibrils. Analysis of in situ ECG recordings showed a markedly shortened QT interval and decreased QRS amplitude with increased heart rate. We also observed increased oxidative stress and decreased antioxidant defense characterized by decreases in serum total thiol level and attenuated paraoxonase and arylesterase activities. Our data indicate that increased heart rate and a shortened QT interval concomitant with higher left ventricular developed pressure in response to β-adrenoreceptor stimulation as a result of less cyclic AMP release could be regarded as a natural compensation mechanism in overweight rats with MetS. In addition to the persistent insulin resistance and obesity associated with MetS, one should consider the decreased heart work, increased heart rate, and shortened QT interval associated with high carbohydrate intake, which may have more deleterious effects on the mammalian heart.

Clinical and experimental studies have shown an association between intracellular free Zn

Recent studies have implicated a relationship between RhoA/ROCK activity and defective Ca

The Zn

There is evidence that phenolic structure may have biological functions. Ellagic acid (EA), a phenolic compound, has been suggested to have cardioprotective effects. EA effects were investigated on cardiac Ca currents and contractility in rat ventricular myocytes to elucidate the underlying mechanisms. Freshly isolated ventricular myocytes from rat hearts were used. EA dose-dependently reduced Ca currents (ICaL) with EC50 = 23 nM, whereas it did not affect the inactivation and reactivation parameters. Inhibition of adenylate cyclase by SQ-22536 (10 μM) and probucol (5 μM) had no effect on EA modulation of ICaL. Nitric oxide synthase block by L-NAME (500 μM) and of guanylate cyclase by ODQ (1 μM) abolished EA inhibitory effects on ICaL. Moreover, EA blunted ventricular myocytes' fractional shortening in a concentration-dependent manner. In conclusion, EA affects ionic and mechanical properties of rat ventricular myocytes starting at nanomolar concentrations. EA suppresses ICaL and exerts negative inotropic effects through activation of NOS-GC-cGMP pathways. Thus, EA may be useful in pathophysiological conditions such as hypertension and ischemic heart diseases.

Recently, sodium tungstate was suggested to improve cardiac performance of diabetic rats in perfused hearts based on its insulinomimetic activity. In this study, we aimed to investigate the cellular and molecular mechanisms underlying this beneficial effect of sodium tungstate. Tungstate was administered (100 mg/kg/day) to diabetic and control rats intragastrically for 6 weeks. Blood glucose levels increased, whereas body weight, heart weight and plasma insulin levels decreased significantly in diabetic animals. Interestingly, none of these parameters was changed by tungstate treatment. On the other hand, fractional shortening and accompanying intracellular Ca(2+) [Ca(2+)](i) transients of isolated ventricular myocytes were measured, and sodium tungstate was found to improve the peak shortening and the amplitude of [Ca(2+)](i) transients in diabetic cardiomyocytes. Potassium and L-type Ca(2+) currents were also recorded in isolated ventricular cells. Significant restoration of suppressed I (to) and I (ss) was achieved by tungstate administration. Nevertheless, L-type calcium currents did not change either in untreated or treated diabetic rats. Tissue biochemical parameters including TBARS, protein carbonyl content, xanthine oxidase (XO) and xanthine dehydogenase (XDH) were also determined, and diabetes revealed a marked increase in TBARS and carbonyl content which were decreased significantly by tungstate treatment. Conversely, although XO and XDH activities didn't change in untreated diabetic rats, a remarkable but insignificant decrease was detected in treated animals. In conclusion, tungstate treatment improved diabetes-induced contractile abnormalities via restoration of dysregulated [Ca(2+)](i) and altered ionic currents. This beneficial effect is due to antioxidant property of sodium tungstate rather than normalization of hyperglycemia.