Production of dissociated sensory neuron cultures and

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1、PROTOCOLProductionofdissociatedsensoryneuronculturesandconsiderationsfortheiruseinstudyingneuronalfunctionandplasticitySachaAMalin,BrianMDavis&DerekCMolliverDepartmentofMedicine,DivisionofGastroenterology,HepatologyandNutrition,UniversityofPittsburgh,3350TerraceStreet,Pittsburgh,Pennsylvania1

2、5261,USA.CorrespondenceshouldbeaddressedtoS.A.M.(sam64@pitt.edu).Publishedonline15February2007;doi:10.1038/nprot.2006.461sDissociatedprimarysensoryneuronsarecommonlyusedtostudygrowthfactordependentcellsurvival,axonoutgrowth,differentiationandbasicmechanismsofsensoryphysiologyandpain.Spinalort

3、rigeminalsensoryneuronscanbecollectedfromembryos,neonatesoradults,treatedwithenzymesthatdegradetheextracellularmatrix,trituratedandgrownindefinedmediawithorwithoutgrowthfactorsandadditionalanimalsera.Productionofculturescantakeaslittleas2.5h.Cellscanbeusedalmostnatureprotocol/immediatelyormain

4、tainedforaslongas1month.Easeofproductionandtheabilitytocontrolgrowthconditionsmakesensorymoc.neuroncultureapowerfulmodelsystemforstudyingbasicneurobiologyofcentralandperipheralnervoussystems.erutan.INTRODUCTIONw24,25wDissociatedrodentsensoryneuronsareamongthemostwidelyspecificsignalingpathways

5、.Adultandneonatalsensorycul-w//:usedneuronalcelltypesforprimaryculture.Sensoryneuronsaretureshavebeenroutinelyusedforphysiologicalstudiesexploringptlocatedinspinalganglia(alsoknownasdorsalrootganglia,ormolecularmechanismsforthedetectionofphysical,chemicalandthDRGs)orincranialsensorygangliaass

6、ociatedwithcranialnervesthermalstimuli.Inaddition,isolatedafferentshavebeenusedtopuV,VII,VIII,IXandX.Thegreatmajorityoftheseneuronsarestudyprocessesimportantforbothcentralandperipheralnervousorderivedfromtheneuralcrest,althoughsomecomefromplacodalsystemfunction,suchasregeneration26,27andthero

7、leofaxonalG132830gtissue(e.g.,thenodoseganglionfoundoncranialnerveX).translationofproteinsinmodulatingsynapticfunction.niSensoryneuronscanbeobtainedfromDRGs,locatedinthehislintervertebralforamenalongthespinalcolumn(seeFig.1),andStrengthsandwe