Overexpression of Na+/Ca2+ exchanger gene attenuates postinfarction myocardial dysfunction

We monitored myocardial function in postinfarcted wild-type (WT) and transgenic (TG) mouse hearts with overexpression of the cardiac Na(+)/Ca(2+) exchanger. Five weeks after infarction, cardiac function was better maintained in TG than WT mice [left ventricular (LV) systolic pressure: WT, 41 +/- 2; TG, 58 +/- 3 mmHg; P < 0.05; maximum rising rate of LV pressure (+dP/dt(max)): WT, 3,750 +/- 346; TG, 5,075 +/- 334 mmHg/s; P < 0.05]. The isometric contractile response to beta-adrenergic stimulation was greater in papillary muscles from TG than WT mice (WT, 13.2 +/- 0.9; TG, 16.3 +/- 1.0 mN/mm(2) at 10(-4) M isoproterenol). The sarcoplasmic reticulum (SR) Ca(2+) content investigated by rapid cooling contractures in papillary muscles was greater in TG than WT mouse hearts. We conclude that myocardial function is better preserved in TG mice 5 wk after infarction, which results from enhanced SR Ca(2+) content via overexpression of the Na(+)/Ca(2+) exchanger.     

Defective intracellular Ca(2+) signaling contributes to cardiomyopathy in Type 1 diabetic rats

The goal of the study was to determine whether defects in intracellular Ca(2+) signaling contribute to cardiomyopathy in streptozotocin (STZ)-induced diabetic rats. Depression in cardiac systolic and diastolic function was traced from live diabetic rats to isolated individual myocytes. The depression in contraction and relaxation in myocytes was found in parallel with depression in the rise and decline of intracellular free Ca(2+) concentration ([Ca(2+)](i)). The sarcoplasmic reticulum (SR) Ca(2+) store and rates of Ca(2+) release and resequestration into SR were depressed in diabetic rat myocytes. The rate of Ca(2+) efflux via sarcolemmal Na(+)/Ca(2+) exchanger was also depressed. However, there was no change in the voltage-dependent L-type Ca(2+) channel current that triggers Ca(2+) release from the SR. The depression in SR function was associated with decreased SR Ca(2+)-ATPase and ryanodine receptor proteins and increased total and nonphosphorylated phospholamban proteins. The depression of Na(+)/Ca(2+) exchanger activity was associated with a decrease in its protein level. Thus it is concluded that defects in intracellular Ca(2+) signaling caused by alteration of expression and function of the proteins that regulate [Ca(2+)](i) contribute to cardiomyopathy in STZ-induced diabetic rats. The increase in phospholamban, decrease in Na(+)/Ca(2+) exchanger, and unchanged L-type Ca(2+) channel activity in this model of diabetic cardiomyopathy are distinct from other types of cardiomyopathy.     

The Na+/Ca2+ exchanger is active and working in the reverse mode in human umbilical artery smooth muscle cells

The data presented in this work suggest that in human umbilical artery (HUA) smooth muscle cells, the Na(+)/Ca(2+) exchanger (NCX) is active and working in the reverse mode. This supposition is based on the following results: (i) microfluorimetry in HUA smooth muscle cells in situ showed that a Ca(2+)-free extracellular solution diminished intracellular Ca(2+) ([Ca(2+)](i)), and KB-R7943 (5microM), a specific inhibitor of the Ca(2+) entry mode of the exchanger, also decreased [Ca(2+)](i) (40.6+/-4.5% of Ca(2+)-free effect); (ii) KB-R7943 produced the relaxation of HUA rings (-24.7+/-7.3gF/gW, n=8, p<0.05); (iii) stimulation of the NCX by lowering extracellular Na(+) increases basal [Ca(2+)](i) proportionally to Na(+) reduction (Delta fluorescence ratio=0.593+/-0.141 for Na(+)-free solution, n=8) and HUA rings' contraction (peak force=181.5+/-39.7 for 130mM reduction, n=8), both inhibited by KB-R7943 and a Ca(2+)-free extracellular solution. In conclusion, the NCX represents an important Ca(2+) entry route in HUA smooth muscle cells.     

Altered E-C coupling in rat ventricular myocytes from failing hearts 6 wk after MI

Excitation-contraction (E-C) coupling was investigated in rat hearts 6 wk after induction of myocardial infarction (MI) by ligation of the left coronary artery. Heart weight was increased by 74% and left ventricular end-diastolic pressure was 23 +/- 2 mmHg in MI compared with 8 +/- 2 mmHg in sham-operated controls (Sham, P < 0.001). Cell shortening was measured in voltage-clamped myocytes at 36 degrees C. In solutions where Cs(+) had been replaced by K(+), the voltage dependence of contraction was sigmoidal between -20 and +100 mV in Sham and MI cells. Verapamil (20 microM) blocked L-type Ca(2+) current and reduced contraction in Sham cells by approximately 50% (P < 0.01) but did not decrease contraction significantly in MI cells at test potentials above +10 mV. Verapamil-insensitive contractions were blocked by Ni(2+) (5 mM). Na(+)/Ca(2+) exchange current was doubled in MI compared with Sham cells at test potentials between -20 and +80 mV (P < 0.05), whereas mRNA and protein expression increased by 30-40%. Finally, voltage dependence of contraction was bell shaped in Na(+)-free solutions, but contraction was significantly increased in MI cells over a wider voltage range (P < 0.05). The insensitivity to Ca(2+) channel block in MI cells may result from an increased contribution of the Na(+)/Ca(+) exchanger to triggering of E-C coupling. These results suggest significant changes in E-C coupling in the hypertrophy and failure that develop in response to extensive MI.     

Modification of sarcolemmal Na+-K+-ATPase and Na+/Ca2+ exchanger expression in heart failure by blockade of renin-angiotensin system

The activities of both sarcolemmal (SL) Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchanger, which maintain the intracellular cation homeostasis, have been shown to be depressed in heart failure due to myocardial infarction (MI). Because the renin-angiotensin system (RAS) is activated in heart failure, this study tested the hypothesis that attenuation of cardiac SL changes in congestive heart failure (CHF) by angiotensin-converting enzyme (ACE) inhibitors is associated with prevention of alterations in gene expression for SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchanger. CHF in rats due to MI was induced by occluding the coronary artery, and 3 wk later the animals were treated with an ACE inhibitor, imidapril (1 mg.kg(-1).day(-1)), for 4 wk. Heart dysfunction and cardiac hypertrophy in the infarcted animals were associated with depressed SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities. Protein content and mRNA levels for Na(+)/Ca(2+) exchanger as well as Na(+)-K(+)-ATPase alpha(1)-, alpha(2)- and beta(1)-isoforms were depressed, whereas those for alpha(3)-isoform were increased in the failing heart. These changes in SL activities, protein content, and gene expression were attenuated by treating the infarcted animals with imidapril. The beneficial effects of imidapril treatment on heart function and cardiac hypertrophy as well as SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities in the infarcted animals were simulated by enalapril, an ACE inhibitor, and losartan, an angiotensin receptor antagonist. These results suggest that blockade of RAS in CHF improves SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities in the failing heart by preventing changes in gene expression for SL proteins.     

Targeted disruption of Na+/Ca2+ exchanger gene leads to cardiomyocyte apoptosis and defects in heartbeat

Ca(2+), which enters cardiac myocytes through voltage-dependent Ca(2+) channels during excitation, is extruded from myocytes primarily by the Na(+)/Ca(2+) exchanger (NCX1) during relaxation. The increase in intracellular Ca(2+) concentration in myocytes by digitalis treatment and after ischemia/reperfusion is also thought to result from the reverse mode of the Na(+)/Ca(2+) exchange mechanism. However, the precise roles of the NCX1 are still unclear because of the lack of its specific inhibitors. We generated Ncx1-deficient mice by gene targeting to determine the in vivo function of the exchanger. Homozygous Ncx1-deficient mice died between embryonic days 9 and 10. Their hearts did not beat, and cardiac myocytes showed apoptosis. No forward mode or reverse mode of the Na(+)/Ca(2+) exchange activity was detected in null mutant hearts. The Na(+)-dependent Ca(2+) exchange activity as well as protein content of NCX1 were decreased by approximately 50% in the heart, kidney, aorta, and smooth muscle cells of the heterozygous mice, and tension development of the aortic ring in Na(+)-free solution was markedly impaired in heterozygous mice. These findings suggest that NCX1 is required for heartbeats and survival of cardiac myocytes in embryos and plays critical roles in Na(+)-dependent Ca(2+) handling in the heart and aorta.     

The reverse mode of the Na(+)/Ca(2+) exchanger provides a source of Ca(2+) for store refilling following agonist-induced Ca(2+) mobilization

Agonist-induced contraction of airway smooth muscle (ASM) can be triggered by an elevation in the intracellular Ca(2+) concentration, primarily through the release of Ca(2+) from the sarcoplasmic reticulum (SR). The refilling of the SR is integral for subsequent contractions. It has been suggested that Ca(2+) entry via store-operated cation (SOC) and receptor-operated cation channels may facilitate refilling of the SR. Indeed, depletion of the SR activates substantial inward SOC currents in ASM that are composed of both Ca(2+) and Na(+). Accumulation of Na(+) within the cell may regulate Ca(2+) handling in ASM by forcing the Na(+)/Ca(2+) exchanger (NCX) into the reverse mode, leading to the influx of Ca(2+) from the extracellular domain. Since depletion of the SR activates substantial inward Na(+) current, it is conceivable that the reverse mode of the NCX may contribute to the intracellular Ca(2+) pool from which the SR is refilled. Indeed, successive contractions of bovine ASM, evoked by various agonists (ACh, histamine, 5-HT, caffeine) were significantly reduced upon removal of extracellular Na(+); whereas contractions evoked by KCl were unchanged by Na(+) depletion. Ouabain, a selective inhibitor of the Na(+)/K(+) pump, had no effect on the reductions observed under normal and zero-Na(+) conditions. KB-R7943, a selective inhibitor of the reverse mode of the NCX, significantly reduced successive contractions induced by all agonists without altering KCl responses. Furthermore, KB-R7943 abolished successive caffeine-induced Ca(2+) transients in single ASM cells. Together, these data suggest a role for the reverse mode of the NCX in refilling the SR in ASM following Ca(2+) mobilization.     

Effects of sprint training on contractility and [Ca(2+)](i) transients in adult rat myocytes.

The effects of 6-8 wk of high-intensity sprint training (HIST) on rat myocyte contractility and intracellular Ca(2+) concentration ([Ca(2+)](i)) transients were investigated. Compared with sedentary (Sed) myocytes, HIST induced a modest (5%) but significant (P < 0.0005) increase in cell length with no changes in cell width. In addition, the percentage of myosin heavy chain alpha-isoenzyme increased significantly (P < 0.02) from 0.566 +/- 0.077% in Sed rats to 0.871 +/- 0.006% in HIST rats. At all three (0.6, 1.8, and 5 mM) extracellular Ca(2+) concentrations ([Ca(2+)](o)) examined, maximal shortening amplitudes and maximal shortening velocities were significantly (P < 0.0001) lower and half-times of relaxation were significantly (P < 0.005) longer in HIST myocytes. HIST myocytes had significantly (P < 0.0001) higher diastolic [Ca(2+)](i) levels. Compared with Sed myocytes, systolic [Ca(2+)](i) levels in HIST myocytes were higher at 0.6 mM [Ca(2+)](o), similar at 1.8 mM [Ca(2+)](o), and lower at 5 mM [Ca(2+)](o). The amplitudes of [Ca(2+)](i) transients were significantly (P < 0.0001) lower in HIST myocytes. Half-times of [Ca(2+)](i) transient decline, an estimate of sarcoplasmic reticulum (SR) Ca(2+) uptake activity, were not different between Sed and HIST myocytes. Compared with Sed hearts, Western blots demonstrated a significant (P < 0.03) threefold decrease in Na(+)/Ca(2+) exchanger, but SR Ca(2+)-ATPase and calsequestrin protein levels were unchanged in HIST hearts. We conclude that HIST effected diminished myocyte contractile function and [Ca(2+)](i) transient amplitudes under the conditions studied. We speculate that downregulation of Na(+)/Ca(2+) exchanger may partly account for the decreased contractility in HIST myocytes.     

Enhanced calcium mobilization in rat ventricular myocytes during the onset of pressure overload-induced hypertrophy

Early cardiovascular changes evoked by pressure overload (PO) may reveal adaptive strategies that allow immediate survival to the increased hemodynamic load. In this study, systolic and diastolic Ca(2+) cycling was analyzed in left ventricular rat myocytes before (day 2, PO-2d group) and after (day 7, PO-7d group) development of hypertrophy subsequent to aortic constriction, as well as in myocytes from time-matched sham-operated rats (sham group). Ca(2+) transient amplitude was significantly augmented in the PO-2d group. In the PO-7d group, intracellular Ca(2+) concentration ([Ca(2+)](i)) was reduced during diastole, and mechanical twitch relaxation (but not [Ca(2+)](i) decline) was slowed. In PO groups, fractional sarcoplasmic reticulum (SR) Ca(2+) release at a twitch, SR Ca(2+) content, SR Ca(2+) loss during diastole, and SR-dependent integrated Ca(2+) flux during twitch relaxation were significantly greater than in sham-operated groups, whereas the relaxation-associated Ca(2+) flux carried by the Na(+)/Ca(2+) exchanger was not significantly changed. In the PO-7d group, mRNA levels of cardiac isoforms of SR Ca(2+)-ATPase (SERCA2a), phospholamban, calsequestrin, ryanodine receptor, and NCX were not significantly altered, but the SERCA2a-to-phospholamban ratio was increased 2.5-fold. Moreover, greater sensitivity to the inotropic effects of the beta-adrenoceptor agonist isoproterenol was observed in the PO-7d group. The results indicate enhanced Ca(2+) cycling between SR and cytosol early after PO imposition, even before hypertrophy development. Increase in SR Ca(2+) uptake may contribute to enhancement of excitation-contraction coupling (augmented SR Ca(2+) content and release) and protection against arrhythmogenesis due to buildup of [Ca(2+)](i) during diastole.     

Enhanced gene expression of Na(+)/Ca(2+) exchanger attenuates ischemic and hypoxic contractile dysfunction

Enhanced gene expression of the Na(+)/Ca(2+) exchanger in failing hearts may be a compensatory mechanism to promote influx and efflux of Ca(2+), despite impairment of the sarcoplasmic reticulum (SR). To explore this, we monitored intracellular calcium (Ca(i)(2+)) and cardiac function in mouse hearts engineered to overexpress the Na(+)/Ca(2+) exchanger and subjected to ischemia and hypoxia, conditions known to impair SR Ca(i)(2+) transport and contractility. Although baseline Ca(i)(2+) and function were similar between transgenic and wild-type hearts, significant differences were observed during ischemia and hypoxia. During early ischemia, Ca(i)(2+) was preserved in transgenic hearts but significantly altered in wild-type hearts. Transgenic hearts maintained 40% of pressure-generating capacity during early ischemia, whereas wild-type hearts maintained only 25% (P < 0.01). During hypoxia, neither peak nor diastolic Ca(i)(2+) decreased in transgenic hearts. In contrast, both peak and diastolic Ca(i)(2+) decreased significantly in wild-type hearts. The decline of Ca(i)(2+) was abbreviated in hypoxic transgenic hearts but prolonged in wild-type hearts. Peak systolic pressure decreased by nearly 10% in hypoxic transgenic hearts and >25% in wild-type hearts (P < 0.001). These data demonstrate that enhanced gene expression of the Na(+)/Ca(2+) exchanger preserves Ca(i)(2+) homeostasis during ischemia and hypoxia, thereby preserving cardiac function in the acutely failing heart.     

Ca(2+) handling in isolated human atrial myocardium

Physiologically, human atrial and ventricular myocardium are coupled by an identical beating rate and rhythm. However, contractile behavior in atrial myocardium may be different from that in ventricular myocardium, and little is known about intracellular Ca(2+) handling in human atrium under physiological conditions. We used rapid cooling contractures (RCCs) to assess sarcoplasmic reticulum (SR) Ca(2+) content and the photoprotein aequorin to assess intracellular Ca(2+) transients in atrial and ventricular muscle strips isolated from nonfailing human hearts. In atrial myocardium (n = 19), isometric twitch force frequency dependently (0. 25-3 Hz) increased by 78 +/- 25% (at 3 Hz; P < 0.05). In parallel, aequorin light signals increased by 111 +/- 57% (P < 0.05) and RCC amplitudes by 49 +/- 13% (P < 0.05). Similar results were obtained in ventricular myocardium (n = 13). SR Ca(2+) uptake (relative to Na(+)/Ca(2+) exchange) frequency dependently increased in atrial and ventricular myocardium (P < 0.05). With increasing rest intervals (1-240 s), atrial myocardium (n = 7) exhibited a parallel decrease in postrest twitch force (at 240 s by 68 +/- 5%, P < 0.05) and RCCs (by 49 +/- 10%, P < 0.05). In contrast, postrest twitch force and RCCs significantly increased in ventricular myocardium (n = 6). We conclude that in human atrial and ventricular myocardium the positive force-frequency relation results from increased SR Ca(2+) turnover. In contrast, rest intervals in atrial myocardium are associated with depressed contractility and intracellular Ca(2+) handling, which may be due to rest-dependent SR Ca(2+) loss (Ca(2+) leak) and subsequent Ca(2+) extrusion via Na(+)/Ca(2+) exchange. Therefore, the influence of rate and rhythm on mechanical performance is not uniform in atrial and ventricular myocardium.     

DMA增加正常大鼠心肌细胞钙瞬变和收缩

实验观察了钠氢交换或钠钙交换抑制剂 5 (N ,N 二甲基 )氨氯吡咪 (DMA)对正常和心肌肥厚大鼠分离心室肌细胞钙瞬变和细胞收缩的影响。通过负载荧光染料Fura 2 /Am ,应用离子影像分析系统 (IonImagingSystem)同步测定离体大鼠心肌细胞钙瞬变和细胞长度。结果表明 :DMA 10 μmol/L分别使钙瞬变和细胞缩短从对照组的 2 0 9.6 0± 5 4.96和 3.0 7± 0 .97μm增加到 2 38.5 0± 80 .41和 4.0 7± 1.0 2 μm (P <0 .0 5 ,n =7)。应用特异性反向钠钙交换阻断剂KB R7943可完全阻断DMA的激动作用。DMA还可使尼卡地平抑制L 型钙通道后的钙瞬变和细胞收缩增加。在肥厚心肌细胞 ,DMA表现出相同的药理作用 ,但对钙瞬变和细胞缩短的刺激作用更强。结果表明 :DMA可通过反向钠钙交换途径增加正常和肥厚大鼠心肌细胞钙瞬变和细胞收缩 ,且对肥厚心肌细胞的影响比对正常心肌细胞大。     

The effects of overexpression of the Na+/Ca2+ exchanger on calcium regulation in hypertrophied mouse cardiac myocytes

In cardiac hypertrophy and failure it has been shown that the amount of Na/Ca exchanger protein can increase. Several studies have investigated this modification in overt heart failure. However, the role of Na/Ca exchanger overexpression during the development of hypertrophy is unknown. To address this question we investigated Ca2+ regulation in an early stage of cardiac hypertrophy before signs of heart failure occurred and evaluated the role of Na/Ca exchanger overexpression. Cardiac hypertrophy was induced by a constant infusion of angiotensin II (Ang, 1 microg/min/kg) via an osmotic pump for 14 days. Thereafter, ventricular myocytes from either wild type (NON) or transgenic mice overexpressing the Na/Ca exchanger (TR) were isolated. Myocytes were loaded with indo-1 AM or fluo-4 AM to monitor cytoplasmic [Ca2+] with all experiments performed at 37 degrees C. In myocytes exposed to Ang there was an increase in cell capacitance of more than 20% indicating cellular hypertrophy. Ca2+ transients were prolonged in hypertrophied NON myocytes but not in TR myocytes. Action potentials had a less negative plateau in TR myocytes. Sarcoplasmic reticulum (SR) Ca2+ content, measured using rapid caffeine application, was greater in TR myocytes but unaffected by hypertrophy. Ca2+ spark frequency was significantly greater in TR. Na/Ca exchanger overexpression prevented the prolongation of the Ca2+ transient observed in hypertrophy and maintained a similar SR Ca2+ leak suggesting a compensatory role in Ca2+ regulation in hypertrophied cardiac myocytes from transgenic mice. We suggest this compensatory effect is mediated by increased SR Ca2+ content and faster Ca2+ removal via the Na/Ca exchanger.     

Modification of intracellular calcium concentration in cardiomyocytes by inhibition of sarcolemmal Na+/H+ exchanger

Although the Na(+)/H(+) exchanger (NHE) is considered to be involved in regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) through the Na(+)/Ca(2+) exchanger, the exact mechanisms of its participation in Ca(2+) handling by cardiomyocytes are not fully understood. Isolated rat cardiomyocytes were treated with or without agents that are known to modify Ca(2+) movements in cardiomyocytes and exposed to an NHE inhibitor, 5-(N-methyl-N-isobutyl)amiloride (MIA). [Ca(2+)](i) in cardiomyocytes was measured spectrofluorometrically with fura 2-AM in the absence or presence of KCl, a depolarizing agent. MIA increased basal [Ca(2+)](i) and augmented the KCl-induced increase in [Ca(2+)](i) in a concentration-dependent manner. The MIA-induced increase in basal [Ca(2+)](i) was unaffected by extracellular Ca(2+), antagonists of the sarcolemmal (SL) L-type Ca(2+) channel, and inhibitors of the SL Na(+)/Ca(2+) exchanger, SL Ca(2+) pump ATPase and mitochondrial Ca(2+) uptake. However, the MIA-induced increase in basal [Ca(2+)](i) was attenuated by inhibitors of SL Na(+)-K(+)-ATPase and sarcoplasmic reticulum (SR) Ca(2+) transport. On the other hand, the MIA-mediated augmentation of the KCl response was dependent on extracellular Ca(2+) concentration and attenuated by agents that inhibit SL L-type Ca(2+) channels, the SL Na(+)/Ca(2+) exchanger, SL Na(+)-K(+)-ATPase, and SR Ca(2+) release channels and the SR Ca(2+) pump. However, the effect of MIA on the KCl-induced increase in [Ca(2+)](i) remained unaffected by treatment with inhibitors of SL Ca(2+) pump ATPase and mitochondrial Ca(2+) uptake. MIA and a decrease in extracellular pH lowered intracellular pH and increased basal [Ca(2+)](i), whereas a decrease in extracellular pH, in contrast to MIA, depressed the KCl-induced increase in [Ca(2+)](i) in cardiomyocytes. These results suggest that NHE may be involved in regulation of [Ca(2+)](i) and that MIA-induced increases in basal [Ca(2+)](i), as well as augmentation of the KCl-induced increase in [Ca(2+)](i), in cardiomyocytes are regulated differentially.     

Electrogenic Na(+)/Ca(2+) exchange. A novel amplification step in squid olfactory transduction

Olfactory receptor neurons (ORNs) from the squid, Lolliguncula brevis, respond to the odors l-glutamate or dopamine with increases in internal Ca(2+) concentrations ([Ca(2+)](i)). To directly asses the effects of increasing [Ca(2+)](i) in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca(2+) from internal stores. We observed an inward current response to caffeine. Monovalent cation replacement of Na(+) from the external bath solution completely and selectively inhibited the caffeine-induced response, and ruled out the possibility of a Ca(2+)-dependent nonselective cation current. The strict dependence on internal Ca(2+) and external Na(+) indicated that the inward current was due to an electrogenic Na(+)/Ca(2+) exchanger. Block of the caffeine-induced current by an inhibitor of Na(+)/Ca(2+) exchange (50-100 microM 2',4'-dichlorobenzamil) and reversibility of the exchanger current, further confirmed its presence. We tested whether Na(+)/Ca(2+) exchange contributed to odor responses by applying the aquatic odor l-glutamate in the presence and absence of 2', 4'-dichlorobenzamil. We found that electrogenic Na(+)/Ca(2+) exchange was responsible for approximately 26% of the total current associated with glutamate-induced odor responses. Although Na(+)/Ca(2+) exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.     

NHE-1 participates in isoproterenol-induced downregulation of SERCA2a and development of cardiac remodeling in rat hearts

Impaired Ca(2+) handling is one of the main characteristics in heart failure patients. Recently, we reported abnormal expressions of Ca(2+)-handling proteins in isoproterenol (ISO)-induced hypertrophied rat hearts. On the other hand, Na(+)/H(+) exchanger (NHE)-1 inhibitor has been demonstrated to exert beneficial effects in ischemic-reperfusion injury and in the development of cardiac remodeling. The aims of the present study are to investigate the role of NHE-1 on Ca(2+) handling and development of cardiac hypertrophy in ISO-infused rats. Male Wistar rats were randomly divided into vehicle [control (CTL)] and ISO groups without or with pretreatment with a selective NHE-1 inhibitor, BIIB-723. ISO infusion for 1 wk significantly increased the ratios of heart to body weight and left ventricle (LV) to body weight and collagen accumulation. All of these increases were antagonized by coadministration with BIIB-723. The ISO-induced significant increase in LV wall thickness was suppressed significantly (P < 0.05) by BIIB-723. ISO-induced decreases in cardiac stroke volume and a total mechanical energy per beat index, systolic pressure-volume area at midrange LV volume, were normalized by BIIB-723. The markedly higher expression of NHE-1 protein in the ISO group than that in CTL group was suppressed (P < 0.05) by BIIB-723. Surprisingly, ISO induced downregulation of the important Ca(2+)-handling protein sarcoplasmic reticulum Ca(2+)-ATPase 2a, the expression of which was also normalized by BIIB-723 without changes in phosphorylated phospholamban (PLB)/PLB expression. We conclude that NHE-1 contributes to ISO-induced abnormal Ca(2+) handling associated with cardiac hypertrophy. Inhibition of NHE-1 ameliorates cardiac Ca(2+)-handling impairment and prevents the development of cardiac dysfunction in ISO-infused rats.     

In situ SR function in postinfarction myocytes.

Previous studies have shown lower systolic intracellular Ca(2+) concentrations ([Ca(2+)](i)) and reduced sarcoplasmic reticulum (SR)-releasable Ca(2+) contents in myocytes isolated from rat hearts 3 wk after moderate myocardial infarction (MI). Ca(2+) entry via L-type Ca(2+) channels was normal, but that via reverse Na(+)/Ca(2+) exchange was depressed in 3-wk MI myocytes. To elucidate mechanisms of reduced SR Ca(2+) contents in MI myocytes, we measured SR Ca(2+) uptake and SR Ca(2+) leak in situ, i.e., in intact cardiac myocytes. For sham and MI myocytes, we first demonstrated that caffeine application to release SR Ca(2+) and inhibit SR Ca(2+) uptake resulted in a 10-fold prolongation of half-time (t(1/2)) of [Ca(2+)](i) transient decline compared with that measured during a normal twitch. These observations indicate that early decline of the [Ca(2+)](i) transient during a twitch in rat myocytes was primarily mediated by SR Ca(2+)-ATPase and that the t(1/2) of [Ca(2+)](i) decline is a measure of SR Ca(2+) uptake in situ. At 5.0 mM extracellular Ca(2+), systolic [Ca(2+)](i) was significantly (P 相似文献   

In dialyzed squid axons oxidative stress inhibits the Na+/Ca2+ exchanger by impairing the Cai2+-regulatory site

The Na(+)/Ca(2+) exchanger, a major mechanism by which cells extrude calcium, is involved in several physiological and physiopathological interactions. In this work we have used the dialyzed squid giant axon to study the effects of two oxidants, SIN-1-buffered peroxynitrite and hydrogen peroxide (H(2)O(2)), on the Na(+)/Ca(2+) exchanger in the absence and presence of MgATP upregulation. The results show that oxidative stress induced by peroxynitrite and hydrogen peroxide inhibits the Na(+)/Ca(2+) exchanger by impairing the intracellular Ca(2+) (Ca(i)(2+))-regulatory sites, leaving unharmed the intracellular Na(+)- and Ca(2+)-transporting sites. This effect is efficiently counteracted by the presence of MgATP and by intracellular alkalinization, conditions that also protect H(i)(+) and (H(i)(+) + Na(i)(+)) inhibition of Ca(i)(2+)-regulatory sites. In addition, 1 mM intracellular EGTA reduces oxidant inhibition. However, once the effects of oxidants are installed they cannot be reversed by either MgATP or EGTA. These results have significant implications regarding the role of the Na(+)/Ca(2+) exchanger in response to pathological conditions leading to tissue ischemia-reperfusion and anoxia/reoxygenation; they concur with a marked reduction in ATP concentration, an increase in oxidant production, and a rise in intracellular Ca(2+) concentration that seems to be the main factor responsible for cell damage.     

Amiloride和Ni2+抑制缺血/复灌引起的大鼠心肌细胞内钙的增加

本文旨在研究Na+/H+交换以及Na+/Ca2 +交换对模拟缺血 /复灌引起的大鼠心肌细胞内游离钙水平变化的调节作用。分别利用模拟缺血液和正常台氏液对大鼠心肌细胞进行缺血 /复灌处理 ,在缺血期间分别应用Na+/H+交换抑制剂阿米洛利 (amiloride)、Na+/Ca2 +交换抑制剂NiCl2 以及无钙液 ,观察它们对细胞内游离Ca2 +浓度变化的影响。利用Zeiss LSM 5 10激光共聚焦显微镜检测、采集细胞内游离Ca2 +的指示剂Fluo 3 AM的荧光信号 ,计算出相对于正常(缺血前 )的相对荧光强度 ,以表示胞内游离Ca2 +浓度的变化。结果显示 ,模拟缺血引起大鼠心肌细胞内游离Ca2 +持续上升 ,缺血前的相对荧光强度值为 10 0 % ,模拟缺血 5min后为 140 3± 13 0 % (P <0 0 5 ) ,复灌 15min后为 142 8±15 5 % (P <0 0 5 )。经 10 0 μmol/Lamiloride、5mmol/LNiCl2 和无钙液分别预处理 ,模拟缺血 5min后的相对荧光强度分别为 10 1 4± 16 3 % (P <0 0 5 )、110 4± 11 1% (P <0 0 5 )和 10 7 1± 10 8(P <0 0 5 ) ;复灌 15min后则分别为 97 8±14 3 % (P <0 0 5 )、10 6 2± 14 5 % (P <0 0 5 )和 10 6 6± 15 7(P <0 0 5 )。另外 ,与对照组细胞相比 ,再灌注期间NiCl2和无钙液处理的细胞钙振荡的产生幅度明显减弱 ,amilorid