Effect of Au NanoStars/collagen matrix material on oxidative stress damage in ventricular myocytes of newborn rats / 军事医学

Objective To verify antioxidation of Au NanoStars/collagen ( AuNSs/Col ) for ventricular myocytes of newborn rats(NRVMs) by in vitro studies.Methods (1)Different concentrations of AuNSs/Col composite materials were created.The optimum concentration of the material was selected by Live /dead staining and Cell Counting Kit-8 (CCK-8) and Col was used for subsequent experiments .( 2 ) NRVMs were randomly divided into Col group , AuNSs/Col group, H2O2-induced Col group, and H2O2-induced AuNSs/Col group.After 6 h treatment, apoptotic cell morphology and early cell apoptosis rate were observed with Annexin Ⅴ-FITC/propidium iodide ( PI)/4′,6-diamidino-2-phenylindole ( DAPI) and the expressions of apoptosis related proteins-B-cell lymphoma-2 ( Bcl-2 ) and Bcl-2 associated x protein ( Bax ) were detected by Western blotting .Results ( 1 ) Both Live/dead and CCK-8 experiments indicated that the AuNSs/Col composite material with 0.1 mg/ml was nontoxicity to NRVMs and could further promote their proliferation .(2) Compared with the uninduced group , the early apoptosis rate of the Col group and the AuNSs /Col group after H2O2 induction was significantly increased , while the Bcl-2/Bax ratio was decreased , indicating that the oxidative stress damage model was established.After H2O2 induction, compared with the Col group , the early apoptosis rate of the AuNSs/Col group was decreased , but the Bcl-2/Bax ratio was increased .Conclusion AuNSs/Col composite material has protective effect on the oxidative damage of cardiomyocytes cultured in vitro.

Multiple voltage-gated sodium channel αsubunits expressed in adult rat ventricular myocytes / 西安交通大学学报(医学版)

Objective To observe the expressions of voltage-gated sodium channel (NaCh)αsubunits in adult rat ventricular myocytes.Methods Single ventricular myocytes were isolated from adult rat heart.Expressions of various αsubunits (Nav1.1,Nav1.2,Nav1.3,Nav1.5,Nav1.6 and Nav1.7)of NaCh in the ventricular myocytes were detected by immunocytochemistry staining.Sodium current was recorded by whole-cell patch clamp method. Results The neuronal subunits Nav1.1,Nav1.6 and Nav1.7 as well as the cardiac subunit Nav1.5 of NaCh were expressed in adult rat ventricular myocytes.Nav1.1,Nav1.5 and Nav1.7 were distributed along the cell membrane of the ventricular myocytes and around the transverse tubule;Nav1.6 was labeled along the cell membrane by lengthways.All these subunits were not colocalized with Cx43 at the intercalated disc.Both transient sodium current (I Na,T )and late sodium current (I Na,L )were recorded from adult rat ventricular myocytes.Conclusion Various neuronal subunits (Nav1.1,Nav1.6 and Nav1.7)as well as cardiac subunit (Nav1.5 )of NaCh were expressed in adult rat ventricular myocytes,which is important for normal function of I Na,T and I Na,L .

Effects of site specific antibody of Na+-Ca2+exchanger on calcium transient in single ventricular myocytes of rats / 中国免疫学杂志

Objective:To investigate the effects of site specific (124 HNFTAGDLGP STIVGSAAFNMF145 ) antibody of Sodium calcium exchanger ( NCX) on calcium transient in single ventricular myocytes of normal adult rats .Methods: Isolated adult rat hearts were perfused using Langendorff method and single ventricular myocytes were then obtained .The ventricular myocytes were incubated with Fuar-2/AM and 2% bovine serum albumin for about 40 min and then,the fluorescence images were recorded when excitation wavelengths were 340 nm and 380 nm using ion imaging system.Fluorescence value F340/F380,length of cell shortening ,time to 90%restore( TR90 ) and calcium sensitivity ( ratio of F340/F380 and cell shortening ) were calculated.Results:The site specific antibody of NCX increased F340/F380 and decreased TR90 in single ventricular myocytes ,but had no more significant effect on calcium sensitivi-ty.Pretreatment with KB-R7943 or Nicardipine could significantly inhibit the TR 90 decrease or F340/F380 increase induced by the anti-body.Pretreating ventricular myocytes with combination of KB-R7943 and Nicardipine ,the antibody had no more significant effects on calcium transient.Conclusion:Site specific ( 124 HNFTAGDLGPSTIVGSAAFNMF145 ) antibody of NCX could increase calcium transient and accelerate the decrease of intracellular calcium during diastole ,which mainly related to its effects of activating L-type Ca2+channel and NCX.

Multiple voltage-gated sodium channel αsubunits expressed in adult rat ventricular myocytes / 西安交通大学学报(医学版)

Objective To observe the expressions of voltage-gated sodium channel (NaCh)αsubunits in adult rat ventricular myocytes.Methods Single ventricular myocytes were isolated from adult rat heart.Expressions of various αsubunits (Nav1.1,Nav1.2,Nav1.3,Nav1.5,Nav1.6 and Nav1.7)of NaCh in the ventricular myocytes were detected by immunocytochemistry staining.Sodium current was recorded by whole-cell patch clamp method. Results The neuronal subunits Nav1.1,Nav1.6 and Nav1.7 as well as the cardiac subunit Nav1.5 of NaCh were expressed in adult rat ventricular myocytes.Nav1.1,Nav1.5 and Nav1.7 were distributed along the cell membrane of the ventricular myocytes and around the transverse tubule;Nav1.6 was labeled along the cell membrane by lengthways.All these subunits were not colocalized with Cx43 at the intercalated disc.Both transient sodium current (I Na,T )and late sodium current (I Na,L )were recorded from adult rat ventricular myocytes.Conclusion Various neuronal subunits (Nav1.1,Nav1.6 and Nav1.7)as well as cardiac subunit (Nav1.5 )of NaCh were expressed in adult rat ventricular myocytes,which is important for normal function of I Na,T and I Na,L .

Src/ROS signal transduction mechanism of low concentration ouabain on contraction in rat cardiocytes / 中国药学杂志

OBJECTIVE: To observe whether low concentration (1×10-8 mol·L-1) of ouabain (OUA) can increase the contractility in rat cardiocytes and investigate the Na/K pump signal transduction pathways related to positive inotropic action following the low concentration of OUA. METHODS: On enzymatic isolation of rats ventricular myocytes, the Na+/K+ pump current (Ip) was by whole-cell patch-clamp, in order to observe the low concentration of OUA on Ip. The contraction of a single myocyte was assessed by a video-based motion edge-detection system. (1) To detect and compare the potentiations of 1×10-8-1×10-3 mol·L-1 OUA on the contractility in rat cardiocytes. (2) The cardiocytes were pre-treated with PP2 (1 μmol·L-1), NAC (100 μmol·L-1), PD98059(50 μmol·L-1) for 5 min, and the effects of the signals transduction inhibitors on the positive inotropic effect of 1×10-8 mol·L-1 OUA was recorded. RESULTS: The 1×10-8-1×10-3 mol·L-1 OUA increased the contractility of rat cardiocytes (P0.05). CONCLUSION: The 1×10-8-1×10-3 mol·L-1 OUA could increase the contraction amplitude of cardiocytes in rats in concentration-dependent manner. Positive inotropic effect of OUA in low concentration is related to Na/K pump signal transduction. Multiple signal pathways regulate the positive inotropic effect of 1×10-8 mol·L-1 of OUA, including the Src/ROS signal pathway.

The effects of H-89 on membrane ion currents in rat ventricular myocytes / 中国药理学通报

Aim To assess the effects of N -[2-p-bromo-cinnamylamino-ethyl]-5-isoquinoline-sulfonamide (H-89), a potentially selective inhibitor of Protein Kinase A (PKA), on cardiac membrane ion channels and transporters, which will further fulfill our understanding of pharmacology of PKA inhibitors.Methods Whole-cell patch clamp was used to investigate the effects of H-89 on cardiac L-type Ca~(2+) current (I_(Ca-L)), Na~+ current (I_(Na)), inward rectifier K~+ current (I_(K1)), transient outward K~+ current (I_(to)) and Na~+-Ca~2+ exchanger current (I_(Na/Ca)) in enzymatic dissociated SD rat ventricular myocytes.Results H-89 at 1～10 μmol·L~(-1) could inhibit I_(Ca-L) , I_(Na) , and Ito in a concentration-relative manner (P <0.05). At low concentra-tion (5 μmol·L~(-1)), H-89 completely inhibited I_(K1) (P <0.05) just as the action of 0.5 mmol·L~(-1) BaCl_2.Further, H-89 at 1～10 μmol·L~(-1) had no significant effect on I_(Na/Ca) (P >0.05).Conclusion The direct or PKA-mediated indirect action maybe involved in the effects of H-89 on ion currents and transporter.

A Carbohydrate Fraction, AIP1, from Artemisia Iwayomogi Reduces the Action Potential Duration by Activation of Rapidly Activating Delayed Rectifier K+ Channels in Rabbit Ventricular Myocytes

We investigated the effects of a hot-water extract of Artemisia iwayomogi, a plant belonging to family Compositae, on cardiac ventricular delayed rectifier K+ current (I(K)) using the patch clamp technique. The carbohydrate fraction AIP1 dose-dependently increased the heart rate with an apparent EC(50) value of 56.1+/-5.5 microgram/ml. Application of AIP1 reduced the action potential duration (APD) in concentration-dependent fashion by activating I(K) without significantly altering the resting membrane potential (IC(50) value of APD(50): 54.80+/-2.24, IC(50) value of APD(90): 57.45+/-3.47 microgram/ml). Based on the results, all experiments were performed with 50 microgram/ml of AIP1. Pre-treatment with the rapidly activating delayed rectifier K+ current (I(Kr)) inhibitor, E-4031 prolonged APD. However, additional application of AIP1 did not reduce APD. The inhibition of slowly activating delayed rectifier K+ current (I(Ks)) by chromanol 293B did not change the effect of AIP1. AIP1 did not significantly affect coronary arterial tone or ion channels, even at the highest concentration of AIP1. In summary, AIP1 reduces APD by activating I(Kr) but not I(Ks). These results suggest that the natural product AIP1 may provide an adjunctive therapy of long QT syndrome.

Cardiomyopeptidin decrease transient outward potassium current of rat ventricular myocytes / 基础医学与临床

Objective To determine the effect of cardiomyopeptidin on transient outward potassium current(I_(to) of rat ventricular myocytes and its action mechanism on the ion channels of myocardium.Methods Single ventricular myocytes of rats were obtained by enzymatic dissociation.The whole-cell patch-clamp recording technique was used to record the change of transient outward potassium current(I_(to) by different dosages of cardiomyopeptidin.Results Cardiomyopeptidin decreased I_(to) in a dose-dependent manner.Cardiomyopeptidin in dose of 10,50,100,250 and(500 mg/L) decreased I_(to %) by 4,13,22,32 and 38 respectively.Cardiomyopeptidin 50 mg/L moved the current density-voltage curve of I_(to) down,but the shape of the curve had no changes.Cardiomyopeptidin 50 mg/L did not change the steady state activation curve of I_(to).Conclusions Cardiomyopeptidin decreases the I_(to) of rat ventricular myocytes,which might be one of the mechanisms of its antiarrhythmic effect.

Opening of ATP-sensitive K+ channel by pinacidil requires serine/threonine phosphorylation in rat ventricular myocytes

The influences of specific protein phosphatase and protein kinase inhibitors on the ATP-sensitive K+ (KATP) channel-opening effect of pinacidil were investigated in single rat ventricular myocytes using patch clamp technique. In cell-attached patches, pinacidil (100 muM) induced the opening of the KATP channel, which was blocked by the pretreatment with H-7 (100 muM) whereas enhanced by the pretreatment with genistein (30 muM) or tyrphostin A23 (10 muM). In inside-out patches, pinacidil (10 muM) activated the KATP channels in the presence of ATP (0.3 mM) or AMP-PNP (0.3 mM) and in a partial rundown state. The effect of pinacidil (10 muM) was not affected by the pretreatment with protein tyrosine phosphatase 1B (PTP1B, 10 mug ml-1), but blocked by the pretreatment of protein phosphatase 2A (PP2A, 1 U ml-1). In addition, pinacidil (10 muM) could not induce the opening of the reactivated KATP channels in the presence of H-7 (100 muM) but enhanced it in the presence of ATP(1 mM) and genistein (30 muM). These results indicate that the KATP channel-opening effect of pinacidil is not mediated via phosphorylation of KATP channel protein or associated protein, although it still requires the phosphorylation of serine/threonine residues as a prerequisite condition.

Effects of Cl- channel blockers on the cardiac ATP-sensitive K+ channel

To explore whether Cl- channel blockers interact with the ATP-sensitive K+ (KATP) channel, I have examined the effect of two common Cl- channel blockers on the KATP channel activity in isolated rat ventricular myocytes using patch clamp techniques. In inside-out patches, 4,4'-diisothio-cyanatostilbene-2,2'-disulfonic acid (DIDS) and niflumic acid applied to bath solution inhibited the KATP channel activity in a concentration-dependent manner with IC50 of 0.24 and 927 muM, respectively. The inhibitory action of DIDS was irreversible whereas that of niflumic acid was reversible. Furthermore, DIDS-induced block was not recovered despite exposure to ATP (1 mM). In cell-attached and inside-out patches, DIDS blocked the pinacidil- or 2,4-dinitrophenol (DNP)-induced KATP channel openings. In contrast, niflumic acid did not block the pinacidil-induced KATP channel openings in inside-out patches, but inhibited it in cell-attached patches. DIDS and niflumic acid produced additional block in the presence of ATP and did not affect current-voltage relationship and channel kinetics. All these results indicate that DIDS among Cl- channel blockers specifically blocks the cardiac KATP channel.

EFFECT OF CHITOSAN ON DELAYED OUTWARD POTASSIUM CURRENT IN SINGLE VENTRICULAR MYOCYTES OF GUINEA PIG / 中国海洋药物

To study the effect of chitosan on delayed outward potassium current(IK) in single ventricular myocytes of guinea pig and investigate its antiarrhythmic mechanism from ion channel view. Patch clamp technique with whole-cell configuration. Holding potential was -40mV,commanding potential was -60- + 70mV ,step pulse +10mV,stimulating frequency 1 Hz,duration 300 ms and stimulating interval 6s. The result showed that Chitosan inhibited IK in a dose -dependent manner. Conclusion :Chitosan can inhibite IK in single ventricular myocytes of guinea pig.