Medchemexpress/3-Methyladenine(Synonyms: 3-MA)/HY-19312/10mM*1mL in DMSO

3-MethyladenineisaselectivePI3KinhibitorwithIC50of25μMand60μMforVps34andPI3Kγ,respectively,anditcaninhibitallPtdIns3Ksat10mM.

IC50:25μM(Vps34),60μM(PI3Kγ)

3-methyladenineshowssomelimitedVps34preferenceinvitro,withanIC50of25µMforVps34ascomparedwith60µMforPtdIns3Kγ,and3-methyladenine(10mM)caninhibitallPtdIns3Ks[1].3-methyladenine(3-MA,5mM)suppressesautophagyinHeLacellsunderbothglucose-freeconditionsandnormalconditions.3-methyladenineinducescaspase-dependentcelldeathinHeLacells,andthedeathoccursindependentlyoftheinhibitionofautophagy.3-MA(1mM)significantlyshortensthedurationofnocodazole-induced-prometaphasearrest[2].Alargenumberofneuronsfromthe3-MAgroupareintheearlystageofapoptosis,displayingcellshrinkage,nuclearandchromatincondensationandaggregationattheedgeofthenucleus[3].

3-methyladenine-treatmentsignificantlyaggravatesneurologicalsymptomsandapoptoticcelldeathafterSAH[3].

• [1].MillerS,etal.FindingafittingshoeforCinderella:searchingforanautophagyinhibitor.Autophagy.2010Aug;6(6):805-807.

  [2].HouH,etal.Inhibitorsofphosphatidylinositol3"-kinasespromotemitoticcelldeathinHeLacells.PLoSOne.2012;7(4):e35665.

  [3].JingCH,etal.Autophagyactivationisassociatedwithneuroprotectionagainstapoptosisviaamitochondrialpathwayinaratmodelofsubarachnoidhemorrhage.Neuroscience.2012Jun28;213:144-153.

CellviABIlityisdeterminedbyatrypanblueexclusionassay.Cellsareculturedinthemediumwith3-Methyladenine.BothadherentandfloatingcellsarecollectedandsUSPendedinphosphatebufferedsaline(PBS,pH7.4)atafinaldensityof1-2×106/mL.Anequalvolumeof0.4%trypanbluesolution(w/v,inPBS)isaddedtothecellsuspensionandmixedthoroughly.Afterincubationatroomtemperaturefor3min,cellcountingisperformedusingahemacytometer.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

3-Methyladenineisdissolvedinnormalsaline.

Animalsarepositionedinastereotacticframeandtreatedwith5-μLintracerebralventricularinjectionsofautophagyinducerRAP(20 pmol),inhibitor3-Methyladenine(3-MA,400 nmol)orvehicle20 minbeforeSAHonset.Intracerebralventricularinjectionsaremadeintotheright lateralventricle atcoordinatesof0.8 mmanteriortoposteriorbregma,1.5 mmmidtolateral,3.8 mmdorsaltoventral.Aftertheinjection,thesyringeisleftinthesitefor30 sbeforeremoval,andtheburrholeisfilledwathbonewax.Drugdosesareselectedonthebasisofpreviouspublishedstudiesandpilotexperimentsinourlaboratory.3-MAisdissolvedinnormalsalinebyheatingthesolutionto60-70°Cimmediatelybeforetreatment.RAPisfirstdissolvedinethanolandthendilutedwithnormalsaline(finalethanolconcentration<2%).=""mce=""has=""not=""independently=""confirmed=""the=""accuracy=""of=""these=""methods.=""they=""are=""for=""reference="">

• [1].MillerS,etal.FindingafittingshoeforCinderella:searchingforanautophagyinhibitor.Autophagy.2010Aug;6(6):805-807.

  [2].HouH,etal.Inhibitorsofphosphatidylinositol3"-kinasespromotemitoticcelldeathinHeLacells.PLoSOne.2012;7(4):e35665.

  [3].JingCH,etal.Autophagyactivationisassociatedwithneuroprotectionagainstapoptosisviaamitochondrialpathwayinaratmodelofsubarachnoidhemorrhage.Neuroscience.2012Jun28;213:144-153.

149.15

C₆H₇N₅

5142-23-4

RoomtemperatureincontinentalUS;mayvaryelsewhere

DMSO:≥1.5mg/mL;DMSO:6.8mg/mL(Needultrasonic);H2O:8.82mg/mL(Needultrasonic)

*"<1 mg/ml"="" means="" slightly="" soluble="" or="" insoluble.="" "≥"="" means="" soluble,="" but="" saturation="">