Cholinergic neuronotrophic factors: VI. Age-dependent requirements by chick embryo ciliary ganglionic neurons☆

[摘要] Duringdevelopment,ciliaryganglionicneuronsbecomepostmitoticandextendneuritesinapparentindependenceofthepresenceoftheirfutureintraocularinnervationtargets.Afterreachingtheirperipheralinnervationterritory,however,theseneuronsbecometargetdependentandabouthalfofthemdie.Wehavepreviouslyreportedthatchickembryointraoculartargettissuescontainaciliaryneuronotrophicfactor(CNTF),whichcanbeextractedandpartiallypurifiedinasolubleformandwhichensuresnear-totalsurvivalof8-daychickembryociliaryganglionicneuronsinmonolayercultures.InthisstudywehavedissociatedandculturedciliarygangliafromembryonicDay(ED)5through14,andexamineddependenceandresponsivenessoftheirneuronstoexogenouslyaddedCNTF.Twocellclasses(darkandbright)couldbedistinguishedbyphasemicroscopyanddifferentiallycountedincelldissociatesfromED7–14,butnotinED5–6ones.Darkcellnumberperganglionincreasedfrom6000to78,000overthisdevelopmentaltimeperiod.Incontrast,brightcells(putativeneurons)declinedfromamaximumofabout10,000to6000,suggestingacorrelationwiththeexpectedneuronalcelldeathinvivo.DissociatedcellsfromED5–14gangliawereseededonapolyornithinesubstratumcoatedwithneuritepromotingfactor,culturedfor24hrwithorwithoutaddedCNTF,andnumericallyexaminedforsurvivalandneuriticdevelopment.CulturesfromED7–14gangliashowedtwocellcategories:(i)flatnonneuronalelementsdramaticallyincreasedinnumberwithganglionicage(therebycorrelatingwiththeincreasingnumberofdarkcellsinthedissociates)and(ii)large,brightcells(oftendisplayingneuriteoutgrowth)decreasedinnumberinparallelwithbrightcellnumberinthedissociate.ThesurvivaloftheseneuronalelementswasstrictlydependentonexogenouslyaddedCNTFbetweenED7and10,butbecameprogressivelyindependentwitholderages.ED14neurons(fullycapableofsurvivingfor24hrwithoutaddedCNTF)continuedtorequireCNTFforneuriteextension,thusdisplayingretainedsensitivitytothisfactor.AlthoughtheED5–6culturescontainedwell-recognizableflatcells,thedominantcategorycomprisedcellswithvariablemorphology,practicallyallofwhichexhibitedneurite-likeprocesses.Boththesurvivalandneuriteextensionofthesecells,whichwetentativelyinterpretasimmatureneuronswereindependentofthepresenceofaddedCNTF.

[学科分类] 生物科学（综合）