Search

Fat-rich diet (FRD) triggers health complications like hypertension, dyslipidemia, hyperglycemia, insulin resistance and non-alcoholic fatty liver disease, known as the risk factors of metabolic syndrome (MetS), which may result in neurological deficits. The impact of MetS on neuronal functions and brain morphology are poorly understood. We induced MetS-like conditions by exposing hypertriacylglycerolemic (HTG) rats to FRD for eight weeks with the aim to study possible neurological dysfunctions. HTG-FRD rats were compared to HTG rats and Wistar rats on standard diet. The physiological status of the animals was monitored by body, liver and kidney weight. Morphology of the liver, vessel wall and hippocampus were investigated. Basal neurotransmission and synaptic plasticity were measured in the hippocampus ex-vivo. A marked increase of liver weight with marks of steatosis was found in the HTG-FRD group. FRD induced an increase of aortic intima-media thickness. Extracellular recording revealed FRD-induced impairment of long-term potentiation (LTP) at Cornu Ammonis (CA)3-CA1 synapse, contrary to increased presynaptic fiber volley (pV). Reduced thickness of pyramidal cell layer at the CA1 area was found morphometrically. LTP was directly associated with kidney weight and inversely associated with liver weight, pV directly correlated with liver weight, liver/body wt ratio and aortic intima-media thickness. Our results suggest correlations between altered physiological status due to MetS-like conditions and neurological deficits, which may be related with consecutive development of so-called metabolic cognitive syndrome.

Intercellular connexin-43 (Cx43) channels are essential for electrical coupling and direct cardiac cell to cell communication to ensure heart function. Expression of Cx43 is altered due to stressful conditions and also affected by the alterations in extracellular matrix. We aimed to explore the effect of chest irradiation on myocardial expression of Cx43 and miR-1 which regulates GJA1 gene transcription for Cx43. Implication of miR-21 that regulates expression of extracellular matrix proteins and PKC signalling that may affect Cx43-mediated coupling was examined as well. Western blot and real-time PCR analyses revealed that six weeks after the exposure of healthy Wistar rats chest to single irradiation of 25 Gy significant myocardial alterations were observed: 1)/ increase of total Cx43 protein expression and its functional phosphorylated forms; 2) suppressed levels of miR-1; 3) enhanced expression of PKCε which phosphorylates Cx43; 4) increase of miR-21 levels; 5) increase of PKCδ expression. These results suggest that irradiation causes post-transcriptional regulation of myocardial Cx43 expression by miR-1 possibly through miR-21 and PKC signalling. We conclude that single dose of irradiation has the potential to enhance myocardial intercellular communication that might be beneficial for the heart that needs to be investigated in details in further studies.

The purpose of this study was to test our hypothesis that red palm oil (RPO) intake may affect abnormalities of myocardial connexin-43 (Cx43) and protein kinase Cε (PKCε) signaling, and consequently the propensity of the spontaneously hypertensive rat heart (SHR) heart to arrhythmias. SHR and Wistar–Kyoto (WKY) rats fed a standard rat chow plus red palm oil (200 µL/day) for 5 weeks were compared with untreated rats. Cytosolic but not particulate PKCε expression as well as Cx43-mRNA, total Cx43 proteins, and its phoshorylated forms were increased, and disordered localization of Cx43 was attenuated in the left ventricle of RPO-fed SHR compared with untreated rats. These alterations were associated with suppression of early post-ischemic-reperfusion-related ventricular tachycardia and electrically inducible ventricular fibrillation. However, the treatment dose of RPO caused down-regulation of myocardial Cx43, but did not alter its cell membrane distribution or overall PKCε expression in WKY rats. It was, however, associated with poor arrhythmia protection, suggesting overdosing. Results indicate that SHR benefit from RPO intake, particularly because of its apparent anti-arrhythmic effects. This protection can be, in part, attributed to the preservation of cell-to-cell communication via up-regulation of myocardial Cx43, but not with PKCε activation.

This study examined whether triiodo-L-thyronine (T3) affects the expression of the major intercellular channel protein, connexin-43, and contractile protein alpha-sarcomeric actin. Cultured cardiomyocytes from newborn rats were treated on day three in culture with 10 or 100 nM T3 and examined 48 and 72 h thereafter. Treated and untreated cells were examined by immunofluorescence and electron microscopy. Expression levels of Cx43 and sarcomeric alpha-actin were monitored by Western blot analysis. Immunofluorescence labeling showed cell membrane location of Cx43 in punctuate gap junctions, whereby fluorescence signal area was significantly higher in cultured cardiomyocytes exposed to T3. This correlated with electron microscopical findings showing increased numbers and size of gap junction profiles, as well as with a significant dose-dependent increase of Cx43 expression detected by Western blot. Immunofluorescence of sarcomeric a-actin was enhanced and its expression increased dose- and time-dependently in T3-treated cultured heart myocytes. However, exposure to the higher dosage (100 nM) of T3 caused mild disintegration of sarcomeric a-actin in some myocytes, suggesting an over-dosage. The results indicate that T3 up-regulates Cx43 and accelerates gap junction formation in cultured neonatal cardiomyocytes. They suggest that thyroid status cannot only modulate the mechanical function of cardiomyocytes but also cell-to-cell communication essential for myocardial electrical and metabolic synchronizations.

The aim of this study was to characterise the arrhythmogenic mechanisms involved in hypokalaemia-induced sustained ventricular fibrillation (SVF), in hypertensive rats. The hearts from rats with hypertension induced by the nitric oxide synthase inhibitor L-NAME, and age-matched normotensive controls, were perfused in Langendorff mode with oxygenated Krebs-Henseleit solution followed by a K + -deficient solution. In additional experiments, free intracellular Ca 2 + concentration ([Ca 2 + ] i ) was measured using fura-2 in conjunction with an epicardial optical probe. The epicardial electrocardiogram was continuously monitored during all experiments. The gap junction protein connexin-43 and the ultrastructure of the cardiomyocytes were examined, and selected enzyme activities were measured in situ. There was a higher incidence of low-K + -induced SVF in the hearts of hypertensive compared to normotensive rats (83 % vs. 33 %, P < 0.05). Perfusion with a low-K + -containing solution lead to elevation of diastolic [Ca 2 + ] i that was accompanied by premature beats, bigeminy, ventricular tachycardia and transient ventricular fibrillation. These events occurred earlier with increased incidence and duration in the hearts of hypertensive rats (arrhythmia scores: hypertensive, 4.9 ′ 0.7; normotensive, 3.1 ′ 0.1; P < 0.05), which exhibited apparent remodelling accompanied by a significant decrease in the density of connexin-43-positive gap junctions. Moreover, low-K + -related myocardial changes, including local impairment of intermyocyte junctions, ultrastructural alterations due to Ca 2 + overload and intercellular uncoupling, and decreased enzyme activities were more pronounced and more dispersed in hypertensive than normotensive rats. In conclusion, nitric oxide-deficient hypertension is associated with decreased myocardial coupling at gap junctions. The further localised deterioration of junctional coupling, due to low-K + -induced Ca 2 + disturbances, as well as spatial heterogeneity of myocardial alterations including interstitial fibrosis, probably provide the mechanisms for re-entry and sustaining ventricular fibrillation.

We hypothesized that the pineal hormone melatonin, which exhibits cardioprotective effects, might affect myocardial expression of cell-to-cell electrical coupling protein connexin-43 (Cx43) and protein kinase C (PKC) signaling, and hence, the propensity of the heart to lethal ventricular fibrillation (VF). Spontaneously hypertensive (SHR) and normotensive Wistar rats fed a standard rat chow received melatonin (40 μg/mL in drinking water during the night) for 5 weeks, and were compared with untreated rats. Melatonin significantly reduced blood pressure and normalized triglycerides in SHR, whereas it decreased body mass and adiposity in Wistar rats. Compared with healthy rats, the threshold to induce sustained VF was significantly lower in SHR (18.3 ± 2.6 compared with 29.2 ± 5 mA; p < 0.05) and increased in melatonin-treated SHR and Wistar rats to 33.0 ± 4 and 32.5 ± 4 mA. Melatonin attenuated abnormal myocardial Cx43 distribution in SHR, and upregulated Cx43 mRNA, total Cx43 protein, and its functional phosphorylated forms in SHR, and to a lesser extent, in Wistar rat hearts. Moreover, melatonin suppressed myocardial proapoptotic PKCδ expression and increased cardioprotective PKCε expression in both SHR and Wistar rats. Our findings indicate that melatonin protects against lethal arrhythmias at least in part via upregulation of myocardial Cx43 and modulation of PKC-related cardioprotective signaling.

Omega-3 fatty acids (omega-3 FA) and statins exhibit besides lipid-lowering effects the antiarrhythmic ability in clinic, while definite mechanisms are not yet elucidated. Our goal was to examine whether these compounds can modulate inducibility of hypertriglyceridemic (HTG) rat heart to ventricular fibrillation (VF) and myocardial cell-to-cell coupling protein connexin-43 (Cx43). HTG and healthy Wistar rats were orally treated with omega-3 FA(30 mg/100 g/day/2 mth) and atorvastatin (Ato, 0.5 mg/100 g/day/2 mth) and compared to untreated rats. Susceptibility of the heart to electrically-inducible VF and functional parameters were monitored using Langendorff-perfused isolated heart. Ventricular tissues from treated and untreated HTG and Wistar rat hearts were processed for ultrastructure examination as well as for analysis of myocardial Cx43 distribution and expression using antiCx43 MAB, immunofluorescence and immunoblotting. Both, omega-3 FA and atorvastatin reduced elevated blood pressure, triglycerides and heart rate in HTG rats. Compared to Wistar the threshold to induce VF was lower in HTG rat hearts, which exhibited abnormal Cx43 distribution, decreased immunostaining and elevated phosphorylated form of Cx43. In contrast, an enhancement of immunostaining of Cx43, suppression of hyperphosphorylation of Cx43 and improvement of cardiomyocyte and intercellular junction integrity by omega-3 FA and atorvastatin was associated with a significant increase of threshold for VF. Moreover, treatment resulted in up-regulation of myocardial Cx43 and increase of VF threshold in healthy rats that was associated with up-regulation of Cx43. Results indicate that antiarrhythmic effects of omega-3 FA and atorvastatin are linked with modulation of expression and/or phosphorylation of Cx43 and protection of cardiomyocyte and cell-to-cell junction integrity. As both compounds are ligands for PPAR, a possible regulation of Cx43 gene expression and pathways involved in Cx43 phosphorylation should be investigated.

Insufficient growth and rarefaction of capillaries, followed by endothelial dysfunction may represent one of the most critical mechanisms involved in heart damage. In this study we examined histochemical and ultrastructural changes in myocardial capillary endothelium in two models of heart failure streptozotocin-induced diabetes mellitus (STZ) and NO-deficient hypertension in male Wistar rats. Diabetes was induced by a single i.v. dose of STZ (45 mg/kg) and chronic 9-week stage was analysed. To induce NO-deficient hypertension, animals were treated with inhibitor of NO synthase L-nitroarginine methylester (L-NAME) (40 mg/kg) for 4 weeks. Left ventricular tissue was processed for enzyme catalytic histochemistry of capillary alkaline phosphatase (AlPh), dipeptidyl peptidase IV (DPP IV), and endothelial NO synthase/NADPH-diaphorase (NOS) and for ultrastructural analysis. In diabetic and hypertensive rats, lower/absent AlPh and DPP IV activities were found in focal micro-areas. NOS activity was significantly reduced and persisted only locally. Quantitative evaluation demonstrated reduction of reaction product intensity of AlPh, DPP and NOS by 49.50%, 74.36%, 20.05% in diabetic and 62.93%, 82.71%, 37.65% in hypertensive rats. Subcellular alterations of endothelial cells were found in heart of both groups suggesting injury of capillary function as well as compensatory processes. Endothelial injury was more significant in diabetic animals, in contrast the adaptation was more evident in hypertensive ones. CONCLUDING: both STZ-induced diabetes- and NO-deficient hypertension-related cardiomyopathy were accompanied by similar features of structural remodelling of cardiac capillary network manifested as angiogenesis and angiopathy. The latter was however, predominant and may accelerate disappearance of capillary endothelium contributing to myocardial dysfunction.

The aim of the present study was to investigate the effect of dietary supplementation with the pyridoindole antioxidant stobadine on kidney status and function in streptozotocin-induced diabetic rats. Diabetic male Wistar rats were fed a standard diet for 32 weeks or a diet supplemented with stobadine (0.05% w/w). The diabetic state was characterized by significantly elevated plasma levels of glucose, HbA1c and urea, severe reduction of total body weight and relatively enlarged kidneys. Elevated levels of conjugated dienes were recorded in the diabetic kidney confirming the presence of oxidative stress in diabetic animals. All diabetic rats showed marked proteinuria and albuminuria along with elevated excretion of the enzyme N-acetyl-beta-D-glucosaminidase. Long-term treatment of diabetic animals with stobadine significantly reduced total proteinuria, albuminuria and enzymuria, yet left the overall physical and glycemic status unaffected. It reduced oxidative damage of kidney tissue as shown by decreased conjugated diene level, and decreased matrix collagen cross-linking, as indicated by decreased breaking time values of rat tail tendons. These beneficial effects of stobadine, supported also by histological findings, may be brought about by virtue of the combination of its antioxidant potential with other effects, e.g., the postulated cholesterol-lowering ability or its ability to alter vascular reactivity and reduce the vascular tone.

Previous studies have indicated that isosmolar, but not hyperosmolar, ethanol induces in vitro gonadotropin-releasing hormone secretion from the basal hypothalamus, presumably by causing cell swelling. Moreover, ethanol reduces secretion of another hypothalamic neuropeptide vasopressin. We have studied the acute effect of ethanol on specific hypophysiotropic basal and K + -stimulated thyrotropin-releasing hormone secretion in vitro especially in relation to cell swelling. Isosmotic 40–160 mM ethanol increased thyrotropin-releasing hormone release from the hypothalamic paraventricular nucleus and median eminence in a dose-dependent manner. Both a 30% decrease of osmolarity and isosmotic 80 mM ethanol induced 12% swelling of hypothalamic neurons. Hyperosmotic 80 mM or 160 mM ethanol induced release of thyrotropin-releasing hormone from both hypothalamic structures but did not cause cell swelling (80 mM) or even induced cell shrinkage (160 mM). Depletion of medium Ca 2+ did not affect thyrotropin-releasing hormone secretion caused by either isosmotic or hyperosmotic ethanol. Our data indicate that both iso- and hyperosmotic ethanol stimulated release of hypophysiotropic thyrotropin-releasing hormone despite opposite effects on neuron volume. The mechanism of ethanol action appears complex and variable depending on the type of cell and neuropeptide affected.

Metabolism, monitored via in situ catalytic enzyme histochemistry and fine structure, was studied in the myocardium of chronic diabetic male Wistar rats administered L-arginine (12.8 mg/100 g/day) for 24 weeks. Diabetes was induced with a single i.v. injection of 55 mg/kg streptozotocin. After 6 months, the tissue of the left ventricle was processed for electron microscope examination and transmural tissue blocks were frozen for enzyme histochemistry. In diabetic myocardium, heterogeneous ischemia-like subcellular alterations of cardiomyocytes and capillaries were observed, together with interstitial fibrosis. This structural remodeling was accompanied by significantly decreased activity of endothelial nitric oxide synthase (NOS) and heterogeneously decreased activities of glycogen phosphorylase (GlPh), hydroxybutyrate dehydrogenase (HBDH) and adenosine triphophatases (ATPases) throughout the myocardium. In arginine-treated diabetic rats, there was evidence of protected structural integrity of endothelial cells and attenuated structural disturbances of cardiomyocytes. This was associated with the markedly preserved histochemical activities of all detected enzymes in comparison with nontreated diabetic rats (NOS 98.7 +/- 10.5% vs. 35.4 +/- 4.1%; ATPases 82.7 +/- 9.1% vs. 69.3 +/- 5.2%; GlPh 65.2 +/- 8.3% vs. 45.5 +/- 3.8%; HBDH 68.9 +/- 8.5% vs. 44.1 +/- 6.7% of control values). The results indicate that long-term supplementation of L-arginine may account for the reduction of diabetes-induced myocardial structural remodeling.

Hypertension-induced myocardial metabolic, structural and electrophysiological remodeling deteriorates with aging and contributes to both heart failure and occurrence of malignant arrhythmias. It has been shown in clinical trials that n-3 polyunsaturated fatty acids (n-3 PUFA) reduce the incidence of cardiovascular diseases and sudden cardiac death. We investigated the cardioprotective effects of n-3 PUFA in aged spontaneously hypertensive rats (SHR) and possible cellular mechanisms involved. Male and female 14-moth-old SHR were fed with n-3 PUFA (Vesteralens, Norway, 20 mg/day for two months) and compared with untreated SHR. Results showed that n-3 PUFA supplementation led to 1) significant decline of blood pressure; 2) suppression of inducible ventricular fibrillation (VF) by 57 % (male) and 67 % (female), although the arrhythmogenic substrates, like fibrosis, hypertrophy and abnormal gap junctions distribution were not eliminated; 3) preservation of the cardiomyocytes and the integrity of their junctions; 4) enhancement of energetic metabolism enzyme activity; 5) augmentation of capillary density associated with increased alkaline phosphatase and decreased dipeptidyl peptidase-4 (DPP4) activity and 6/ increase in gap junction channel connexin-43 expression. Thus, aged male as well as female SHR benefit from n-3 PUFA supplementation that results in decrease in VF susceptibility, partly due to an improvement of myocardial metabolic state, cardiomyocyte and cell-to-cell junctions integrity and Cx43 up-regulation.

Genetic component represents an important factor in the development of hypertension, which is known to be associated with changes in expression of vascular gap junction protein connexin 43 (Cx43). The aim of the study was to examine the distribution and expression of Cx43 in the aortic endothelium of adult normotensive Wistar rats (W), borderline hypertensive rats (BHR) and spontaneously hypertensive rats (SHR). Rings of the thoracic aorta were processed for immunofluorescence and Western blot analysis of endothelial Cx43 and for electron microscopy. Both, BHR and SHR exhibited significantly increased blood pressure vs. W (132±2 mm Hg and 185±3 mm Hg vs. 110±2 mm Hg). Reduced Cx43 immunofluorescence was observed in the endothelium of BHR and these alterations were more pronounced in SHR. Western blot analysis showed significant suppression of Cx43 expression in the aorta of both BHR (p