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.

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 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.

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.

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.

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

苄基四氢巴马汀为四氢巴马汀的衍化物，本文采用标准膜片钳全细胞记录技术，研究了苄基四氢巴马汀对豚鼠心室肌单细胞动作电位及延迟整流钾通道（Ｉｋ）两种成分（Ｉｋｓ和Ｉｋｒ）的选择性作用。结果表明：（１）３０μｍｏｌ·Ｌ－１苄基四氢巴马汀可显著延长无钙细胞外液条件下动作电位的ＡＰＤ９０（从１４３±１６ｍｓ到１８４±２１ｍｓ，Ｐ＜００１，ｎ＝５），而对ＲＰ，ＡＰＡ无明显影响，其对ＡＰＤ９０的延长作用不依赖于胞外钙。（２）苄基四氢巴马汀对延迟整流钾通道两种成分均有不同程度的阻滞作用，其阻滞Ｉｋｒ的ＩＣ５０（半数抑制浓度）为７９μｍｏｌ·Ｌ－１，阻滞Ｉｋｓ的ＩＣ５０为２２４μｍｏｌ·Ｌ－１。

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.

Characterization and Differentiation into Adipocytes and Myocytes of Porcine Bone Marrow Mesenchymal Stem Cells

Bone marrow mesenchymal stem cells （BMSCs） could differentiate into various cell types including adipocytes and myocytes, which had important scientific significance not only in the field of tissue regeneration, but also in the field of agricultural science. In an attempt to exhibit the characterization and differentiation into adipocytes and myocytes of porcine BMSCs, we isolated and purified porcine BMSCs by red blood cell lysis method and percoll gradient centrifugation. The purified cells presented a stretched fibroblast-like phenotype when adhered to the culture plate. The results of flow cytometry analysis and immunofluorescence staining demonstrated that the isolated cells were positive for mesenchymal surface markers CD29, CD44 and negative for hematopoietic markers CD45 and the adhesion molecules CD31. Cells were induced to differentiate into adipocytes with adipogenic medium containing insulin, dexamethasone, oleate and octanoate. Oil Red O staining demonstrated that the porcine BMSCs successfully differentiated to adipocytes. Moreover, the findings of real-time PCR and Western blotting indicated that the induced cells expressed adipogenic marker genes （PPAR-y, C/EBP-c~, perilipin, aP2） mRNA or proteins （PPAR-3,, perilipin, aP2）. On the other hand, porcine BMSCs were induced into myoctyes with myogenic medium supplemented with 5-azacytidine, basic fibroblast growth factor, chick embryo extract and horse serum. Morphological observation by hochest 33342 staining showed that the induced cells presented as multi-nucleus muscular tube structure. And myogenic marker genes （Myf5, desmin） mRNA or proteins （MyfS, MyoD, myogenin, desmin） were found in the induced cells. In addition, the results of immunofluorescence staining revealed that myogenic marker （Myf5, MyoD, myogenin, desmin, S-MyHC） proteins was positive in the induced cells. Above all, these results suggested that the isolated porcine BMSCs were not only consistent with the characterization of mesenchymal stem cells, but also exhibited the multipotential capacity to form adipocytes and myocytes, which provided the basis to investigate the regulation mechanism involved in the selective differentiation of porcine BMSCs.

Antioxidant Effect of Selenium-containing Glutathione S-Transferase in Rat Cardiomyocytes

As one of the most important antioxidant enzymes, glutathione peroxidase（GPX） protects cells and tissues from oxidative damage, and plays an important role in cardiovascular and cerebrovascular injuries induced by oxida- tive stress. The antioxidant effect of selenium-containing glutathione S-transferase（Se-GST）, a mimic of GPX was investigated on rat cardiomyocytes. To explore the protection function of Se-GST in hydrogen peroxide（H202） chal- lenged rat cardiomyocytes, we examined malondialdehyde（MDA）, lactate dehydrogenase（LDH）, superoxide dismu- tase（SOD） and cell apoptosis. The results demonstrate exposure of rat cardiomyocytes to H202 for 6 and 12 h induced the significant increases of MDA, LDH and apoptosis rate of cardiomyocytes, but pretreatment of rat cardiomyocytes with Se-GST at 0.0005 or 0.001 unit/mL prevents oxidative stress induced by H202 with the decreases of cell apopto- sis. All the results hint Se-GST has antioxidant activity for oxidative stress challenged rat cardiomyocvtes.

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.

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 Ik(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 IK(Ca)in a dose-dependent manner. The increasing potency of AA, LA, and oleic acid (OA) on Ik(Ca) at the same concentration(10 μmol/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(10 μmol/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.

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.

Effects ofβ_2-Adrenergic Antagonist on Cytosolic Ca^(2+) in Ventricular Myocytes from Infarcted Rat Heart

Objectives To investigate the effects of β2-adrenergic antagonist on cytosolic Ca^2 ＋ （[Ca^2＋ ]i） in ventricular myocytes from infarcted rat heart. Methods A ligature was placed around left anterior descending coronary artery of rat hearts. Rats in the control group were sham-operated. Cardiomyocytes were dissociated at two, four, eight weeks after myocardial infarction （MI） and [Ca^2＋]i was measured via fura-2 fluorescence. The response of cardiomyocytes to isoproterenol in presence or absence of betal-adrenergic antagonist atenolol, beta2-adrenergic antagonist ICI118, 551 or non-selective β1, 2- adrenergic antagonists propranolol was examined. Results The followings were found that ICI 118, 551 had no significant effects on the rise of [Ca^2＋]i induced by isoproterenol in normal ventricular myocytes （P 〉 0.05）, ICI118, 551 only significantly attenuated the rise of [Ca^2＋]i induced by isoproterenol at four weeks and eight weeks after MI （24.5%±5.7% vs 57.8% ± 13.2%, P〈 0.01; 12.2%±7.9% vs 44.6%±11.3%, P〈 0.01）. Atenolol had suppressive effects only in the control group and the post-MI group of two weeks （P 〈 0.05）, and propranolol had suppressive effects in the control and all the three post-MI groups （P 〈 0.01）. Conclusions Beta2-adrenergic antagonist ICI118, 551 may exert negative effects on Ca^2＋ overload initiated by sympathetic stimulation after MI.

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-κB in 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 control group. 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 abolished the protective effects of EPO pretreatment. NF-κB was markedly activated during EPO pretreatment and PDTC inhibited 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.

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

目的将在心脏的 myocytes 在信号小径由浆液和在染色体十上删除的磷酸酶和 tensin 相当或相同的事物(PTEN ) 的角色导致了的有教养的老鼠的肥大上学习 simvastatin 的效果。方法有教养的新生的 Sprague-Dawley (SD ) 老鼠心脏的 myocytes 与 15% 胎儿的牛的浆液被对待，或没有 simvastatin 的浆液，或不同 consentrations。图象分析系统被用来测量心脏的 myocytes 表面区域。myocytes 的蛋白质合成被测量经由[3H ] 白氨酸加入方法。atrial natriuretic 的表示水平在 myocytes 的肽(ANP ) mRNA 与反向的抄写聚合酶链反应(RT-PCR ) 被决定。在心脏的 myocytes 的 PTEN 的 mRNA 和蛋白质表示层次分别地与 RT-PCR 和西方的污点被调查。在 24 个小时的结果，心脏的 myocytes 表面区域在 15% 浆液组是显著地更高的(1611.

The Effect of Levobunolol Hydrochloride on the Calcium andPotassium Channels in Isolated Ventricular Myocytes of Guinea Pig

The effects of levobunolol hydrochlorid (Bun) on the type L calciumchannel currents (Ica) and delayed rectifier potassium channel currents (Ik) in isolated ventricular myocytes of guinea pig were studied by using patch clamp wholecell recording techniques. The results were showed that: 1) Bun caused a dosedependent decrease in Ica and a dose-dependent increase in Ik of the ventricular myocytes.The threshold concentrations of Bun for Ica and Ik were 10-8 mol/L and10-7 mol/L respectively. The maximum effective concentration of Bun for both Ica and Ik was 3 × 10-5 mol/L, and half-maximal concentration was 3 × 10-6 mol/L;2 ) Ik was blocked by 2× 10-6mol/L tetraethylammonium (TEA). A concentration of 3 × 10-6 mol/L Bun showed a decreasing effect on the Ica as revealed by the current-voltage relationship curve, i. e., Bun caused an elevation of the curve; 3)When Ica was blocked by 2 × 10-6 mol/L Isoptin (Verapamil), at a concentrationof 3 × 106- mol/L Bun showed an increasing effect on Ik and the effect could be blocked by TEA. The above-mentioned results indicated that Bun had an inhibito-ry effect on Ica and a fascilitatory effect on Ik The results suggested that themolecular mechanisms of antihypertensive, heart rate slowing and β-receptorblocking effects of Bun might be due to decrease of Ica and increase of Ik.

Protein Kinase C Enhances the Swelling-Induced Chloride Current in Human Atrial Myocytes

Swelling-activated chloride currents（ICl.swell） are thought to play a role in several physiologic and pathophysiologic processes and thus represent a target for therapeutic approaches. However, the mechanism of ICl.swell regulation remains unclear. In this study, we used the whole-cell patch-clamp technique to examine the role of protein kinase C（PKC） in the regulation of ICl.swell in human atrial myocytes. Atrial myocytes were isolated from the right atrial appendages of patients undergoing coronary artery bypass and enzymatically dissociated. ICl.swell was evoked in hypotonic solution and recorded using the whole-cell patch-clamp technique. The PKC agonist phorbol dibutyrate（PDBu） enhanced ICl.swellin a concentration-dependent manner, which was reversed in isotonic solution and by a chloride current inhibitor, 9-anthracenecarboxylicacid. Furthermore, the PKC inhibitor bis-indolylmaleimide attenuated the effect and 4α-PDBu, an inactive PDBu analog, had no effect on ICl.swell. These results, obtained using the whole-cell patch-clamp technique, demonstrate the ability of PKC to activate ICl,swell in human atrial myocytes. This observation was consistent with a previous study using a single-channel patch-clamp technique, but differed from some findings in other species.

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.