The Properties Of Sodium-calcium Exchanger Current Expressed By NCX1.1

Background and purpose:The sodium–calcium exchanger (NCX) is an integral protein that belongsto the superfamily of plasma membrane exchangers. NCX activity in themammalian heart has been reported to be enhanced by β-adrenergicstimulation.The bi-directional and electrogenic property of NCX raises aquestion about whether NCX is involved in firing of heart beat. We studied theelectrophysiological characteristics of NCX current （INCX） of sinoatrialnode(SAN) of newborn rat and INCXexpressed by NCX1.1,to investigate thesignal transduction pathway of β-adrenergic receptor(β-AR) regulating INCX,aswell as the role of INCXin pacemaker current.The First Part The electrogenic property of INCXof newbornrat’s SANMethods:The sino-atrial node cells of newborn wistar rats were enzymaticallydisassociated by the method of enzyme digestion with differentialattachment．Whole cell-patch clamp recording technique was used to recordINCXof SAN.The I-V curve of the INCXresponse to different frequencystimulation and forskolin and the β-AR agonist (isoprenalinehydrochloride)was obtained solely or after treatment with H89or pertussistoxin(PTX) were all used to evaluate current characteristics of INCXof SAN.Results:The spindle-shaped cells constituted the greatest proportion of the culturedSAN cells.The frequency of spontaneous beating of spindle-shaped cell is147.9±9.9beats/min and the spontaneous action potential was recorded inspindle-shaped cells. Under different frequency stimulation(0.25Hz、0.5Hz、1.0Hz、2.0Hz), we observed no obvious effect on INCXof SAN cells.Treatmentwith isoprenaline hydrochloride increased INCXof SAN cells by36.7%(P＜0.05).Treatment with different concentration of isoprenaline hydrochlorideincreased IN C Xof SAN cells accordingly (0.1μmol、1μmol、10μmol) by 36.7%、111%、169%. H89attenuated the rise of INCXinduced byisoprenaline hydrochloride (P＜0．05)．Forskolin and PTX increased therise of INCXby41.27%and12.8%(P＜0．05).Conclusions:1. There is no clear effect of different frequency stimulation on INCXof SANcells2. NCX activity of the SAN cells of newborn rats could be enhanced byisoprenaline hydrochloride stimulation3. There may be obvious effect of isoprenaline hydrochloride and this effectincreases with the increase of the concentration of isoprenalinehydrochloride.4. INCXmight be regulated by β-AR through the pathway of Gs/Gi-cAMP-PKA.The Second Part The electrogenic property of INCXexpressed byNCX1.1Mthods:The NCX expression system was made using Immunomagnetic Beadspositive mark method. The I-V curve of the IN C Xresponse to differentextracellular Na+concentration and forskolin and the β-AR agonist(isoprenaline hydrochloride) was obtained solely or after treatment with H89orpertussis toxin(PTX) were all used to evaluate current characteristics ofINCXof SAN.Reults:Treatment with high concentration of extracellular Na+decreased theinward INCXexpressed by NCX1.1by28.75%(P＜0．05)．In contrasttreatment with low concentration of extracellular Na+increased the inwardIN C Xexpressed by NCX1.1by33.91%(P＜0．05). Treatment withisoprenaline hydrochloride increased the inward INCXof SAN cells by27.67% (P＜0.05).H89attenuated the rise of the inward INCXinduced byisoprenaline hydrochloride (P＜0．05)．Forskolin and PTX increasedthe rise of the inward INCXby81.25%and26.32%(P＜0．05).Conclusion:1. High concentration of extracellular Na+could restrain the forward model ofNCX activity．In contrast, low concentration of extracellular Na+stimulate theforward model of NCX activity of NCX1.1.2.The forward model of NCX activity of NCX1.1could be enhanced byisoprenaline hydrochloride stimulation.3.INCXmight be regulated by β-AR through the pathway of Gs/Gi-cAMP-PKA.