Resveratrol and diabetic cardiac function: focus on recent in vitro and in vivo studies.

Resveratrol, a natural phytoalexin found in wine has the potential to impact a variety of human diseases. Resveratrol like other polyphenols activates many of the same intracellular pathways as those activated by caloric restriction. It can quench reactive oxidative species, ROS and induce eNOS and iNOS expression. Resveratrol also can activate SIRT1, a NAD⁺-dependent deacetylase, that leads an improved in mitochondrial function, and then this procedure turns to activate the transcription factor Nrf2 that coordinates expression of key antioxidant mechanisms by binding to the antioxidant response elements. Resveratrol provides cardioprotection by triggering preconditioning and inducing autophagy. It also presents chemical similarities with estrogen and was reported to activate both nuclear and extranuclear estrogen receptors. Resveratrol treatment alleviated diabetes-induced cardiovascular system disorders via different endogeneous signaling pathways including oxidative stress/antioxidant defense system, glucose/insulin metabolism, overexpression of iNOS/nitrotyrosine, and preconditioning. Resveratrol treatment significantly reduced the blood glucose level in STZ-treated type 1 diabetic animals through insulin-dependent and insulin-independent pathways. Resveratrol triggers some of the similar intracellular insulin signalling components in myocardium such as eNOS, AKT through the AMPK pathway, and plays an essential role in Glut-4 translocation and glucose uptake in STZ-induced diabetic myocardium. However, resveratrol can exhibit hormetic action expressing health benefits at lower doses whereas being detrimental at higher doses. It might also exert antidiabetic effects by activating SIRT1 directly in the brain. This review includes a summary of the role of resveratrol and diabetic cardiac function including a brief discussion on in vitro and in vivo studies as well as our original observations in diabetic rats.

Ahnak1 modulates L-type Ca(2+) channel inactivation of rodent cardiomyocytes.

Ahnak1, a giant 700 kDa protein, has been implicated in Ca(2+) signalling in various cells. Previous work suggested that the interaction between ahnak1 and Cavbeta(2) subunit plays a role in L-type Ca(2+) current (I (CaL)) regulation. Here, we performed structure-function studies with the most C-terminal domain of ahnak1 (188 amino acids) containing a PxxP consensus motif (designated as 188-PSTP) using ventricular cardiomyocytes isolated from rats, wild-type (WT) mice and ahnak1-deficient mice. In vitro binding studies revealed that 188-PSTP conferred high-affinity binding to Cavbeta(2) (K (d) approximately 60 nM). Replacement of proline residues by alanines (188-ASTA) decreased Cavbeta(2) affinity about 20-fold. Both 188-PSTP and 188-ASTA were functional in ahnak1-expressing rat and mouse cardiomyocytes during whole-cell patch clamp. Upon intracellular application, they increased the net Ca(2+) influx by enhancing I (CaL) density and/or increasing I (CaL) inactivation time course without altering voltage dependency. Specifically, 188-ASTA, which failed to affect I (CaL) density, markedly slowed I (CaL) inactivation resulting in a 50-70% increase in transported Ca(2+) during a 0 mV depolarising pulse. Both ahnak1 fragments also slowed current inactivation with Ba(2+) as charge carrier. By contrast, neither 188-PSTP nor 188-ASTA affected any I (CaL) characteristics in ahnak1-deficient mouse cardiomyocytes. Our results indicate that the presence of endogenous ahnak1 is required for tuning the voltage-dependent component of I (CaL) inactivation by ahnak1 fragments. We suggest that ahnak1 modulates the accessibility of molecular determinants in Cavbeta(2) and/or scaffolds selectively different beta-subunit isoforms in the heart.

PKA phosphorylation activates the calcium release channel (ryanodine receptor) in skeletal muscle: defective regulation in heart failure.

The type 1 ryanodine receptor (RyR1) on the sarcoplasmic reticulum (SR) is the major calcium (Ca2+) release channel required for skeletal muscle excitation-contraction (EC) coupling. RyR1 function is modulated by proteins that bind to its large cytoplasmic scaffold domain, including the FK506 binding protein (FKBP12) and PKA. PKA is activated during sympathetic nervous system (SNS) stimulation. We show that PKA phosphorylation of RyR1 at Ser2843 activates the channel by releasing FKBP12. When FKB12 is bound to RyR1, it inhibits the channel by stabilizing its closed state. RyR1 in skeletal muscle from animals with heart failure (HF), a chronic hyperadrenergic state, were PKA hyperphosphorylated, depleted of FKBP12, and exhibited increased activity, suggesting that the channels are "leaky." RyR1 PKA hyperphosphorylation correlated with impaired SR Ca2+ release and early fatigue in HF skeletal muscle. These findings identify a novel mechanism that regulates RyR1 function via PKA phosphorylation in response to SNS stimulation. PKA hyperphosphorylation of RyR1 may contribute to impaired skeletal muscle function in HF, suggesting that a generalized EC coupling myopathy may play a role in HF.

[Transient receptor potential, TRP channels: a new family of channels broadly expressed]. / Les canaux TRP (transient receptor potential): une nouvelle famille de canaux à expression variée.

Transient receptor potential, TRP channels are a new superfamily of functionally versatile non-selective cation channels present from yeast to mammals. On the basis of their structural homology, TRP channels are subdivided in 7 groups : TRPC 1-7 Canonical, TRPV 1-6 Vanilloid, TRPM 1-8 Melastatin, TRPP 1-3 Polycystin, TRPML Mucolipin, TRPA Ankyrin and TRPN (NO mechanotransducer potential C), the latter not expressed in mammals. Their cloning and heterologous expression allowed to demonstrating that these channels are generally weakly voltage-dependent. They are activated by various ligands involving a signal transduction cascade as well as directly by multiple compounds, heat and pH. TRP channels are found in a broad range of cell types. TRP channels are essential in allowing animals to sense the outside world and cells to sense their local environment. Following mutations or anomalous behaviour, these channels have a major role in several human diseases.

Length and protein kinase A modulations of myocytes in cardiac myosin binding protein C-deficient mice.

OBJECTIVE: Beta-adrenergic stimulation modulates cardiac contractility through protein kinase A (PKA), which phosphorylates proteins such as troponin I (cTnI) and C-protein (cMyBP-C). The relative contributions of cTnI and cMyBP-C to the regulation of myofilament Ca(2+) sensitivity are still controversial because of difficulty in targeting specific protein phosphorylation. Recently, impaired relaxation was found in cMyBP-C-deficient mice (KO) in vivo under basal conditions and after beta-adrenergic stimulation. The goal of this study was to analyse the length-dependent and PKA-dependent modulations of the cardiac contractile machinery in a mouse model lacking cMyBP-C. METHODS: In the present work, we studied the PKA effect on myofilament Ca(2+) sensitivity of left ventricular skinned myocytes isolated from 5-week- and 55-week-old wild-type (WT) and cMyBP-C knockout (KO) mice at 1.9 and 2.3 mum sarcomere lengths (SL). The cTnI content and phosphorylation status were examined by Western blot analysis. RESULTS: Without PKA stimulation and at the shorter SL, Ca(2+) sensitivity was higher in KO compared to WT. The difference disappeared at the longer SL. No difference in passive tension or maximal active tension was observed. PKA stimulation induced a desensitization of WT myofilaments at both SL but had almost no effect in KO myofilaments despite similar levels of cTnI phosphorylation. We also observed expression of slow skeletal TnI in KO animals that was not correlated with the PKA effects. CONCLUSION: The results suggest that cMyBP-C contributes to the regulation of cardiac contraction at short sarcomere length and that myofilament desensitization induced by PKA requires the presence of cMyBP-C and does not depend only upon TnI phosphorylation.

Role of T-type Ca2+ channels in the heart.

After the first demonstration 30 years ago that Ca2+ could permeate through two different channels, the occurrence and role of T-type Ca2+ current, ICaT have been the matter of hundreds of publications, including the two 1985' reports in various cardiac tissues and species. Except for its specific biophysical characteristics, ICaT is no longer so easily distinguished from the L-type Ca2+ current, ICaL, since it is also sensitive to multiple compounds and various neuromediators including the beta-adrenergic agonists. Changes in ICaT occur during development, so that while it is recorded in all embryonic and neonatal cells investigated, ICaT has been reported in adult ventricular cells of only few species in control. However, under various pathological conditions, ICaT is often recorded at some phases of remodelling at least in some localized area and one or more of the three channel proteins, Cav3.1-3.3 are clearly re-expressed under the influence of IGF-1, endothelin, and angiotensin II. ICaT contributes to the control of electrical activity including pacemaker and arrhythmia. Furthermore ICaT, and its low-depolarisation window current, participate in Ca2+ entry, so that ICaT has been involved in the release of Ca2+ from internal stores, the Ca2+-induced Ca2+ release mechanism, although at much lower level than ICaL. ICaT contributes also to Ca2+-dependent hormonal secretion. This review further emphasizes the difficulties encountered in analysing this current.

Effects of diabetes on ryanodine receptor Ca release channel (RyR2) and Ca2+ homeostasis in rat heart.

The defects identified in the mechanical activity of the hearts from type 1 diabetic animals include alteration of Ca2+ signaling via changes in critical processes that regulate intracellular Ca2+ concentration. These defects result partially from a dysfunction of cardiac ryanodine receptor calcium release channel (RyR2). The present study was designed to determine whether the properties of the Ca2+ sparks might provide insight into the role of RyR2 in the altered Ca2+ signaling in cardiomyocytes from diabetic animals when they were analyzed together with Ca2+ transients. Basal Ca2+ level as well as Ca2+-spark frequency of cardiomyoctes isolated from 5-week streptozotocin (STZ)-induced diabetic rats significantly increased with respect to aged-matched control rats. Ca2+ transients exhibited significantly reduced amplitude and prolonged time courses as well as depressed Ca2+ loading of sarcoplasmic reticulum in diabetic rats. Spatio-temporal properties of the Ca2+ sparks in cardiomyocytes isolated from diabetic rats were also significantly altered to being almost parallel to the changes of Ca2+ transients. In addition, RyR2 from diabetic rat hearts were hyperphosphorylated and protein levels of both RyR2 and FKBP12.6 depleted. These data show that STZ-induced diabetic rat hearts exhibit altered local Ca2+ signaling with increased basal Ca2+ level.

Transmural stretch-dependent regulation of contractile properties in rat heart and its alteration after myocardial infarction.

The "stretch-sensitization" response is essential to the regulation of heart contractility. An increase in diastolic volume improves systolic contraction. The cellular mechanisms of this modulation, the Frank-Starling law, are still uncertain. Moreover, their alterations in heart failure remains controversial. Here, using left ventricular skinned rat myocytes, we show a nonuniform stretch-sensitization of myofilament activation across the ventricular wall. Stretch-dependent Ca2+ sensitization of myofilaments increases from sub-epicardium to sub-endocardium and is correlated with an increase in passive tension. This passive tension-dependent component of myofibrillar activation is not associated with expression of titin isoforms, changes in troponin I level, and phosphorylation status. Instead, we observe that stretch induces phosphorylation of ventricular myosin light chain 2 isoform (VLC2b) in sub-endocardium specifically. Thus, VLC2b phosphorylation could act as a stretch-dependent modulator of activation tuned within normal heart. Moreover, in postmyocardial infarcted rat, the gradient of stretch-dependent Ca2+ sensitization disappears associated with a lack of VLC2b phosphorylation in sub-endocardium. In conclusion, nonuniformity is a major characteristic of the normal adult left ventricle (LV). The heterogeneous myocardial deformation pattern might be caused not only by the morphological heterogeneity of the tissue in the LV wall, but also by the nonuniform contractile properties of the myocytes across the wall. The loss of a contractile transmural gradient after myocardial infarction should contribute to the impaired LV function.

Ahnak is critical for cardiac Ca(V)1.2 calcium channel function and its beta-adrenergic regulation.

Defective L-type Ca2+ channel (I(CaL)) regulation is one major cause for contractile dysfunction in the heart. The I(CaL) is enhanced by sympathetic nervous stimulation: via the activation of beta-adrenergic receptors, PKA phosphorylates the alpha1C(Ca(V)1.2)- and beta2-channel subunits and ahnak, an associated 5643-amino acid (aa) protein. In this study, we examined the role of a naturally occurring, genetic variant Ile5236Thr-ahnak on I(CaL). Binding experiments with ahnak fragments (wild-type, Ile5236Thr mutated) and patch clamp recordings revealed that Ile5236Thr-ahnak critically affected both beta2 subunit interaction and I(CaL) regulation. Binding affinity between ahnak-C1 (aa 4646-5288) and beta2 subunit decreased by approximately 50% after PKA phosphorylation or in the presence of Ile5236Thr-ahnak peptide. On native cardiomyocytes, intracellular application of this mutated ahnak peptide mimicked the PKA-effects on I(CaL) increasing the amplitude by approximately 60% and slowing its inactivation together with a leftward shift of its voltage dependency. Both mutated Ile5236Thr-peptide and Ile5236Thr-fragment (aa 5215-5288) prevented specifically the further up-regulation of I(CaL) by isoprenaline. Hence, we suggest the ahnak-C1 domain serves as physiological brake on I(CaL). Relief from this inhibition is proposed as common pathway used by sympathetic signaling and Ile5236Thr-ahnak fragments to increase I(CaL). This genetic ahnak variant might cause individual differences in I(CaL) regulation upon physiological challenges or therapeutic interventions.

Increased exchange current but normal Ca2+ transport via Na+-Ca2+ exchange during cardiac hypertrophy after myocardial infarction.

Hypertrophied and failing cardiac myocytes generally show alterations in intracellular Ca2+ handling associated with changes in the contractile function and arrhythmogenicity. The cardiac Na+-Ca2+ exchange (NCX) is an important mechanism for Ca2+ extrusion and cell relaxation. Its possible involvement in changes of excitation-contraction coupling (EC-coupling) with disease remains uncertain. We analyzed the NCX function in rat ventricular myocytes 5 to 6 months after experimental myocardial infarction (PMI) produced by left coronary artery ligation and from sham-operated (SO) hearts. Caged Ca2+ was dialyzed into the cytoplasm via a patch-clamp pipette and Ca2+ was released by flash photolysis to activate NCX and measure the associated currents (I(NaCa)), whereas [Ca2+]i changes were simultaneously recorded with a confocal microscope. I(NaCa) density normalized to the [Ca2+]i jumps was 2.6-fold higher in myocytes from PMI rats. The level of total NCX protein expression in PMI myocytes was also increased. Interestingly, although the I(NaCa) density in PMI cells was larger, PMI and SO myocytes presented virtually identical Ca2+ transport via the NCX. This discrepancy was explained by a reduced surface/volume ratio (34.8%) observed in PMI cells. We conclude that the increase in NCX density may be a mechanism to maintain the required Ca2+ extrusion from a larger cell to allow adequate relaxation.

Ca2+-dependent reduction of IK1 in rat ventricular cells: a novel paradigm for arrhythmia in heart failure?

OBJECTIVES: We investigated the inward rectifier potassium current (I(K1)), which can be blocked by intracellular Ca(2+), in heart failure (HF). METHODS: We used the whole-cell patch-clamp technique to record I(K1) from single rat ventricular myocytes in voltage-clamp conditions. Fluorescence measurements of diastolic Ca(2+) were performed with Indo-1 AM. HF was examined 8 weeks after myocardial infarction (coronary artery ligation). RESULTS: I(K1) was reduced and diastolic Ca(2+) was increased in HF cells. The reduction of I(K1) was attenuated when EGTA was elevated from 0.5 to 10 mM in the patch pipette and prevented with high BAPTA (20 mM). Ryanodine (100 nM) and FK506 (10 microM), both of which promote spontaneous SR Ca(2+) release from ryanodine receptor (RyR2) during diastole, reproduced the effect of HF on I(K1) in normal cells but had no effect in HF cells. The effects of ryanodine and FK506 were not additive and were prevented by BAPTA. Rapamycin (10 microM), which removes FKBP binding proteins from RyR2 with no effect on calcineurin, mimicked the effect of FK506 on I(K1). Cyclosporine A (10 microM), which inhibits calcineurin via cyclophilins, had no effect. In both HF cells and normal cells treated by FK506, the protein kinase C (PKC) inhibitor staurosporine totally restored the inward component of I(K1), but only partially restored its outward component at potentials corresponding to the late repolarizing phase of the action potential (-80 to -40 mV). CONCLUSIONS: I(K1) is reduced by elevated diastolic Ca(2+)in HF, which involves in parallel PKC-dependent and PKC-independent mechanisms. This regulation provides a novel paradigm for Ca(2+)-dependent modulation of membrane potential in HF. Since enhanced RyR2-mediated Ca(2+)release also reduces I(K1), this paradigm might be relevant for arrhythmias related to acquired or inherited RyR2 dysfunction.

The role of ryanodine receptors and consequences of their alterations during cardiac insufficiency.

Congestive heart failure (CHF) is a leading cause of death. Although changes to other components contribute, it is generally agreed that much of the contractile deficit is due to reduced Ca(2+) homeostasis that includes alterations in Ca(2+) current and action potential characteristics, together with reduced Ca(2+) transient amplitude. CHF is also associated with progressive skeletal muscle dysfunction. In both cardiac and skeletal muscles, the global increase in myoplasmic Ca(2+) during depolarization, or Ca(2+) transient, appears to consist of the summation of large numbers of local, unitary Ca(2+) release events (ie, Ca(2+) sparks) resulting from the activity of a cluster of ryanodine receptors (RyRs) (ie, RyR1 or RyR2 in skeletal and cardiac muscles, respectively). RyR2 channels from failing hearts have been shown to be hyperphosphorylated by protein kinase A, leading to dissociation of FK506-binding protein 12.6 and altered RyR2 channel function. After reviewing the alterations occurring in cardiomyocytes, the present report summarizes the intrinsic alterations of Ca(2+) homeostasis in rat extensor digitorum longus skeletal muscle. They include a weaker and prolonged Ca(2+) transient that could be attributed to both a lower synchronization of the individual Ca(2+) sparks and a lower synchronization of these events triggered upon depolarization. As in cardiac muscle, these alterations in sarcoplasmic reticulum function are associated with protein kinase A-induced hyperphosphorylation of RyR1 and a concomitant reduction in FK506-binding protein 12. These specific alterations in RyR1-dependent Ca(2+) release could play a significant role in the specific force decrements in skeletal muscle as well as in the remodelling that occurs secondary to CHF.

Occurrence of a tetrodotoxin-sensitive calcium current in rat ventricular myocytes after long-term myocardial infarction.

OBJECTIVE: To determine the characteristics of a TTX-sensitive Ca(2+) current that occurred only following remodelling after myocardial infarction in Wistar rat. METHODS: Using the whole-cell patch-clamp technique, we studied ionic inward current in myocytes isolated from four different ventricular regions of control Wistar rat hearts, or from hearts 4 to 6 months after ligation of the left coronary artery. Inward current characteristics were also analysed in Xenopus laevis oocytes that heterologously expressed the human sodium channel alpha-subunit Nav1.5. The effects of oxidative stress by hydrogen peroxide or tert-butyl-hydroxyperoxide as well as those of PKA-dependent phosphorylation, which partly mimic the pathological conditions, were investigated on control cardiomyocytes and Nav1.5-expressing oocytes. RESULTS: In Na-free solution, a low-threshold, tetrodotoxin-sensitive inward current was found in 20 out of 78 cells isolated from 16 post-myocardial infarcted (PMI) cardiomyocytes but not in cardiomyocytes from young and sham rat hearts. This current exhibited kinetics and pharmacological properties similar to the I(Ca(TTX)) current previously reported. I(Ca(TTX))-like current was critically dependent on extracellular Na(+) and was reduced by micromolar Na(+) concentrations. Neither in normal rat cardiomyocytes nor in Nav1.5-expressing oocytes could a I(Ca(TTX))-like current be elicited in Na(+)-free extracellular solution, even after oxidative stress or PKA-dependent phosphorylation. CONCLUSIONS: Our data suggest that I(Ca(TTX))-like current in PMI myocytes does not arise from classical Na(+) channels modified by oxidative stress or PKA phosphorylation and most probably represents a different Na(+) channel type re-expressed in some cells after remodelling.

Altered communication between L-type calcium channels and ryanodine receptors in heart failure.

Heart failure (HF) is a progressive syndrome that appears as the final phase of most cardiac diseases and is manifested as a decreased contractile function. Contraction in cardiomyocytes arises by the Ca2+ induced Ca2+ release mechanism, where Ca2+ entry (ICa) through Ca2+ channels (DHPRs) activates Ca2+ release channels (RyRs) in the junctional sarcoplasmic reticulum (SR). This is the base of cardiac excitation-contraction (EC) coupling. To elucidate the mechanisms underlying depressed function of the failing heart, analysis of EC coupling main elements have been undertaken. ICa density is usually maintained in HF. However, failing myocytes show a reduced SR Ca2+ release. Then, if the trigger of SR Ca2+ release is maintained, why is SR Ca2+ release depressed in HF? Analyses of the DHPR-RyR coupling efficiency have revealed a decrease in the ICa efficacy to trigger Ca2+ release in failing myocytes. In terminal heart failure without hypertrophy, a decrease in SR Ca2+ load can account for the decreased SR Ca2+ release. Fewer Ca2+ sparks (elementary units of SR Ca2+ release) are triggered by an equivalent ICa in hypertrophied failing myocytes, suggesting a functional or spatial reorganization of the space T-tubule junctional SR. This theory is supported by new data showing that the T-tubule density is reduced in failing cells.

SR33805, a Ca2+ antagonist with length-dependent Ca2+ -sensitizing properties in cardiac myocytes.

1. This study examined the effects of SR33805, a fantofarone derivative with reported strong Ca(2+) -antagonistic properties, on the contractile properties of intact and skinned rat ventricular myocytes. 2. On intact cells loaded with the Ca(2+)-fluorescent indicator Indo-1, the application of low concentrations of SR33805 enhanced the amplitude of unloaded cell shortening and decreased the duration of cell shortening. Amplitude of the Ca(2+) transient was also decreased. 3. These effects were accompanied with a shortening of the action potential and a dose-dependent blockade of L-type calcium current (IC(50)=2.4 x 10(-8) M). 4. On skinned cardiac cells, the application of a low SR33805 concentration (10(-8) M) induced a significant increase in maximal Ca(2+)-activated force at the two-tested sarcomere lengths (SLs), 1.9 and 2.3 microm. 5. The application of a larger dose of SR33805 (10(-6)-10(-5) M) induced a significant leftward shift of the tension-pCa relation that accounts for Ca(2+)-sensitization of the myofilaments, particularly at 2.3 microm SL. 6. In conclusion, despite its strong Ca(2+)-antagonistic properties SR33805 increases cardiac cell contractile activity as a consequence of its Ca(2+)-sensitizing effects. These effects are attributable to both an increase in the maximal Ca(2+)-activated force and a length-dependent Ca(2+)-sensitization.

ß-blocker timolol prevents arrhythmogenic Ca²âº release and normalizes Ca²âº and Zn²âº dyshomeostasis in hyperglycemic rat heart.

Defective cardiac mechanical activity in diabetes results from alterations in intracellular Ca(2+) handling, in part, due to increased oxidative stress. Beta-blockers demonstrate marked beneficial effects in heart dysfunction with scavenging free radicals and/or acting as an antioxidant. The aim of this study was to address how ß-blocker timolol-treatment of diabetic rats exerts cardioprotection. Timolol-treatment (12-week), one-week following diabetes induction, prevented diabetes-induced depressed left ventricular basal contractile activity, prolonged cellular electrical activity, and attenuated the increase in isolated-cardiomyocyte size without hyperglycemic effect. Both in vivo and in vitro timolol-treatment of diabetic cardiomyocytes prevented the altered kinetic parameters of Ca(2+) transients and reduced Ca(2+) loading of sarcoplasmic reticulum (SR), basal intracellular free Ca(2+) and Zn(2+) ([Ca(2+)]i and [Zn(2+)]i), and spatio-temporal properties of the Ca(2+) sparks, significantly. Timolol also antagonized hyperphosphorylation of cardiac ryanodine receptor (RyR2), and significantly restored depleted protein levels of both RyR2 and calstabin2. Western blot analysis demonstrated that timolol-treatment also significantly normalized depressed levels of some [Ca(2+)]i-handling regulators, such as Na(+)/Ca(2+) exchanger (NCX) and phospho-phospholamban (pPLN) to PLN ratio. Incubation of diabetic cardiomyocytes with 4-mM glutathione exerted similar beneficial effects on RyR2-macromolecular complex and basal levels of both [Ca(2+)]i and [Zn(2+)]i, increased intracellular Zn(2+) hyperphosphorylated RyR2 in a concentration-dependent manner. Timolol also led to a balanced oxidant/antioxidant level in both heart and circulation and prevented altered cellular redox state of the heart. We thus report, for the first time, that the preventing effect of timolol, directly targeting heart, seems to be associated with a normalization of macromolecular complex of RyR2 and some Ca(2+) handling regulators, and prevention of Ca(2+) leak, and thereby normalization of both [Ca(2+)]i and [Zn(2+)]i homeostasis in diabetic rat heart, at least in part by controlling the cellular redox status of hyperglycemic cardiomyocytes.

Ryanodine receptor: a new therapeutic target to control diabetic cardiomyopathy.

Diabetes mellitus is a major risk factor for cardiovascular complications. Intracellular Ca(2+) release plays an important role in the regulation of muscle contraction. Sarcoplasmic reticulum Ca(2+) release is controlled by dedicated molecular machinery, composed of a complex of cardiac ryanodine receptors (RyR2s). Acquired and genetic defects in this complex result in a spectrum of abnormal Ca(2+) release phenotypes in heart. Cardiovascular dysfunction is a leading cause for mortality of diabetic individuals due, in part, to a specific cardiomyopathy, and to altered vascular reactivity. Cardiovascular complications result from multiple parameters, including glucotoxicity, lipotoxicity, fibrosis, and mitochondrial uncoupling. In diabetic subjects, oxidative stress arises from an imbalance between production of reactive oxygen and nitrogen species and capability of the system to readily detoxify reactive intermediates. To date, the etiology underlying diabetes-induced reductions in myocyte and cardiac contractility remains incompletely understood. However, numerous studies, including work from our laboratory, suggest that these defects stem in part from perturbation in intracellular Ca(2+) cycling. Since the RyR2s are one of the well-characterized redox-sensitive ion channels in heart, this article summarizes recent findings on redox regulation of cardiac Ca(2+) transport systems and discusses contributions of redox regulation to pathological cardiac function in diabetes.

Vitamin E in oxidant stress-related cardiovascular pathologies: focus on experimental studies.

The scope of this review is to summarize the important roles of vitamin E family members as protective agents in cardiovascular pathologies of different types of disease states and particularly in diabetes, including some of our research results, to illustrate how this recent knowledge is helping to better understand the roles of the vitamin E family in biology, in animals and humans specifically. Cardiovascular disease, a general name for a wide variety of diseases, disorders and conditions, is caused by disorders of the heart and blood vessels. Cardiovascular disease is the world's largest killer, claiming 17.1 million lives a year. Cardiovascular complications result from multiple parameters including glucotoxicity, lipotoxicity, fibrosis. Obesity and diabetes mellitus are also often linked to cardiovascular disease. In fact, cardiovascular disease is the most life-threatening of the diabetic complications and diabetics are 2- to 4-fold more likely to die of cardiovascular-related causes than non-diabetics. In order to prevent the tendency of cardiovascular disease, primary prevention is needed by modifying risk factors. Several recent studies, besides earlier ones, have reported beneficial effects of therapy with antioxidant agents, including trace elements, vitamins (E and/or C), other antioxidants, against the cardiovascular dysfunction. Hence, the use of peroxisome proliferator activated receptor-α (PPARα) agonists to reduce fatty acid oxidation, of trace elements such as selenium as antioxidant and other antioxidants such as vitamins E and C, contributes to the prevention of these dysfunctions. Moreover, therapy with antioxidants and the above vitamins to prevent or delay the onset and development of cardiovascular complications in diabetic patients and animal models has been investigated although these studies showed inconsistent results.

Intracellular free zinc during cardiac excitation-contraction cycle: calcium and redox dependencies.

AIMS: Zinc exists in biological systems as bound and histochemically reactive free Zn(2+). It is an essential structural constituent of many proteins, including enzymes from cellular signalling pathways, in which it functions as a signalling molecule. In cardiomyocytes at rest, Zn(2+) concentration is in the nanomolar range. Very little is known about precise mechanisms controlling the intracellular distribution of Zn(2+) and its variations during cardiac function. METHODS AND RESULTS: Live-cell detection of intracellular Zn(2+) has become feasible through the recent development of Zn(2+)-sensitive and -selective fluorophores able to distinguish Zn(2+) from Ca(2+). Here, in freshly isolated rat cardiomyocytes, we investigated the rapid changes in Zn(2+) homeostasis using the Zn(2+)-specific fluorescent dye, FluoZin-3, in comparison to Ca(2+)-dependent fluo-3 fluorescence. Zn(2+) sparks and Zn(2+) transients, in quiescent and electrically stimulated cardiomyocytes, respectively, were visualized in a similar manner to known rapid Ca(2+) changes. Both Zn(2+) sparks and Zn(2+) transients required Ca(2+) entry. Inhibiting the sarcoplasmic reticulum Ca(2+) release or increasing the Ca(2+) load in a low-Na(+) solution suppressed or increased Zn(2+) movements, respectively. Mitochondrial inhibitors slightly reduced both Zn(2+) sparks and Zn(2+) transients. Oxidation by H2O2 facilitated and acidic pH inhibited the Ca(2+)-dependent Zn(2+) release. CONCLUSION: It is proposed that Zn(2+) release during the cardiac cycle results mostly from intracellular free Ca(2+) increase, triggering production of reactive oxygen species that induce changes in metal-binding properties of metallothioneins and other redox-active proteins, aside from ionic exchange on these proteins.