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1、SodiumChannelStructure,Function,GatingandInvolvementinDiseaseDavidR.Marks,M.Sc.AnOverviewSodiumChannelStructure-CurrenttheoryandTypesofNa+ChannelsSodiumChannelFunction-Currenttheoryofinactivation-Modulation-Pharmacology-ActivationAnOverviewContdArticle2Na+ChannelGatingArticle1-Na+ChannelsandNeurodeg
2、enerativeDiseaseArticle3Na+channelmutationandphysiologySodiumChannels-StructureComposedof,-1and-2subunits,butthelarge-subunitscarriesmostofthefunctionalproperties4repeatedmotifs,eachwith6transmembranedomainsAlllinkedtogetherContainavoltage“sensor”/ligandbindingdomain(methodofactivation)Thehydrophobi
3、cS4segment(voltage“sensor”)isfoundinallvoltagegatedNa+channelsandisabsentinligandgatedNa+channelsSelectivityfilter(shellofhydration)InactivationgateCartoonrepresentationofthe“typical”voltage-activatedsodiumchannelTypesOfNa+ChannelsVoltagegatedChangesinmembranepolarityopenthechannelLigandgated(nicoti
4、nicacetylcholinereceptor)Ligandbindingalterschannel/receptorconformationandopenstheporeMechanicallygated(stretchreceptor)PhysicaltorsionordeformationopensthechannelporeSodiumChannels-FunctionPlayacentralroleinthetransmissionofactionpotentialsalonganerveCanbeindifferentfunctionalstates(3) -Arestingst
5、atewhenitcanrespondtoadepolarizingvoltagechanges-Activated,whenitallowsflowofNa+ionsthroughthe-Inactivated,whensubjectedtoa“suprathreshold”potential,thechannelwillnotopenThetheoryisthattheinactivationgate“swings”shut,turningoffthechannelPleaseKeepInMindThestructureoftheNa+channelisnot100%solved,henc
6、ea“workingmodel”isdrawnbasedonbiophysical,pharmacological,physiologicalandmolecularassaysZhao(2004)writes“Themechanismofopeningandclosingisunknown,butstructuralstudiessuggest”Na+ChannelModulationPhosphorylationsodiumchannelfunctionismodulatedbyserine/threonineandtyrosinekinasesaswellastyrosinephosph
7、atases(Yuet al,Science1997)Mutationalteredaminoacidsequence/structurecanchangethebiophysicalpropertiesoftheNa+channelPharmacologyblockNa+channeltoreducetheconductanceProteolysis-(cleavage)Proteasesmaycleavespecificresiduesorsequencesthatinactivateachannel,orsignificantlyalterthebiophysicalproperties
8、WhyNa+Channels/ModulationAreImportantNeuronaldepolarization,ActionPotentialNeuronalExcitabilityCardiacExcitabilityMuscleExcitabilityThebasisofneuronal/cardiac/muscularfunctionreliesonthepropagationofactionpotentials,downaxons,sarcolemma,myocardium,aswellasrequiringsynaptictransmission.Differentialex
9、citabilityalterstheelectricalconduction/transmissionpropertiesofthe“circuit”Na+ChannelBlockers/PharmacologicalAgentsTetrodotoxin(TTX)AmioderoneLidocaineProcainamideMexilitineKetamineMany,manyothersSomeNa+ChannelsOutsideTheNervousSystemNaf“FunnyCurrent”inpacemakercellsoftheheart(SAnode/ectopicpacemak
10、ers)Navinthemyocardium,sarcolemma,andT-tubulesandmotorendplateNa+ChannelActivationChangeintransmembranepotentialresultsinaconformationchangeintheNa+channelThefourS4segmentalphahelicestranslocate,thusleadingtotheopeningofthechannelporeTheenergyoftheconformationalchangeinthechannelduringactivationisme
11、diatedbythereductioninoverallentropyofthesystem.Thevoltagesensorisahighlychargedsequenceofaminoacidsthat“aligns”itselfaccordingtotheelectricalfieldpresentAchangeintransmembranepotentialcreatesunfavorableelectrodynamicinteractionforthevoltagesensor,henceaconformationalshiftlowerstheenergyofthesystema
12、ndcreatesmorefavorableconditionsPatchClamping/TransfectionTransfection1.Kv1.3cDNAinPlasmid2.Lipofectaminecomplexing3.AddtoDishes4.Patch28-48hrsafterTransition:AGeneralOverviewofArticlesBeforeDiscussionFromBasicstructure/functionrelationshipstoagatingmechanismThegatingofabacterialNa+channelandapplica
13、tionofNa+channelactivationandbiophysicalpropertiesArticle1AgatinghingeinNa+channels:amolecularswitchforelectricalsignalingProposedconformationalshiftofA-helixcausedbysubstitutionofProlineforG219Prolinesinalphahelicesafterthefirstturn(4thresidue)causeakinkinthehelix.Thiskinkiscausedbyprolinebeingunab
14、letocompletetheH-bondingchainofthehelixandstericorrotamereffectsthatkeepprolinefromadaptingthepreferedhelicalgeometryConservedglycineIntheS6domainNa+ChannelGatingCurrenttheoryholdsthatachangeintransmembranepotential“flips”theconformationofthevoltagesensor,therebyopeningthechannelporeAmutation,G219P,
15、glycine219changedtoprolinealterstheconformationoftheS6domainThemutantchannelnowfavorsastatemuchlikethe“open”stateofawild-typechannelNOTE:thesebacterialNa+channelsarehomotetramersofidenticalsubunitsMutationaltersthebiophysicalpropertiesofthechannelTheG219Pmutantactivatessignificantlyearlier(activates
16、atmuchmorenegativevoltages)thanthewild-typeV:VoltageatwhichofchannelspresentareintheopenstateComparabletoKminthatitisameasureoftheabilityofachanneltoactivateOthermutationstotheNa+channelDonotexertassignificanteffectsintheactivation(V)InfluenceofhybridNa+channelsubunitsongatingandbiophysicalpropertie
17、sArticle2-Na+ChannelsAndNeurodegenerativeDiseaseOverviewMultipleSclerosis(MS)displaysaremission-relapsecourse.Someaxonsareabletomaintainminimalconductionvelocity,whileothersdegeneratecompletely.Definition:Experimentalautoimmuneencephalomytis(EAE)animalmodelofMSMScandisplayremission-relapsingcourse.T
18、hisisbelievedtobetheresultoftheexpressionoftwodistinctisoformsofvoltage-gatedNa+channelsNaV1.2/1.6areexpressedoverlongdistances(10m)B-amyloidarepeptiesassociatedwithneurodegenerativediseases,andcanaccumulateinfibrillaraggregatesWhatisImportantAboutThisArticleNav1.6iscolocalizedwithaNa/CaexchangerNav
19、1.2isNOTcolocalizedwithB-amyloidproteinsNav1.2helprestoreconductionindemyelinatedaxonsNav1.6isseenindegeneratingaxonsAnincreaseinNaV1.6yieldsanIncreaseinNa/Caexchangers,elevatingintracellularCa2+toharmfullevelsArticle3-Na+ChannelsandtheConductionSystemoftheHeartLongQTsyndromediseasewheretheentirecyc
20、leofexcitation-contractioncouplingofthemyocardiumisprolongedPatienthadG-Asubstitutionatcodon1763oftheNav1.5channelgene,whichchangedavaline(GTG)toamethionine(ATG)ThismutationproducedapersistentlyactiveandfastrecoveringNa+channelMutantwasINSENSITIVEtolidocaineArticle3Authorsgeneratedasimilarmutantbysi
21、te-directedmutagenesisExaminedthemutantinaheterologousexpressionsystemtoobtainbiophysicalandotherpropertiesTheNav1.5V1763MmutantisSensitivetoTTX,butresistanttolidocaineTTXeliminateslidocaine-insensitivecurrentWhy this is important:Otherthantraumaticcardiacarrest,arrhythmiasdegenerateintoventricularf
22、ibrillationorventriculartachycardias.“circusmovement”wherebytissuebecomes“hyper-excitable”ExtensionandApplicationofNa+ChannelPropertiesandFunctionRelatingtoArticle3AdvancedCardiacLifeSupport(ACLS)TargetsNa+ChannelsExtensively“PleaseShockShockShock,EverybodyShock,AndLetsMakePatientsBetter”Thepurposeo
23、fdefibrillationofventriculararrhythmiasistoapplyacontrolledelectricalshocktotheheart,whichleadstodepolarizationoftheentireelectricalconductionsystemoftheheart.Whentheheartrepolarizes,thenormalelectricalconductionmayrestoreitselfDepolarizationtheoreticallyinactivatesallvoltage-gatedNa+channels,andall
24、owsVoltage-gatedpotassiumchannelstoactivate,andhelphyperpolarizethemembrane+40mv-70mvAfter AdministrationOf ProcainamideV FIB/V TACHAfterphosphorylation/phosphatecleavageUse-dependent block of sodium channels.Blocks potassium channels.Blocks alpha-adrenergic receptors.Blocks muscarinic receptors.Use
25、dtoattempttoterminatepersistentreentrantarrhythmiasReducesautomaticityofALLpacemakers(boththeSAnodeandANYtissuecapableofgeneratingapacemakerpotential)SlowsDownConductionofdepolarizationinALLtissuesoftheheartanddecreasescardiacexcitabilityThis is your last resort.Givingthisdrugmaystopthearrhythmia,bu
26、tmakeitalmostimpossibleforthehearttospreadimpulsesafterSummaryFortheLectureNa+channelsarecomprisedsubunits,theAlphaof4repeatingmotifs,eachmotifwith6transmembranedomainsTherearevoltage,ligand,andmechanically-gatedNa+channelsNa+channelsareinvolvedinthedepolarizationofexcitablemembranesNa+channelshavem
27、ultiplemodalitiesofmodulation,whichcanalterneuronal/membraneexcitabilityNa+channelsarethetargetofamultitudeofpharmacologicalagentsSummaryNa+channelsAreinvolvedintheremission-relapseofMSNa+channelgatingcanbesignificantlyaffectedbymodulation(phosphorylation,mutation,proteolyticcleavage)MutationinNav1.5isimplicatedinLongQTsyndrome,generatingpersistentandslowinactivatingsodiumcurrent
