Background

Zebrafish,Brachydaniorerio,haveadistinctembryonicdevelopment.Theembryogoesthroughaprocesscalledepiboly.Epiboly,asdefinedinthesixtheditionofDevelopmentalBIOLOGy(Gilbert2000),isthemovementofepithelialsheets(usuallyoftheectodermcells)thatspreadasaunit,ratherthanindividually,toenclosethedeeperlayersoftheembryo.Epibolyenablesanembryotoprotecttheinnercellsfromextracellularvariations.AccordingtoProfessorRichardFluck,whoworkswithmedakaandisaDr.E.PaulandFrancesH.ReiffProfessorofBiologyatFranklinandMarshallCollege,itishisexperiencethatmedakaembryosaremoresusceptIBLetoexteriorenvironmentalchangepriortocompleteepiboly.Zebrafisharesupposedtobeevenmoresensitivetoenvironmentalchangethanmedaka.

Objective

ThestudyofacidicpHlevelseffectingthedevelopmentofzebrafish.Thisisbeingdonetodetermineifacidrainhasanymajoreffectondevelopingembryos.Rainfall(pH5.6)hasamoreacidicvaluethanthatofnormalwater(pH7orneutral).TheEmbryomedium,whichisusedfordevelopingdechorionatedembryos,aswellastheotherHank"sstandardsolutionscanbeseenhere.AddinglessNaOHbuffersothatitwillbelessthanapH7.2levelwillchangetheembryomedium.Thisissupposedlyonlypertinentifthechorionhasbeenremoved.HopefullythechorionwillnothinderpHinteractions.TheslowadditionofNaOHwillenableforafairrangeofacidictooptimalpHreADInglevels.BytakingpicturesoftheembryospriortoexposuretovariouspHsolutionsandcomparingthesetopicturesofthesameembryosfurtheralongdevelopmentitwillhopefullyshowthatpHlevelhaseneffectonthedevelopmentoftheembryos.

Procedure1)Zebrafishweremarbledthepreviousnightbeforecollectionofembryos-Themarblesinthetankbottompromotespawningwhenthenextdawncomes2)Embryoswereselectedfortwovaryingcellstagesofdevelopment-firstsetwerebetweenthe2-4cellstageofdevelopment-secondsetwasofembryosbetween64-128cellsindevelopment3)pHlevelsusedwere:7.2,6.6,6,5,and4.-pH7.2isthecontrolexperiment

Note:EachofthesepHlevelswasappliedtoeachofthetwocellstagespreviouslymentioned.AnimagewastakenoftheinitialembryostocomparewiththedevelopingembryosinthevariouspHsolutions.Embryoswereobservedovera48hourperiod.

Results

ItwasfoundthatthedifferenceinpHlevelsthatwereuseddidnothavestopdevelopment.TheonlysignificanteffectwasthatthelowerpHlevelscausedthezebrafishtodevelopslower.Thiswasobservedoveraperiodof48hours.

Discussion

ThezebrafishshowednosignsofaltereddevelopmentduetothelevelsofpHusedinthisexperiment.Thismayhaveoccurredbecausetheembryoswerenotdechorionated.Perhapsdechorionatingthemwouldhaveproducedviableresults.AsitstandszebrafishareimpervioustothepHlevelsthatwereusedinthisexperiment.ItalmostcertainthatmuchmoreacidicpHlevelswouldhavedrasticeffectsonthedevelopmentofzebrafishbecausetheproteinlayersoftheembryoswouldbedisturbed.