Seite - 95 - in Advances in Neuroimmunology

BrainSci. 2016,6, 25 explore thesevariouspossibilities iswarranted, includingcombinationsofdifferent intensities, times, andtypesofstresswithchronicalcoholchallenges, circadianrhythms,andgeneticbackground. Toassessthepossiblecontributionofothercomponentstoneuroimmuneactivationpathwaysthat result in theeffectsobservedhere,TLR4mRNAwasalsoassessedafter restraintstress.Asignificant increase inTLR4mRNA,was observedonly at 4 h in cortical tissue following stress, returned to control levelby8h, andremainedat this level through48hafter thestress (Figure2D).This fairly tight time-response relative to the othermRNAswas one of the reasons for focusing on 29 h of withdrawal(4hpoststressrelativetonon-stressedalcoholwithdrawnrats)asthetargetforcomparison. Aconsiderationhere is thatprior assessments [38] focusedmostlyonalcoholwithdrawal-derived mRNAdatafromthe24thhourofwithdrawalalthoughtheyshowedthatmRNAscanremainelevated for longerperiods, thus it seemsunlikely thatmRNAlevelswouldbemeaningfullydifferentacross these twotimepointswithin thepresentstudy.Regardless, theseresultsagreewithBlandinoetal. [18] whofoundnochange incorticalTLR4mRNAat their2h timepointpost footshockorLPStreatment. It isnotable that stressorWCEelevatedTLR4mRNAonly in thecortex. Thereasons for this specific effectonTLR4areunknown,butsuggestsdifferentialneuroimmuneregulationandpossibly lower thresholds foractivationacrossbrainregions. TheTLR4receptorcomplex isaprominentdriver in neuroimmuneprocesses ingeneralandinalcoholism(e.g., see [48])andanimalmodels inparticular (e.g., [38,49,50]). In fact, theTLR4receptormayplayarole inapositive feedback loopthatamplifies the intensity of overall neuroimmune activation in alcoholism [51]. Such reduced thresholds for neuroimmuneactivationmayberepresentedmostprofoundly followingendotoxinwhereactivation progresses toneurodegeneration in thesubstantianigrawithcorrespondingbehavioraldeficits [52]. WhileWCE alone generally elevated cortical cytokines (and see [38]) and hippocampal CCL2 and IL-1β, WCE or stress alone did not consistently do so for some mRNAs in some regions (e.g., thehippocampusandhypothalamus,Figures4and5).The limitedTNFα response in theseregions contrastswiththeeffectsofpain-associatedstresssuchasfootshockreportedbyBlandinoetal. [18]and thussupportstheideathatthetypeofstressmaybeimportantincytokineinduction.Again,thepattern of results suggests thatuniquemechanismsareoperatingacrossbrainregionsanditwould likelybe productive to furtherexamineneuroimmunemRNAsmoregenerallyonaregionbyregionbasisand thusspeaktotherelativelyunderstudiedyetcritical issueofhowneuroimmunechangesacrossregions ornetworkscouldproduceneuropathology. Thepotentialdifferential inductionofanti-inflammatory cytokinessuchas IL-10couldalsobe informative. These futurestudiesshouldelucidatehowstress induction of some cytokine mRNAs, but not others, contributes to the profile of neuroimmune activation inmodelsofalcoholism[3–5,9]. One potentially important focus in identifying mechanisms of cytokine regulation in this experimental context relates to thecorticotropin-releasingfactor (CRF)system.Thedatahereinshow that theCRF1RantagonistCP154,526was inactiveagainst stress inductionofneuroimmunemRNAs. Thiseffectprovidesaninterestingcomparisonwithpreviouswork[38] that focusedontheeffectof this drugoncytokinemRNAselevatedbyWCE.Themechanismsthatexplainhowthedrugcomestoexert aneffectononechallengeandnot theotherareunknown,butdifferential engagementofadaptive mechanismsmightbeonepossibility. That is, thedrugmayaffectarecruitedprocessuniquetorats experiencingWCE.This interestingpossibility is reminiscentof theworkofKoobandcolleagueswho notedthatCRFreceptorantagonisteffectsweregenerallynotmanifestunless ratsweredependenton alcohol (e.g., [53]). It is important tonote that theCP154,526studyhereinwas limited in its scopeand couldbeexpanded in future studies toexaminerelatedquestions. Forexample, thedrugcouldbe employed inexamining theeffectoncytokinemRNAs in thecontextof combinedstressandWCE which isarguablyaveryrelevantexperience insomealcoholabusers. While thecurrent researchcorroborates thedemonstrationbyWhitmanetal. [38] thatcytokine mRNAsare increased in thecortex24hafterWCEandextendsour inquiry intootherbrainsitesand to theeffectsof stress, thestudiesdonotaddress thispotentially interestingcombinatorialeffectsof the twochallenges.Whatourresultsdoshowis thateffectsof thesechallengesareeachthemselves 95