SiO_(2) Induces Iron Overload and Ferroptosis in Cardiomyocytes in a Silicosis Mouse Model

Objective The aim of this study was to explore the role and mechanism of ferroptosis in SiO_(2)-induced cardiac injury using a mouse model.Methods Male C57BL/6 mice were intratracheally instilled with SiO_(2) to create a silicosis model.Ferrostatin-1(Fer-1)and deferoxamine(DFO)were used to suppress ferroptosis.Serum biomarkers,oxidative stress markers,histopathology,iron content,and the expression of ferroptosis-related proteins were assessed.Results SiO_(2) altered serum cardiac injury biomarkers,oxidative stress,iron accumulation,and ferroptosis markers in myocardial tissue.Fer-1 and DFO reduced lipid peroxidation and iron overload,and alleviated SiO_(2)-induced mitochondrial damage and myocardial injury.SiO_(2) inhibited Nuclear factor erythroid 2-related factor 2(Nrf2)and its downstream antioxidant genes,while Fer-1 more potently reactivated Nrf2 compared to DFO.Conclusion Iron overload-induced ferroptosis contributes to SiO_(2)-induced cardiac injury.Targeting ferroptosis by reducing iron accumulation or inhibiting lipid peroxidation protects against SiO_(2) cardiotoxicity,potentially via modulation of the Nrf2 pathway.

Intermittent hypoxia attenuates ischemia/reperfusion induced apoptosis in cardiac myocytes via regulating Bcl-2/Bax expression

Intermittent hypoxia has been shown to provide myocardial protection against ishemia/reperfusion-induced injury.Cardiac myocyte loss through apoptosis has been reported in ischemia/reperfusion injury. Our aim was to investigate whether intermittent hypoxia could attenuate ischemia/reperfusion-induced apoptosis in cardiac myocytes and its potential mechanisms. Adult male Sprague-Dawley rats were exposed to hypoxia simulated 5000 m in a hypobaric chamber for 6 h/day, lasting 42 days. Normoxia group rats were kept under normoxic conditions. Isolated perfused hearts from both groups were subjected to 30 min of global ischemia followed by 60 min reperfusion.Incidence of apoptosis in cardiac myocytes was determined by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) and DNA agarose gel electrophoresis. Expressions of apoptosis related proteins,Bax and Bcl-2, in cytosolic and membrane fraction were detected by Western Blotting. After ischemia/reperfusion,enhanced recovery of cardiac function was observed in intermittent hypoxia hearts compared with normoxia group.Ischemia/reperfusion-induced apoptosis, as evidenced by TUNEL-positive nuclei and DNA fragmentation, was significantly reduced in intermittent hypoxia group compared with normoxia group. After ischemia/reperfusion,expression of Bax in both cytosolic and membrane fractions was decreased in intermittent hypoxia hearts compared with normoxia group. Although ischemia/reperfusion did not induce changes in the level of Bcl-2 expression in cytosolic fraction between intermittent hypoxia and normoxia groups, the expression of Bcl-2 in membrane fraction was upregulated in intermittent hypoxia group compared with normoxia group. These results indicated that the cardioprotection of intermittent hypoxia against ischemia/reperfusion injury appears to be in part due to reduce myocardial apoptosis. Intermittent hypoxia attenuated ischemia/reperfusion-induced apoptosis via increasing the ratio of Bcl-2/Bax, especially in membrane fraction.

Protective function of tocilizumab in human cardiac myocytes ischemia reperfusion injury

Objective:To investigate the protective function of tocilizumab in human cardiac myocytes ischemia-reperfusion injury.Methods:The human cardiac myocytes were treated by tocilizumab with different concentrations(1.0 mg/mL,3.0 mg/mL,5.0 mg/mL) for 24 h.then cells were cultured in ischemia environment for 24 h and reperfusion environment for 1 h.The MTT and flow cytometry were used to detect the proliferation and apoptosis of human cardiac myocytes,respectively.The mRNA and protein expressions of Bcl-2 and Bax were measured by qRT-PCR and western blot,respectively.Results:Compared to the negative group,pretreated by tocilizumab could significantly enhance the proliferation viability and suppress apoptosis of human cardiac myocytes after suffering ischemia reperfusion injury(P<0.05).The expression of Bcl-2 in tocilizumab treated group were higher than NC group(P<0.05).while the Bax expression were lower(P<0.05).Conclusions:Tocilizumab could significantly inhibit apoptosis and keep the proliferation viability of human cardiac myocytes after suffering ischemia reperfusion injury.Tocilizumab may obtain a widely application in the protection of ischemia reperfusion injury.

EFFECT OF PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR ACTIVATORS ON TUMOR NECROSIS FACTOR-αEXPRESSION IN NEONATAL RAT CARDIAC MYOCYTES

Objective To investigate the effect of peroxisome proliferator-activated receptor-α(PPARα) and PPARγactivators on tumor necrosis factor-α(TNFα) expression in neonatal rat cardiac myocytes. Methods Primary cultures of cardiac myocytes from 1- to 3-day-old Wistar rats were prepared, and myocytes were ex-posed to lipopolysaccharide (LPS) and varying concentrations of PPARαor PPARγactivator (fenofibrate or pioglitazone).RT-PCR and ELISA were used to measure TNFα, PPARα, and PPARγexpression in cultured cardiac myocytes. Transient tr-ansfection of TNFαpromoter with or without nuclear factor-kappaB (NF-κB) binding site to cardiac myocytes was performed. Results Pretreatment of cardiac myocytes with fenofibrate or pioglitazone inhibited LPS-induced TNFαmRNA and protein expression in a dose-dependent manner. However, no significant changes were observed on PPARαor PPARγmRNA expression when cardiac myocytes were pretreated with fenofibrate or pioglitazone. Proportional suppression of TNFαpromoter activity was observed when myocytes was transiently transfected with whole length of TNFαpromoter (-721/+17) after being stimulated with LPS and fenofibrate or pioglitazone, whereas no change of promoter activity was observed with transfection of TNFαreporter construct in deletion of NF-κB binding site (-182/+17). Conclusions PPARαand PPARγactivators may inhibit cardiac TNFαexpression but not accompanied by change of PPARαor PPARγmRNA expression. Therefore PPARαand PPARγactivators appear to play a role in anti-inflammation. The mechanism may partly be involved in suppression of the NF-κB pathway.

Effect of Cholic Acid on Fetal Cardiac Myocytes in Intrahepatic Choliestasis of Pregnancy

This study examined the effect of cholic acid （CA） on cultured cardiac myoeytes （CMs） from neonatal rats with an attempt to explore the possible mechanism of sudden fetal death in intra- hepatic cholestasis of pregnancy （ICP）. Inverted microscopy was performed to detect the impact of CA on the beating rates of rat CMs. MTT method was used to study the effect of CA on the viability of CMs. CMs cultured in vitro were incubated with 10 ~maol/L Ca2＋-sensitive fluorescence indicator fluo-3/AM. The fluorescence signals of free calcium induced by CA were measured under a laser scanning confocal microscope. The results showed that CA decreased the beating rates of the CMs in a dose-dependent manner. CA could suppress the activities of CMs in a time- and dose-dependent manner. CA increased the concentration of intracellular free calcium in a dose-dependent manner. Our study suggested that CA could inhibit the activity of CMs by causing calcium overload, thereby leading to the sudden fetal death in ICP.

Cardiac disease modeling using induced pluripotent stem cell-derived human cardiomyocytes

Causative mutations and variants associated with cardiac diseases have been found in genes encoding cardiac ion channels, accessory proteins, cytoskeletal components, junctional proteins, and signaling molecules. In most cases the functional evaluation of the genetic alterationhas been carried out by expressing the mutated proteins in in-vitro heterologous systems. While these studies have provided a wealth of functional details that have greatly enhanced the understanding of the pathological mechanisms, it has always been clear that heterologous expression of the mutant protein bears the intrinsic limitation of the lack of a proper intracellular environment and the lack of pathological remodeling. The results obtained from the application of the next generation sequencing technique to patients suffering from cardiac diseases have identified several loci, mostly in non-coding DNA regions, which still await functional analysis. The isolation and culture of human embryonic stem cells has initially provided a constant source of cells from which cardiomyocytes(CMs) can be obtained by differentiation. Furthermore, the possibility to reprogram cellular fate to a pluripotent state, has opened this process to the study of genetic diseases. Thus induced pluripotent stem cells(i PSCs) represent a completely new cellular model that overcomes the limitations of heterologous studies. Importantly, due to the possibility to keep spontaneously beating CMs in culture for several months, during which they show a certain degree of maturation/aging, this approach will also provide a system in which to address the effect of long-term expression of the mutated proteins or any other DNA mutation, in terms of electrophysiological remodeling. Moreover, since i PSC preserve the entire patients' genetic context, the system will help the physicians in identifying the most appropriate pharmacological intervention to correct the functional alteration. This article summarizes the current knowledge of cardiac genetic diseases modelled with i PSC.

Beyond cardiomyocytes:Cellular diversity in the heart's response to exercise

Cardiomyocytes comprise~70%to 85%of the total volume of the adult mammalian heart but only about 25%to 35%of its total number of cells.Advances in single cell and single nuclei RNA sequencing have greatly facilitated investigation into and increased appreciation of the potential functions of non-cardiomyocytes in the heart.While much of this work has focused on the relationship between non-cardiomyocytes,disease,and the heart's response to pathological stress,it will also be important to understand the roles that these cells play in the healthy heart,cardiac homeostasis,and the response to physiological stress such as exercise.The present review summarizes recent research highlighting dynamic changes in non-cardiomyocytes in response to the physiological stress of exercise.Of particular interest are changes in fibrotic pathways,the cardiac vasculature,and immune or inflammatory cells.In many instances,limited data are available about how specific lineages change in response to exercise or whether the changes observed are functionally important,underscoring the need for further research.

In Vitro Study on Mesenchymal Stem Cells:Anti-apoptotic Effects on Cardiac Myocytes

Objectives To investigate the anti-apoptotic effects of mesenchymal stem cells （MSCs） on hypoxic injured cardiac myocytes in vitro. Methods MSCs were isolated from bone marrow of Sprague-Dawley （SD） rats, and cardiac myocytes from neonatal rats. The rat cardiac myocytes were co-cultured with MSCs or MSC-conditioned media in anoxia （95% N2 ±5% CO2） for 72 hours. Cell apoptosis was measured by Hoechst 33258 staining. The expression of Bcl-2 and Bax in cardiac myocytes was tested by Western Blot. Results The apoptotic rate was 51.6% ± 2.4% when cardiac myocytes were cultured in continuous hypoxia and was significantly decreased when cardiac myocytes were cocultured with MSCs or MSC-conditioned media （ 15.1% ± 5.4% and 24. 0% ± 4.2% respectively, P 〈 0. 001 ）. The decreased expression of Bax in the cardiac myocytes was greatly related to the decreasing of apoptosis, but there was no difference in Bcl-2 expression among these groups. Conclusions Co-cultured MSCs showed significant anti-apoptotic effects on cardiac myocytes in continuous hypoxia. The mechanism may be the interact of cell to cell and paracrine of cytokines which effected the expression of Bax in the cardiac myocytes.

Antioxidant Effect of Human Selenium-containing Single-chain Fv in Rat Cardiac Myocytes

Reactive oxygen species（ROS） plays a key role in human heart diseases. Glutathione peroxidase（GPX） functions as an antioxidant as it catalyzes the reduction of hydroperoxide. In order to investigate the antioxidant effect of human selenium-containing single-chain Fv（Se-scFv-B3）, a new mimic of GPX, a model system of hydrogen peroxide（H202）-induced rat cardiac myocyte damage was established. The cardiac myocyte damage was characte- rized in terms of cell viability, lipid peroxidation, cell membrane integrity, and intracellular H202 level. The Se-scFv-B3 significantly reduced H2O2-induced cell damage as shown by the increase of cell viability, the decline of malondialdehyde（MDA） production, lactate dehydrogenase（LDH） release, and intracellular H2O2 level. So Se-scFv-B3 may have a great potential in the treatment of human heart diseases induced by ROS.

Role of NF-κB in protection of EPO pretreatment on neonatal rat cardiac myocytes with hypoxia/reoxygenation injury

Objective：To observe the protective effects of erythropoietin （EPO） pretreatment on cardiac myocyte with hypoxia/reoxygenation （H/R） injury and the role of NF-κBin this effects. Methods：After the H/R model of cardiac myocytes of neonatal rats was established, the cultured cardiac myocytes were divided into 4 groups, including EPO pretreatment group （ EPO 10 U/ml 24 h before H/R）, EPO pretreatment ＋ PDTC group（EPO 10 U/ml and PDTC 5 μg/ml 24 h before H/R）, PDTC group （PDTC 5 μg /ml 24 h before H/R） and eomrolgroup. Before and after the H/R, assay of LDH concentration in the culture medium, the survival rate of the myocytes tested by MTT chromatometry and the apoptosis by flow cytometry were undertaken. Activation of NF-κB was determined by EMSA before and after H/R. Results：EPO pretreatment markedly reduced the LDH concentration in the medium, elevated the survival rate of myocytes and inhibited the apoptosis after H/R. Addition of PDTC during the pretreatment abol- ished the protective effects of EPO pretreatment. NF-κB was markedly activated during EPO pretreatment and PDTCinhibited the activation. However, after H/R, the activity of NF-κB in myocytes with EPO pretreatment was significantly inhibited compared to the other myocytes. Conclusion：NF-κB is significantly activated during EPO pretreatment, but is inhibited after H/R, which is correlated with the protective effects of EPO pretreatment on cardiac myocytes with H/R. This phenomenon can be explained as the negative feedback mechanism of the activation of NF-κB.

Detection of cardiac myosin binding protein-C (cMyBP-C) by a phospho-specific PKD antibody in contracting rat cardiomyocytes

Protein phosphorylation plays an important role in physiological processes, such as muscle contraction. Phospho-specific antibodies have become powerful tools to study these processes. Cardiac myosin binding protein-C (cMyBP-C) is one of the proteins that make up the contractile apparatus of cardiomyocytes. Phosphorylation of cMyBP-C is essential for normal cardiac function, since dephosphorylation of this protein leads to its degradation and has been associated with cardiomyopathy. One of the upstream kinases, which phosphorylate cMyBP-C, is protein kinase D (PKD). While studying the role of PKD in cMyBP-C phosphorylation, we tried to analyze phosphorylation of PKD with a phospho-specific PKD-Ser744/748 antibody. Contrary to the expected 115 kDa, a signal was found for a 150-kDa protein. By MALDI-TOF mass spectrometry, we identified this protein to be cMyBP-C. These data were confirmed by immunostaining using the p-PKD-Ser744/748 antibody, which displayed a striated pattern similar to the one observed for a regular cMyBP-C antibody. To our knowledge there are no antibodies commercially available for phosphorylated cMyBP-C. Thus, the p-PKD-Ser744/748 antibody can accelerate research into the role of cMyBP-C phosphorylation in cardiomyocytes.

Effects of simvastatin on hypertrophy and PTEN expression of rat cardiac myocytes

Objective To study the effects of simvastatin on the hypertrophy of cultured rat cardiac myocytes induced by serum and the role ofphosphatase and tensin homolog deleted on chromosome ten （PTEN） in the signal pathway. Methods Cultured neonatal Sprague- Dawley （SD） rat cardiac myocytes were treated with 15% fetal bovine serum, or without serum, or different consentrations of simvastatin. Image analysis system was used to measure the cardiac myocytes surface area. Protein synthesis of myocytes was measured via [3H]-leucine incorporation method. The expression level of atrial natriuretic peptide （ANP） mRNA in myocytes was determined with reverse transcription polymerase chain reaction （RT-PCR）. The mRNA and protein expression levels of PTEN in cardiac myocytes were investigated with RT-PCR and Western blot respectively. Results At 24 hours, cardiac myocytes surface area was significantly higher in 15% serum group （1611.16± 160.75 lam2） than in serum-free group （538.04±118.60 ±tm2, P〈0.01）. Simvastatin decreased the cell surface area in a concentration dependent manner. The cell surface area in 10-5 and 10-6 mol/L simvastatin groups were 799.84＋ 167.70 ±tm2 and 1076.88± 199.28 um2 respectively, which were both significantly lower than that in 15% fetal bovine serum group （P〈0.01）. Incorporation rate of [3H]-leucine was significantly higher in 15% fetal bovine serum group （2360± 106cpm/well） than that in serum-free group （1305±92 cpm/well, P〈0.01）. Incorporation rate of [3H]-leucine in 10.5 and 10.6 mol/L simvastatin groups were 1707±101 clam/well and 1962±125 cpm/well respectively, which were both lower than that in serum group （P〈0.01）. With the increase of simvastatin concentration, the expression level ofANP mRNA in cardiac myocytes was decreased gradually, which were 0.29±0.03 and 0.40-±0.03 respectively in 10.5 and 10-6 mol/L simvastatin groups, and significantly lower than that in serum group（0.60-±.03, P〈0.01）. Simvastatin increased the expressions of PTEN mRNA and protein in cardiac myocytes in a concentration dependent manner. PTEN mRNA expression level in 10-7, 10-6and 105mol/L simvastatin groups were 0.38±0.03, 0.83±0.04 and 0.85±0.05, respectively, which were all higher than that in 15% fetal bovine serum group （0.29±0.04, P〈0.05）. Similarly, PTEN protein level in 10-7, 10-6 and 10.5 mol/L simvastatin groups （39.25±3.41, 46.35±1.78 and 47.22±2.39 respectively） were also significantly higher than that in 15% fetal bovine serum group （32.21±4.06, P〈0.05）. Conclusion Simvastatin can inhibit the hypertrophy of cultured rat cardiac myoeytes induced by serum, and the increase of expression level of PTEN might be involved in the mechanism （J Geriatr Cardio12010; 7：47-51）.

Effect of post-burn serum on free cytosolic calcium in isolated cardiac myocytes of adult rats

The ohjective of this study was to determine whether the free intracellular calcium concentration ([Ca2+] ) of isolatedcardiac myocytes increased with the stimulation of post-burn serum(PBS) in adult rats. Cardiac myocytes were isolated by collage-nase using Langendorff’s perfusion apparatus, and [Ca2+], was measured using the fluorescent indicator Fain-2. The normal[Ca2+], was 101. 3 ± 21. 3 nmol/L in cardic myocytes. PBS at various postburn home could very significantly increase the[Ca2+]i (P< 0. 01 ) and, 6 h PBS had the strongest effect. However, no significant difference was found between the effects of2 h PBS and 4 h PBS (P >0. 05 ). Both calcium channel antagonist verapamil(30 umol/L) and the inhibitor of ryanodine receptoron sarcoplasmic reticulum procaine (2 mmol/L), very significantly inhibited the action of 6 h PBS, with the inhibition rate of47. 7% and 67. 6% respectively. The inhibiting rate of procaine was significantly greater than that of verapamil (P < 0. 01 ). Theresults suggested that PBS could stimulate the increase of [Ca2+], in isolated cardiac myocytes of adult rats, in which calcium release from intracellular stores might play greater roles. Agents modulating the calcium release from intracellular stores are expectedto have great significance in preventing the organic injuries due to the increases of [ Ca2+]i.

The Effect of Benzyltetrahydropalmatine (BTHP) on Action Potentials and the Two Components of Delayed Rectifying Potassium Currents in Guinea Pig Ventricular Myocytes

The effects of BTHP on Ca 2+ independent action potential and the two components of delayed rectifier potassium currents were studied in guinea pig single ventricular myocytes by using whole cell patch clamp technique. BTHP 30 μmol·L -1 significantly prolonged APD 90 from 143±16 ms to 184±21 ms ( P 【0.01, n=5) without affecting either the RP or APA, and the APD prolonging effects of BTHP were independent of extracellular Ca 2+ . BTHP inhibited both I kr (IC 50 =7 9 μmol·L -1 ) and I ks (IC 50 =22 4 μmol·L -1 ) in a concentration dependent fashion. The results demon strated that BTHP had no obvious selectivity for I kr and I ks .

Characteristics of the Nonselective Cation Current (NSCCs) in Rabbit Left Ventricular Epicardial, Midmyocardial and Endocardial Myocytes

Objectives Recent studies have described regional differences in the electrophysiology and pharmacology of ventric- ular myocardium in canine, feline, rat, guinea pig, and human hearts. This has been shown to be due to a smaller IKs and a lager sodium-calcium exchange current （INa-Ca） and late INa in M region （ deep subepicardial to midmyocardial）. Studies from our laboratory have found a new repolarization current-nonselective cation current （NSCCs） existing in rabbit fight ventricular myocytes. Methods We examined the characteristics of NSCCs in epicardial, M region, and endocardial cells isolated from the rabbit left ventricle with standard microelectrode and whole-cell patch-clamp tech- niques. The permeability to Na^＋ , K^＋ , Li^＋ , Cs^＋ but not to Cl^- indicating that it was a nonselective cation current. Gd^＋3 （0. 1 mmol/1） and La^3＋ （0. 1 retool/1 ） can block the current markedly. Results Further characterization of NSCCs was significantly smaller in M cells than in epicardial and endocardial cells. NSCCs current density was significantly smaller in M cells than in epicardial and endocardial cells. With repolarization to - 80 mV, INa current density was （ -0. 44 ±0. 05） PA/PF in endocardial cells, （ -0. 12 ±0. 05） PA/PF in M cells and （ - 0. 28 ±0. 07） PA/PF in epicardial cells ; and with repolarization to ＋ 30 mV, INa, current density was （ 1.09 ± 0. 29） PA/PF in endocardial cells, （0. 38±0. 09） PA/PF in M cells and （0. 91 ± 0. 32） PA/PF in epicardial cells. Conclusions Transmural dispersion of repolarization was due to the heterogeneity of NSCCs in rabbit left ventricle epicardial, endocardial myocytes and M cells. These findings may advance our understanding of the ionic basis for our understanding of factors contributing to the development of cardiac arrhythmias.

Isolation of Ca^(2+) Tolerant Cardiomyocytes from Aadult Rats for Patch Clamp Studies

Objective: To investigate the factors affecting the viability and Ca 2+ tolerance of isolated rats' cardiac myocytes for patch clamp research. Methods: Hearts were firstly perfused by the Langendorff perfusion apparatus with normal Tyrode's solution, then with Ca 2+-free Tyrode's solution and subsequently with low Ca 2+ enzyme solution containing collagenase 0.1-0.2 g/L. All the solutions were saturated with oxygen and the perfusion temperature was kept at 37 ℃. Finally hearts were washed by Ca 2+-free Tyrode's solution, after which the ventricles were minced into small pieces in KB solution, dispersed and filtered. The isolated myocytes were stored in KB solution at room temperature for 1 h and recovered to normal calcium concentration before patch clamp experiments.Results: When all the factors such as water, enzyme, Ca 2+,pH, and oxygen were well controlled, the well constructed and rod-like cardiac myocytes with a yielding rate of 30%-50% came out.Conclusion: All the factors should be well controlled, which ensured the isolated cells Ca 2+ tolerant and appropriate for patch clamp experiments.

Effects of unsaturated fatty acids on calcium-activated potassium current in gastric myocytes of guinea pigs

AIM: To investigate the effects of exogenous unsaturated fatty acids on calcium-activated potassium current [IK(Ca)] in gastric antral circular myocytes of guinea pigs. METHODS: Gastric myocytes were isolated by collagenase from the antral circular layer of guinea pig stomach. The whole-cell patch clamp technique was used to record /K(Ca) in the isolated single smooth muscle cells with or without different concentrations of arachidonic acid (AA), linoleic acid (LA), and oleic acid (OA). RESULTS: AA at concentrations of 2,5 and 10 μmol/L markedly increased IK(Ca) in a dose-dependent manner. LA at concentrations of 5, 10 and 20 μmol/L also enhanced /K(Ca) in a dose-dependent manner. The increasing potency of AA, LA, and oleic acid (OA) on /K(Ca)at the same concentration (10μmol/L) was in the order of AA>LA>OA. AA (10 μmol/L)-induced increase of Ik(ca) was not blocked by H-7 (10 μmol/L), an inhibitor of protein kinase C (PKC), or indomethacin (10 μmol/L), an inhibitor of the cyclooxygenase pathway, and 17-octadecynoic acid (10 μmol/L), an inhibitor of the cytochrome P450 pathway, but weakened by nordihydroguaiaretic acid (10μmol/L), an inhibitor of the lipoxygenase pathway. CONCLUSION: Unsaturated fatty acids markedly increase Ik(Ca), and the enhancing potencies are related to the number of double bonds in the fatty acid chain. The lipoxygenase pathway of unsaturated fatty acid metabolism is involved in the unsaturated fatty acid-induced increase of IK(Ca) in gastric antral circular myocytes of guinea pigs.

Voltage- and Use-dependent Effect of 7-chlor-benzylte-trahydropalmatine on Sodium Currents in Guinea Pig Ventricular Myocytes

The whole-cell patch-clamp technique was employed to obtain information about the voltage-dependence and kinetics of interaction of 7-chlor-benzylte-trahydropalmatine (7-Cl-BTHP) with cardiac sodium channels. 7-Cl-BTHP (30 mol/L) significantly decreased the peak sodium current (from 7. 8±1. 8 nA to 5. 3±1. 4 nA, P<0. 01, n=5), without producing a shift of the current-voltage curve. It shifted the inactivation curves of sodium current to hyperpolarized potentials, and the V(0.5) was shifted from - (82. 5±2. 5) mV to - (95±2.4) mV (P <0. 05, n=4). 7-Cl-BTHP produced a significant use-dependent effect that was proportional to the duration of the voltage step. In addition, 7-Cl-BTHP slowed the recovery of sodium channel from inactivation, which could explain its use-dependent effects on sodium current. The characteristics of 7-Cl-BTHP blockage suggest that this agent binds preferentially to inactivated sodium channels.

EFFECTS OF ADENOSINE IN SIMULATED ISCHEMIA ANDREPERFUSION ON ISOLATED GUINEA-PIG VENTRICULAR MYOCYTES

Objective To investigate the effects of adenosine （Ado） on myocardiac electrophysiology in simu- lated ischemla and reperfusion in guinea-pig ventricular myocytes. Methods Electrical activity was recorded using standard intracellular microelectrode technique. Right ventricle was superfused with simulated ischemic Tyrode’s so- lution for 15 min, and reperfued with normal Tyrode’s solution for 30 min. Results The results showed Ado had no measurable effects on guinea-pig ventricular myocytes in normal Tyrode’s solution. In the presence of Ado, maximal diastolic potential tended to be more depolarized during ischemia, and action potential （AP） parameters were abbrevi- ated greatly in a concentration-dependent manner. Especially, the concentration of Ado 100 μmol·L-1 had significant effects on AP parameters in ischemic phase [APD30, APD50, and APD90 reduced by （86±8）% versus （65±6）%, （70 ±7）% versus （50±6）%, and （60±6）% versus （42±4）% for control after 15 min, P<0.O5]. During reperfu- sion, AP parameters did not completely return to initial values in presence of Ado. This study illustrated that Ado significantly decreased incidence of arrhythmias induced by ischemia and reperfusion (in presence of Ado 100 μmol· L-1, the incidence of DAD decreased by 17% versus 82% for control during reperfusion). Conclusion Ado has no significant effects on guinea-pig ventricle in normal conditions, abbreviates greatly AP parameters during ischemia with a concentration-dependent manner, and has marked antiarrhythmic effects in ischemia and reperfusion.

Effect of Allocryptopine on Late sodium current of atrial myocytes in spontaneously hypertensive rats

Objective To explore the effect of allocryptopine （All） on the Late sodium current （INa,Late） of atrial myocytes in spontaneously hyper- tensive rats （SHR）. Method The enzyme digestion method was used to separate single atrial myocytes from SHR and Wistar-Kyoto rat （WKY） rats. INa,Late was record by patch-clamp technique and the effect of All on the current was evaluated. Results Comparing with WKY cells, markedly increasing of INa,Late current in SHR myocytes was found from 0.24 ± 0.02 pA/pF of WKY cells to 1.73± 0.04 pA/pF of SHR cells （P 〈 0.01, n = 15）. After treament with 30 μmol/L All; the current densities was reduced to 0.92 ± 0.03 pA/pF. The ratio of INa,Late/INa,peak of WKY and SHR were 0.09% ± 0.01% and 0.71% ± 0.02%, INa, Late/INa,peak of SHR was reduced to 0.37% ± 0.02% by 30 μmol/L All （P 〈 0.01, n = 15）. We also determined the effect of All on the gating mechanism of the INa,Late in the SHR cells. It was found that All decreased the INa,Late by alleviating the inactivation of the channels and increasing the window current of sodium channel. Conclusion Increased INa,Late in SHR atrial myocytes and the prolonged APD were inhibited by All coming from Chinese herb medicine.