心脏电生理技术基础(膜片钳).ppt

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1、CELLULARCARDIACELECTROPHYSIOLOGICALTECHNIQUESNORBERTJOST,PhDElectricalmodelofthemembraneStandardintracellularmicroelectrodetechniqueVoltageclamptechniquePatchclamptechniqueG=1/ROhm’slawIonchannelmodelCurrentclampVoltageclampIntracellularmicroelectrodetechniqueRe<

2、KClRe~10-40MOhmThesetupOrganbathd:stimulatingelectrodee:microelectroder:referentelectrodeP:preparationcomputerA/DingerlőamplifierDetectedsignalPder100ms50mV0mVdrug60min20-60min60min0mV20mV100msAPARPAPD50APD9090%50%VmaxWash-outPre-incubationTwomicroelectrodevoltageclampvoltagecommandholdingpotent

3、ialtestpotentialThemacroscopicsodiumcurrentThevoltage-clampcircuitvoltagecommandamplifierCurrentmeasurevoltagemeasurefollowupamplifierPatch-clamp:thespecialcaseofthevoltageclampCellPatch-clamp:thespecialcaseofthevoltageclamp(1)Suckasmallpieceofmembraneontothetipofaglassmicropipette(~1µmindiamete

4、r)Cell(2)“Gigaohm-seal”R>1GOhmPatch-clamp:thespecialcaseofthevoltageclampCell(3)Sensevoltagehere,insidetheelectrode,andusevoltageclamptokeepitconstant.Patch-clamp:thespecialcaseofthevoltageclampclosedopenCell++Patch-clamp:thespecialcaseofthevoltageclamp(3)Sensevoltagehere,insidetheelectrode,andu

5、sevoltageclamptokeepitconstant.closedopenopenCell(3)Turnontheaimedpotentialtheinsidepartofthepipetteandkeepitconstantlybyapplyingthevoltageclamptechnique.Patch-clamp:thespecialcaseofthevoltageclampvoltagecommand10msecPropertiesofindividualvoltage-dependentsodiumchannelsIndividualchannelsareeithe

6、ropenorclosed(nopartialopenings)Propertiesofindividualvoltage-dependentsodiumchannelsIndividualchannelsareeitheropenorclosed(nopartialopenings)Eachchannelopeningisonlyabriefeventcomparedtothetotaldurationofthewholecellvoltage-dependentsodiumcurrent.ThemacroscopicsodiumcurrentPropertiesofindividu

7、alvoltage-dependentsodiumchannelsIndividualchannelsareeitheropenorclosed(nopartialopenings)Eachchannelopeningisonlyabriefeventcomparedtothetotaldurationofthewholecellvoltage-dependentsodiumcurrent.Channelopeningandclosingisv