Fusion-Activated Ca2+ Entry An “Active Zone” of Elevated Ca2+ during the Postfusion Stage of Lamellar Body Exocytosis in Rat Type II Pneumocytes 英文参考文献.docVIP

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Fusion-Activated Ca2+ Entry An “Active Zone” of Elevated Ca2+ during the Postfusion Stage of Lamellar Body Exocytosis in Rat Type II Pneumocytes 英文参考文献.doc

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Fusion-ActivatedCa2EntryAn“ActiveZone”ofElevatedCa2duringthePostfusionStageofLamellarBodyExocytosisinRatTypeIIPneumocytes英文参考文献

Fusion-ActivatedCa2+Entry:An‘‘ActiveZone’’of ElevatedCa duringthePostfusionStageofLamellar BodyExocytosisinRatTypeIIPneumocytes 2+ PikaMiklavc1.,ManfredFrick1,2.,OliverH.Wittekindt1,ThomasHaller2,PaulDietl1* 1InstituteofGeneralPhysiology,UniversityofUlm,Ulm,Germany,2DepartmentofPhysiologyandMedicalPhysics,MedicalUniversityofInnsbruck,Innsbruck,Austria Abstract Background:Ca2+isessentialforvesiclefusionwiththeplasmamembraneinvirtuallyalltypesofregulatedexocytoses. 2+ 2+ However,incontrasttothewell-knowneffectsofahighcytoplasmicCa concentration([Ca ]c)intheprefusionphase,the occurrenceandsignificanceofCa signalsinthepostfusionphasehavenotbeendescribedbefore. 2+ Methodology/Principal Findings: We studied isolated rat alveolar type II cells using previously developed imaging techniques. These cells release pulmonary surfactant, a complex of lipids and proteins, from secretory vesicles (lamellar bodies)inanexceptionallyslow,Ca -andactin-dependentprocess.Measurementsoffusionporeformationbydarkfield 2+ 2+ scattered light intensity decrease or FM 1-43 fluorescence intensity increase were combined with analysis of [Ca ]c by ratiometricFura-2orFluo-4fluorescencemeasurements.Wefoundthatthemajorityofsinglelamellarbodyfusionevents 2+ werefollowedbyatransient(t1/2 ofdecay=3.2s)riseoflocalized[Ca ]c originatingatthesiteoflamellarbodyfusion. 2+ [Ca ] increasefollowedwithadelayof 0.2–0.5s(method-dependent)andinthemajorityofcasesthissignalpropagated , c throughoutthecell(at,10mm/s).RemovalofCa from,oradditionofNi2+totheextracellularsolution,stronglyinhibited 2+ 2+ 2+ these[Ca ]ctransients,whereasCa storedepletionwiththapsigarginhadnoeffect.Actin-GFPfluorescencearoundfused 2+ 2+ LBsincreasedseveralsecondsaftertheriseof[Ca ]c.Botheffectswerereducedbythenon-specificCa channelblocker SKF96365. Conclusions/Significance:Fusion-activatedCa2+entry(FACE)isanewmechanismthatleadsto[Ca2+]ctransientsatthesite ofvesiclefusion.Substantialevidencefromthisand