Malnutrition and brain development: An analysis of the effects of inadequate diet during different stages of life in rat

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Neuroscience and Biobehavioral Reviews

j o ur n a l ho m e p ag e :w w w . e l s e v i e r . c o m / l o c a t e / n e u b i o r e v

Review

Malnutrition and brain development: An analysis of the effects of inadequate diet during different stages of life in rat

Meryem Alamy, Wail A. Bengelloun

^∗

FacultyofScience,MohammedV-AgdalUniversity,Rabat,Morocco

a r t i c l e i n f o

Articlehistory:

Received5October2011

Receivedinrevisedform19March2012 Accepted25March2012

Keywords:

Proteinmalnutrition Undernutrition Bodyweight

Neuroanatomicalaspects Behavioraleffects Rat

a b s t r a c t

Proteinmalnutritionorundernutritioncanresultinabnormaldevelopmentofthebrain.Depending ontype,ageatonsetandduration,differentstructuralandfunctionaldeﬁcitscanbeobserved.Inthe presentreview,wediscusstheneuroanatomical,behavioral,neurochemicalandoxidativestatuschanges associatedwithproteinmalnutritionorundernutritionatdifferentagesduringprenatalandimmediately postnatalperiodsaswellasinadultrat.Analysisofalldatasuggeststhatproteinmalnutritionaswellas undernutritioninducedimpairedlearningandretentionwhenimposedduringtheimmediatelypostnatal periodandinadulthood,whereashyperactivityincludingincreasedimpulsivenessandgreaterreactivity toaversivestimulioccurredwhenmalnutritionorundernutritionwasimposedeitherpreorpostnatally.

Thisgeneralstateofhyperreactivitymaybelinkedessentiallytoanalterationindopaminergicsystem.

Hence,thepresentreviewshowsthatinspiteoftheattentiondevotedintheliteraturetoprenataleffects, cognitivedeﬁcitsaremoreseriousfollowingmalnutritionorundernutritionafterbirth.Wethusclearly establishaspecialvulnerabilitytomalnutritionafterweaninginrats.

Contents

1. Introduction... 1464

2. Effectofmalnutritionorundernutritiononbodyweight(Tables1and2). . ... 1465

2.1. Prenatalmalnutritionorundernutrition ... 1465

2.2. Postnatalmalnutritionorundernutrition. . ... 1465

3. Neuroanatomicalanddevelopmentaleffectsofmalnutritionorundernutrition(Tables3and4) ... 1466

3.1. Prenatalmalnutritionorundernutrition . . ... 1467

3.2. Postnatalmalnutritionorundernutrition... 1470

4. Behavioraleffectsofmalnutritionorundernutrition(Tables5and6)... 1471

5. Neurochemicaleffectsofmalnutritionorundernutrition(Tables7and8)... 1473

5.1. Effectofmalnutritionondopaminergicsystem... 1473

5.1.1. Prenatalmalnutritionorundernutrition... 1473

5.1.2. Postnatalmalnutritionorundernutrition... 1474

5.2. Effectonserotoninergicsystem ... 1474

5.2.1. Prenatalmalnutritionorundernutrition... 1474

5.2.2. Postnatalmalnutritionorundernutrition... 1475

6. Oxidativestatusaftermalnutritionorundernutrition(Tables9and10)... 1475

7. Discussion... 1476

Acknowledgments... 1477

References... 1477

∗Correspondingauthor.Tel.:+212661166277;fax:+212537671401.

E-mailaddress:[email protected](W.A.Bengelloun).

http://dx.doi.org/10.1016/j.neubiorev.2012.03.009

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1. Introduction

Nutritionplaysacrucialroleinthematurationandfunctional developmentof thecentralnervous system.Theeffects ofmal- nutritionor undernutrition havebeenrepeatedly demonstrated over thepast40 years in humans(Galleret al.,1997; Prakash, 2006;Rosenberg,2007;ScrimshawandGordon,1968;Waltonand Allen,2011;Winnick,1969)and inexperimentalstudiesinrats (Dhopeshwarkar,1983;Dobbing,1987,1968;Galleretal.,1997;

Morganeetal.,1992,1978;Tonkissetal.,1993).Differenttypesof malnutritionmayaffectmaturationofthebrainandthedevelop- mentofcognitivefunction,resultinginbehavioralabnormalities and disturbances in learning and memory. In thelast decades, improvementofinvestigative techniquessuchas immunohisto- chemicallabeling,dosageofproteins,enzymes,receptors,mRNA levelsandreactiveoxygenenzymesassessingoxidativestress,has enabledabetterunderstandingoftheeffectsofmalnutritionand undernutritiononfunctionaldevelopmentofthebrain.

The term“malnutrition” implies that one or more essential nutrimentismissingorpresentbutininappropriateproportions.

Althoughthelackofmicronutrientssuchasvitaminsorminerals mayaffectbrainmaturation,proteinsappeartobethemostcritical (Morganeetal.,2002).“Undernutrition”isaformofmalnutrition whereallnutrimentsrequiredbythespeciesareavailableinthe dietbutininsufﬁcientamounts(Morganeetal.,2002).

Malnutritionisoneofthemostcommonhealthconditionsfound inhumanchildren(Galleretal.,1997).Clinically,severeinfantmal- nutritionisgenerallyclassifiedasmarasmus(insufficientcaloric intake),kwashiorkor(proteindeficiency)orasacombinationofthe twosyndromes(protein–caloriemalnutrition)(Bengelloun,1990).

Inbothmarasmusandkwashiorkor,deﬁcienciesinmicronutrients, includingvitamins,minerals,andtraceelements,oftenaccompany theenergyorproteindeﬁcits.ThecurrentfamineinSomaliaisan exampleofinadequatenutritionwhoseeffectsmaylastforsome timetocome.

Intheliterature,severalattemptshavebeenmadetostudythe long-termeffectsofmalnutritiononintellectualperformanceand learningabilityinhumans.Themoststrikinglong-termeffectsof earlymalnutritionwerebehavioralandcognitivewithimpaired ﬁnemotor skills,lower IQscoresand attentiondeﬁcitdisorder (Galleretal.,1997,1990,1987).

Theeffectsofmalnutritiononcognitivedevelopmenthavebeen studiedinbothhumansandanimals,althoughtheanimalstudies beganearlierthanthehumanstudiesandaremoreabundantinthe literature(Lausetal.,2011).Researchinvolvinghumansconcerns mainlychildren indevelopingcountries;thehighestprevalence ofdisadvantagedchildrenisinsub-SaharanAfricaandSouthAsia (Rosenberg,2007).Experimentalanimal studiesareparticularly usefultounderstandinghowmalnutritionaffectsbraindevelop- ment.Earlyexperimentalstudiesfocusedontheeffectsofdeﬁcient dietonbodyandbrainweight,(Bengelloun,1990;Fernandezetal., 1985;Winnick,1969).Othersinvestigatedtheeffectsofmalnutri- tiononcognitivefunctions(Bengelloun,1990;Tonkissetal.,1994, 1993,1991,1990a,b).Later,worksaddressedimpairmentsofneu- rotransmittersystems(Alamyetal.,2005;Almeidaetal.,1996a) andchanges inproteinphosphorylationand oxidativestatus in brain(Bonattoetal.,2005;Feolietal.,2006).

Nutritionalinadequacyremainsoneofthemajornongenetic factorsaffectingdevelopmentofbrain.Itcontinuestobeofinterest duetothewidespreadincidenceoffoetalandinfantilenutritional deﬁciencies and thegrowing body of evidence that theeffects ofnutritionalinsultonthedevelopingbrainarelong-lastingand lead to permanent deﬁcits in learning and behavior (Morgane etal.,2002).Maternalmalnutritioninduceddeleteriouseffectson brainstructures(Kingetal.,2004,2002;Soto-Moyanoetal.,1999) resultinginbraindysfunctionanddecreasedbrainweightinpups,

particularlyintheneonatalandearlypostnatalperiods(Joshietal., 2003;Souzaetal.,2011,2008).Moreover,inexperimentalstud- iesinwhichfemaleratsunderwentrehabilitationforoneortwo generationsonanadequatelevelofdietaryproteinfollowingahis- toryofintergenerationalmalnutrition,norecoveryoccurredafter twogenerationsofdietaryrehabilitationformeasuresofmaternal behaviorincludingactiveandpassivenursing,puporientedbehav- ior,timespentinthenestandtimeincontactwiththeyoung(Galler andPropert,1981).

Prenatalproteinmalnutritioncausedalterationsinthedevelop- mentaltimecourseofdendategyrus(Debassioetal.,1996,1994), inthemorphologyofhippocampalcells(Díaz-Cintraetal.,1991) andinthenumberanddistributionofneurotransmitterreceptors (Almeidaetal.,1996a).Inadditiontosuchalterations,itisalso well-establishedthatproteindeprivationaffectscathecolaminer- gic,serotoninergic(Wigginsetal.,1984),glutamatergic(Rottaetal., 2003)and gabaergic(Steigeret al.,2003,2002)neurotransmis- sion.Proteinorprotein–caloriemalnutritionimposedearlyinlife (postnatal)producedalterationsinthebrain,includingneurophys- iological(CastroandRudy,1987;Fukudaetal.,2002;Lukoyanov andAndrade,2000),morphological (Bedi,1987; Debassioetal., 1996,1994;Díaz-Cintra etal.,1991)andneurochemical(Alamy etal., 2005;Dietrichet al.,2004;Leahyet al.,1978)aspectsof developingbrainfunction.

Dietaryproteinisanimportantsourceofessentialaminoacids thatcanserveasintracellularantioxidant.Therefore,itsrestriction mayleadtoanincreaseinoxidativedamagebydiminishingantiox- idantdefencesof tissue (Bonattoet al.,2005).Freeradicals are physiologicallygeneratedbybiologicalsystemsduringmetabolism andcaninduceoxidativedamageinorganicmolecules.Themain enzymaticdefencesagainstfreeradicalsaresuperoxidedismutase (SOD)andcatalase(CAT).Thiscoupledenzymaticactivityisable toreducetheionsuperoxidetoH₂O(HalliwellandGutterdridge, 2006).Whenfreeradicalsincrease,cellulardamageoccurs.Because ofitshighrateofoxidativemetabolicactivity,highcontentofphos- pholipidsandmoderatelevelsofantioxidant,thebrainisespecially vulnerabletofreeradicals(Garciaetal.,2005).

Boththeageatwhichmalnutritionoccursanditsdurationare criticalfactorsin determiningitseffects onthebrain (Morgane etal.,2002).Thereisevidencethattheearlierthedietaryinsult,the moresevereandpermanentareitseffects(Morganeetal.,1992).

Manystudiesdescribedtheeffectsofprenatalorperinatalmalnu- tritiononthedevelopmentofthebrain.Itshouldbenotedthatthe gestationalperiodcomprisesmostoftheprocessofneurogenesis, whereas inthepostnatal period myelinization,dendritic prolif- erationandsynaptogenesistakeplace.Malnutritionduringthese phasescanthusproducelong-lastingorpermanentalterationsin differentneurotransmitterssystems,whichmayberesponsiblefor behavioralabnormalitiesaswellaschangesinreactivitytodiffer- entpharmacologicaltreatments(Almeidaetal.,1996a;Morgane etal.,1992).Early(gestational)restrictioninterferedwithcelldivi- sionandinducedadeﬁcitinthenumberofcellsinallorganseven afteradequaterefeeding,whereaslatemalnutritionresultedina decreaseintheprotein/DNAratio(cellsize)withrecoveryafter refeeding(Winnick,1969).

Atweaning,proteinmalnutritioninduceddepressedweightand rateofgrowth.Theratioofbrainweight/bodyweightwasgreater inmalnourishedthan inwell-nourishedrat (Bengelloun, 1990).

Althoughit waspossibletorehabilitatemorphological parame- ters,thereisalastingeffectinbehavioraldisturbances.Moreover, undernutritionatweaningbyreductionofamountofnutriments inducedbehavioralandbiochemicaleffects(Alamyetal.,2005).

Inadult,contrarytoearlierreports(Morganeetal.,1992),there wasalsoevidencethatprolongedproteindeprivationresultedin markedstructuralalterationinthebrainalongsidethebehavioral effects(LukoyanovandAndrade,2000).

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Therearedifferencesbetweenspeciesduringbrainmaturation withrespecttoneurogenesis,neuronmigration,synapseformation andtheelaborationofneuronalcircuits.Theentiredevelopmen- tal period appears to be critical for the final outcome of the mature brain since it contains multiple overlapping and sepa- ratecritical periods(Morgane et al., 2002).According tothese authors,aclearunderstandingoftheeffectsofmalnutritionmust include considerationof thetime ofmalnutrition inrelationto thescheduleof brain developmentin thespecies. It shouldbe stressedthatpostnatalbraindevelopmentismarkedlyinfluenced byprenatalnutritionalstatus,andthatgestationaldevelopmentis greatlyinfluencedbythepregestationalnutritionalstatusofthe mother.

Malnutrition or undernutrition appears toaffect a sequence ofeventsthatextendsthroughout braindevelopmentincluding cellbirth(neurogenesis and gliogenesis),cellmigration, andall aspects ofdifferentiation, includingmyelinization and synapto- genesis. Inadequate diet may exert its effects on any of these developmentalphases.Duringembryonicandfetaldevelopments, neurons and glia are generated at speciﬁc time periods from embryonic cells. Afterbirth, youngneuronsmigratealong spe- ciﬁcpathwaysinrelationwithotherdevelopmentaleventsinthe brain.If cellproliferation or migration is delayedneuronsmay notarriveintime toreceivetheirappropriateinputs. Thismay resultinalteredfunctionalorganizationofthebrainareasinvolved.

Thedevelopmentof neurotransmitterfunction alsooccursdur- ingtheperiodofdifferentiation,throughsynthesisofregulatory enzymes,neurotransmittersandspeciﬁcreceptors(Galleretal., 1997).

Inconsideringthebodyofdatacitedinthisreview,theeffectsof malnutritionorundernutritionmaypersistlaterinlifebecauseof changestheyinducedinthemechanismsofneurotransmitterand receptorsynthesisandsynapsesgrowth.

The present review focused on experimental studies in rat, to determine and brieﬂy describe the effects of malnutrition andundernutritioninducedatdifferentageperiods.In arecent review, limited to the cognitive deﬁcits of malnutrition in humans and rodents, Laus et al. (2011) stressed that protein andprotein–caloriemalnutritionduringtheprenatalperiodand early in life can lead to learning and memory impairments.

Thehypothesis addressed in thepresent review wasthat malnutrition induces behavioral and cognitive deﬁcits which are age-dependentandthat theseeffects aremoreimportantwhen malnutrition occurs during the postnatal period and in adulthood.

Analysisoftherelevantneuroanatomical,behavioralandneu- rochemical studies in rat no doubt contributes to a better understandingoftheeffectsofmalnutritioninhumans.Salientdata areexhaustivelypresentedinTables1through10.

2. Effectofmalnutritionorundernutritiononbodyweight (Tables1and2)

Thereareseveralmethodstoproducequalitativemalnutrition throughselectivenutrimentdeprivationsuchaslowproteindiets (proteinmalnutrition).Undernutrition(quantitativemalnutrition orcalorierestriction),maybeinducedthroughinadequateamounts ofnutrimentsresultingfromlargelitters,removalofthepupsfrom thedamsorreducingrations.

Typicalexperimentalparadigmsinvolvedeithermalnutritionor undernutritionatdifferentperiodsinlife.Irrespectivetothetech- niqueused,malnutritionorundernutritioninducedbodyweight deﬁcits.Theseverityofthiseffectwasdifferentasafunctionof thetype(malnutritionorundernutrition)andtheageatonsetof restriction.

2.1. Prenatalmalnutritionorundernutrition

In the case of prenatal malnutrition, the effects of protein malnutrition were studied (Debassio et al., 1994; Galler and Tonkiss, 1991; Tonkiss et al., 1994, 1991) by providing nulli- parousfemalerats,5weekspriortomating,accesstooneofthe twoisocaloricdiets,adequateprotein(25%casein)orlowprotein (6%casein)content.Thenutritionaltreatmentcontinuedthrough- out pregnancyuntilparturition. Atbirth,all littersofboth diet groupswereculledtoeightmalepupsandfosteredtodamsfed 25% casein diet that had given birth nomore than 24h previ- ously.

Prenatal severe protein malnutrition produced a significant decreaseinanimals’bodyweights(Hernandesetal.,2005).Pre- natally malnourished rats weighed significantly less than the well-nourishedratsatbirth,andinspiteofanormallactation,this continuedtobethecaseforupto27days.Byday70,bodyweights of controlandexperimentalgroups werenolongerstatistically distinguishable(Tonkissetal.,1994).Prenatalproteinmalnutri- tionledtobodyweightdeficitsrangingfrom5%to21%(Almeida etal.,1996b,c;Chenetal.,1997;Kehoeetal.,2001;Tonkissetal., 1994).

Prenatalproteinrestriction(5%versus20%caseinindiet)initiated atconception,andmaintainedduringtheﬁrst2weeksofratges- tation(Gressensetal.,1997)resultedinsmallerfoetalbodyweight andreducedbraincorticalthicknessonembryonicdayE15atthe endofthemalnutritionperiod.However,refeedingthemalnour- isheddamsduringthelastweekofpregnancyallowedacomplete normalizationofgrowthparametersinoffspring.

Prenatalundernutritionwasinducedbyrestrictingfoodintake ofdamstohalfofthequantityingestedbycontroldams(Jahnke andBedi,2007).Thisundernutritioninduceda73%ofbodyweight lossinpupsat21daysofage,whendamswererestrictedduring pregnancyandlactation(Jahnkeand Bedi,2007).Uterine-artery ligationonembryonicday17inducedadecreaseinbrainweight andan8%decreaseinbodyweightatbirth(Olivieretal.,2007).

Perinatalundernutritionwasproducedbyreducingproteinand fattyacidsintakeina multideﬁcientdietinfemalesthroughout pregnancyandlactation,aswellasintheiroffspringafterweaning frompostnataldays22to70.Inmalnourishedoffspringbothbody andbrainweightswereseverelyaffected(Souzaetal.,2008).This malnutritionproducedan83%bodyweightlossanda25%decrease inbrainweightatday70.In therehabilitatedgroupprovideda controldietafterweaningfrompostnataldays22to70,thebody andbrainmassdeﬁcitswereentirelycompensated(Souzaetal., 2008).

2.2. Postnatalmalnutritionorundernutrition

Proteinmalnutritionimposedearlyinlife(atbirth)wasdescribed byFukudaetal.(2002).Duringthelactationperiodandafterwean- inguntil49daysofage,damsandpupswereplacedincageswith proteindeﬁcientdietcontaining6%casein,whereasthenormal proteindietcontained16%protein.Thismalnutritioninduceda 40%bodyweightlossatday70.

Atweaning,protein malnutritionwasinducedby submitting ratstolow(8%)proteindiet whilenormalsubjectsweremain- tainedona27%proteindiet(Bengelloun,1990).Suchmalnutrition inducedduringthepostnatalperiodatbirth(Fukudaetal.,2002), oratweaning(Bengelloun,1990)ledtobodyweightdecreasesof respectively40%and50%ofcontrols.

Adultrats2monthsofagewerefedstandardlaboratorychow containingprotein(17%) supplementedwithlysine,methionine andcystein,whereasamalnourishedgroupwerefedalowpro- tein dietcontainingcasein(8%) supplementedwithmethionine

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Table1

Malnutritioninrats:bodyweighteffect.

Dietarymanipulation Ageoftesting(days) Results References

Malnutrition

Type Timing

Protein,6%caseindiet Prenatal P0

P45 P70

−15%

−5%

Nodifference

Almeidaetal.(1996b)

P21 AfterP21

−21%

−11%

Nodifference

Chenetal.(1997)

Protein,6%caseindiet Prenatal P9 −9%, Kehoeetal.(2001)

Protein,6%caseindiet Lactation(P0–P21) P21 P70

−60%

−20%

Santuccietal.(1994)

Protein,8%caseindiet P0–P50 P70 −40% Fukudaetal.(2002)

Protein,8%caseindiet P25–P60 P60 −50% Bengelloun(1990)

Protein,6%caseindiet P0–P21 P70 −20% Hernandesetal.(2005)

Protein,8%caseindiet Adult P60–P240 P320 −4% LukoyanovandAndrade(2000)

P0:birth,E:embryonicperiod,P:postnatalperiod.Thedecreaseofbodyweightisrepresentedrelativetocontrolbodyweight(100%).

(LukoyanovandAndrade,2000).Thisproteindeprivationlasting either6or8monthsledtoa4–5%bodyweightloss.

Undernutritionatbirthbylargelitter groupscomposedof 16 pupsfromdamsgivingbirthonthesameday(eightinthewell- nourishedgroup), induceda 16% body weight loss(Hernandes et al., 2005). Another method of undernutrition involved sep- aration, pups being kept half of the day with a non lactating female(Castro and Rudy, 1987)or in an incubator(Hernandes et al., 2005). This undernutrition led respectively to 40% and 18%bodyweightloss(CastroandRudy,1987;Hernandesetal., 2005).

Undernutritionatweaningwasinducedbyreducingamountsof nutrimentssoastomaintainundernourishedpupsat80%ofbody weightofthecontrol.Thisresultedina20%bodyweightreduction (Alamyetal.,2005).

Toourknowledge,therearenoreportedstudiesexaminingthe effectsofpostnatalundernutritioninadult,anaspectwhichmaybe interestingtoinvestigate.However,Levayetal.(2010)showedthat afterlifelongrestrictedfoodintakeat25%(mildrestriction)through preconception,gestation,lactationandpostweaningperiods,rats weighed18%lessthancontroloffspring.

Inconclusion,allthesedatashowedthatmalnutritionaswell asundernutritioninducedadecreaseinbodyweightandthatthis effectwashigherwhenitoccurredatbirth,duringlactationand throughthemonthfollowingweaning.Itshouldbenotedthatall ofthese studiesdescribed a recoveryin bodyweight lossafter refeeding.

3. Neuroanatomicalanddevelopmentaleffectsof malnutritionorundernutrition(Tables3and4)

Maturationaldevelopmentofthebraininvolvesaseriesoftem- porallyoverlappingstages(Morganeetal.,2002).Theimpactof malnutritiononbrainmustbeanalyzedinthecontextthatthe developmentofthecentralnervoussystemoccursinphaseswhich differintermsoftimeofonsetandindurationbetweenregionsof thebraininthesamespeciesandalsofromonespeciestoanother.

Acrossspecies,substantialdifferenceswithrespecttobraindevel- opmentarerepresentedbythetimingofbirthinrelationtothe stageofbrainmaturation(Morganeetal.,2002).Malnutritioncan altertheactivityofenzymesandinterferewithproteinsynthesis andproteinstructureaswiththeincorporationoflipidsintovarious brainstructures.Proteinsdonotcrosstheplacentainasigniﬁcant amountbutallaminoaciddo.Thenutritivevalueofproteinsin thedietresidesessentiallyintheindividualaminoacidsthathave beenabsorbedinthematernaldigestivesystem.Itappearsthatall aminoacidsareessentialforthemotherandthefoetus.Omission ofasingleessentialaminoacidfromthematernaldiet(methionine, phenylalanine,arginine,lysineortryptophan)canhavedeleterious effectsonfoetalbraindevelopmentsimilartothoseproducedby theomissionofproteinsasawhole(Zamenhof,1991).

Malnutritionappearstoaffectbraindevelopmentincludingcell birthand migration,and differentiationincludingmyelinization andsynaptogenesis.Prenatalandpostnatalproteinmalnutrition affected cell number, synapses, myelinization (Debassio et al.,

Table2

Undernutritioninrats:bodyweighteffect.

Dietarymanipulation Ageoftesting(days) Results References

Undernutrition

Type Timing

Undernutrition P2–P18 P22 −40% CastroandRudy(1987)

25%restriction Preconception,E,P P70 −18% Levayetal.(2010)

Uterine-arteryligation E17 P0

P14 P21

−8%

−10%

−13%

Olivieretal.(2007)

50%foodintake P0–P21 P70 −25% Hernandesetal.(2005)

Largelittersize P0–P21 P70 −16% Hernandesetal.(2005)

Undernutritionbyseparation P0–P21 P70 −18% Hernandesetal.(2005)

50%foodintake E0–P21 P21 −73% JahnkeandBedi(2007)

50%foodintake E0–P21 P21

P62

−66%

−33%

Partadiredjaetal.(2008)

Reductioninfoodintake P25–P125 P25–P125 −20% Alamyetal.(2005)

P0:birth,E:embryonicperiod,P:postnatalperiod.

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Table3

Malnutritioninrats:neuroanatomicaleffects.

Dietarymanipulation Ageoftesting

(days)

Test Results References

Malnutrition

Type Timing

Protein,6%caseindiet Prenatal P30 E12–E16–E20

Injectionof tritiatedthymidine

Decreaseingranulecells onE20

Noeffectonpyramidal cellsinhippocampus

Debassioetal.

(1994)

P90

P8–P15 P30 Injectionof tritiatedthymidine

Decreaseinnumberof granulecellsonP8 NoeffectatP15 IncreaseatP30

Debassioetal.

(1996)

Protein,5%caseindiet Prenatal E0–E15gestation

P15 Immunohistochemistry Delayedastrogenesis Abnormalneuronal differentiation

Abnormalsynaptogenesis

Gressensetal.

(1997)

Protein,7%caseindiet Gestationlactation P2 P15 P60

Glialacidicprotein Astrogliosisincerebral cortexandhippocampus

Feolietal.(2008)

P220

Golgi Decreaseincellsize

Decreaseindendritic complexity Decreaseinsynaptic density

Díaz-Cintraetal.

(1991)

Protein,8%caseindiet Adult P60–P180 P60–P360 P60–P540

P180 P360 P540

Numericaldensity Toluidineblue stain

Progressivelossofneurons ingranulecellsandCA3 pyramidalcells Decreaseinthicknessof CA3pyramidallayer

Paula-Barbosaetal.

(1989)

Protein,8%caseindiet Adult P60–P240

P320 Estimationoftotal

numberofneurons andsynapses Giemsastain

Lossofhippocampalcells andsynapses

Lukoyanovand Andrade(2000)

E:embryonicperiod,P:postnatalperiod.

1994; Lukoyanov and Andrade, 2000). More recently a study reported the occurrence of astrogliosis after protein malnutrition (Feoli et al., 2008). Nutritional rehabilitation can reverse somebutnotallofthesestructuralandmorphologicalalterations (Bengelloun,1990;LukoyanovandAndrade,2000;Winnick,1969).

Prenatalproteinrestrictioninitiated atconception,and main- tainedduringtheﬁrst2weeksofratgestation(Gressensetal., 1997)induced a delayed astrogenesis, a delayed production of hyaluronanintheextracellularmatrix,abnormalneuronaldiffer- entiation,decreasedprogrammedcelldeathintheneocortexand inthecerebellum,aswellasabnormalsynaptogenesisinthebasal ganglia3 weeksafterbirth.Analysisofadultrat brainsdidnot revealasigniﬁcantalterationofbrainarchitectureandneuronal

differentiation,suggestingahighlevelofplasticityinthedevelop- ingbrain(Gressensetal.,1997).

Prenatal proteindeprivationledtoabnormalitiesin thehippocampalformation,alteratingthedevelopmentaltimecourseof dendategyrus(Debassioetal.,1996,1994)andthemorphology of hippocampalcells (Díaz-Cintra et al.,1991).Prenatal malnutritioninrats,withchronicproteinrestrictionfor5weeksprior tomatingthatcontinuedduringpregnancy,hadnoeffectonthe generationofpyramidalcellsbutinduceddecreasedneurogenesis ingranulecellsatE20(Debassioetal.,1994)andatP8(Debassio etal.,1996).Theirresultsfurthershowedthatprenatalproteinmal- nutritionalteredthepostnataldevelopmentofgranulecellslong afterrehabilitationatbirth,emphasizingtheimportanceofearly nutritiononlaterdevelopmentalevents.Thereportedincreasein granulecellsinmalnourishedratsonP30 suggestedacompen- satoryincreaseinneurogenesisinthepostnatalperiod.Moreover,

Table4

Undernutritioninrats:neuroanatomicaleffects.

(days)

Undernutrition

Type Timing

50%foodintake Gestation Lactation

P120 Histology Decreaseincelldensityinseptumand

hippocampalﬁelds

JordanandClark (1983)

50%foodintake E16–P30 P70

P212

Histology NoeffectonP70,decreaseingranule cellsonP212

Bedi(1991)

50%foodintake E0–P21 P21 TUNELlabeling Increasedapoptosisindentategyrus JahnkeandBedi

(2007)

Undernutrition E1–P21 P70 Nitricoxide

synthase

Decreasednitricoxidepositivecellsin CA1,CA3anddentategyrusin hippocampus

Zhangetal.(2010a)

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Table5

Malnutritioninrats:behavioraleffects.

Dietarymanipulation Ageoftesting (days)

Malnutrition

Type Timing

Protein Prenatal P91 Visual

discrimination learning

Moreerrors Longertimetolearn

Tonkissetal.(1991)

Protein Prenatal P125

P225

Foodreward Saccharin-reward

Increasedresponsetoreward Tonkissetal.(1990b)

Protein Prenatal P45 Socialbehavior Decreasedsocialbehavior

Increasednon-socialrearing

Almeidaetal.(1996c)

Protein P0–P21 P0-26,36,46, P56,66,76

P26,36,46 P56,66,76

Socialplay Decreasedpinningbehavioral Increasewrestling,walkoverand rearing

CamargoandAlmeida(2005)

Protein Prenatal P130 Openﬁeld Fewerentriesincentralarena Trzcinskaetal.(1999)

Protein Prenatal P70 T-maze Reductionofanxiety Almeidaetal.(1996b)

Protein Prenatal P0–P21

P70 ElevatedT-maze Reductionofanxiety Hernandesetal.(2005)

Protein Prenatal P35 Elevatedplusmaze

andenvironmental stimulation

Anxiolyticeffectofenvironmental stimulationlessevidentin malnourishedrats

Pereira-da-Silvaetal.(2009)

Protein Prenatal P90 Kindlingprocedure Sameeffectonthetwonutritional

group

Shultzetal.(1995)

Protein Prenatal P160 DRLoperanttask Impairedperformanceduring

acquisition

Tonkissetal.(1990b)

Protein Prenatal P16–20

P20–27 P70–71 P220–221

Morrismaze Noimpairmentsineitherproximal orindistalcuediscrimination

Tonkissetal.(1994)

Protein P0–P50 P70 Morrismaze Impairedlearningindistalcuebut

notproximalcuediscrimination

Fukudaetal.(2002)

Protein P25–P60 P60

P109

Openﬁeld Passiveavoidance

Increasedactivity

Nodifferenceinpassiveavoidance

Bengelloun(1990)

Protein P60–P240 P320 Morrismaze

Openﬁeld

Noeffectsduringtraining Retentiondeﬁcits

Reductionemotionalreactivity

LukoyanovandAndrade(2000)

morphometricGolgistudiesofgranulecellsafterprenatalprotein deprivationshowedareductionincellsize,dendriticcomplexity andsynapticspinedensitymeasuredatdifferentagesfromP15 toP220(Díaz-Cintraetal.,1991),withthemostmarkeddeﬁcits notedonP220.Inglutamatergicgranulecells,malnutritionreduced themossyﬁberaxonalarea(Andradeetal.,1991;Granados-Rojas etal.,2002)andthenumberofdendritesandspines(Díaz-Cintra etal.,1994,1991).Similarmorphologicaleffectsofmalnutrition havebeenfoundinCA3pyramidalcells(Cintraetal.,1990;Díaz- Cintraetal.,1991;García-Ruizetal.,1993;Granados-Rojasetal., 2002).Delayedastrocytogenesisandabnormalneuronaldifferen- tiationandsynaptogenesiswerealsoobservedbytheseauthors.

Feolietal.(2008)evaluatedspeciﬁcglialchangesinratsexposed topreandpostnatalproteinmalnutritionandfoundastrogliosis

in thecerebral cortex and hippocampus. Prenatal proteinmalnutrition altered thenumber of GABA interneurons in dentate gyrusandinCA1–3subﬁeldsofhippocampus(Díaz-Cintraetal., 2007).

Undernutritionduringgestationandlactationalteredthenum- berofpyramidalcells(JordanandClark,1983),withadecreased neuronal cell density in the septum and hippocampal ﬁelds CA3, CA4 and in the dentate gyrus at 4 months of age. This effect was extended to offspring as shown by Ranade et al.

(2008) who noted a reduced hippocampal volume in all pups when the mother was subjected to different types of malnutrition duringtheperiod ofgestationand lactation. Jahnkeand Bedi (2007) reported that undernutrition during gestation and lactation can result in an increase in the level of apoptotic

Table6

Undernutritioninrats:behavioraleffects.

Dietarymanipulation Ageoftesting(days) Test Results References

Undernutrition

Type Timing

Undernutrition Prenatal–P25 P105 Electricshock Decreasethresholdofresponse Smartetal.(1975)

Undernutrition P25–P125 P46–P48

P81 P84 P123

Openﬁeld Radialmaze Tailﬂick

Hyperactivity Sameacquisition Sameretention Decreaseinlatency

Alamyetal.(2005)

Undernutrition P2–P18 P22 Morrismaze Impairedlearningindistalcuebut

notproximalcueversion

CastroandRudy(1987)

Undernutrition E1–P21 P70 Morrismaze Impairedlearningandmemory Zhangetal.(2010a)

E:embryonicperiod,P:postnatalperiod.

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Table7

Malnutrition:effectsonbrainneurochemistry.

Dietarymanipulation Ageoftesting (days)

Test Drug Results References

Malnutrition

Type Timing

Protein Prenataland postnatal

P100 Locomotoractivity Amph

Apo

Nodifference Stereotyped behavior

Leahyetal.(1978)

Protein Prenatal P90–P120 Microdyalisis DA

∼5-HT

Mokleretal.(2007)

Protein Prenatal+suckling P62 HPLC 5-HT

5-HIAA

Alfaro-Rodrigez etal.(2006)

Protein Prenatal P3,P5,P8,P11,

P13P16,P21, P30

Spectoﬂuorescence 5-HT

5-HIAA Trp Albumin

Resnickand Morgane(1984)

Protein Prenatal P1,P15,P30,

P45

HPLC DA 5-HT

Noeffecton DAand5-HT Trp Tyr

Chenetal.(1997)

Protein Prenatal+P2–8 stress

P9 HPLC Hypothalamus

DA DOPAC 5-HT +5-HIAA Hippocampus DA 5-HT

Kehoeetal.(2001)

Protein P60–P150 P150 Locomotoractivity Amph

Apo

Locomotoractivity Noeffect

Dietrichetal.

(2004)

Protein Prenatallactation Biochemicalassays Tyr

Trp

Feolietal.(2006)

Protein Prenatal P90 Morrismaze CDP

i.p.

Spatiallearningdeﬁcits Tonkissetal.

(2000b)

Protein Prenatal P110–P180 Morrismaze CDPinseptum Morerapidlatencyto

escape

Learningimpairment Ratsmalnourishedless sensitivetoCDP

Tonkissetal.

(2000a)

Protein Prenatal P7,P11 Ultrasonicvocalization

(USV)

DZ Differencein

sensitivitytoUSV suppresseffectofDZ

TonkissandGaller (2007)

Protein Gestation–P50 P90 Elevatedplusmaze

GABAreceptors

DZ Reductionoftolerance

Increaseuptake

Borgheseetal.

(1998)

Protein Prenatal P35,P56 Prepulseinhibition

Receptorbinding assays

ReducedPPIatP56 Increaseinstriatal NMDAreceptor bindingatP56notP35

Palmeretal.(2004)

P56 P59–P61

Apo-induced stereotypy Amph-induced locomotion Hal-inducedcatalepsy MK801-induced locomotion

Receptorbindingassay Apo Amph Hal MK801

3H-MK801

3H-Hal

3H-GRD-12935 (bindstoDA transportersites)

Nodifference Stereotypicresponse infemalerats ResponsetoAmph inadultdeprived femalerats NodifferenceinHal catalepsyandin MK801locomotion In³MK801-binding inthestriatumand hippocampusand³Hal inthestriatumin femalerats InDAtransporter bindinginthestriatum infemalerats

Palmeretal.(2008)

Protein E14–P40 P80–90 Conditionedeffect Cocaine Responsivenessto

cocaine

Valdomeroetal.

(2006)

Apo:apomorphine;Amph:amphetamine;CDP:chlordiazepoxide;DA:dopamine;DOPAC:dihydroxyphenylaceticacid;DZ:diazepam;E:embryonicperiod;Hal:haloperidol;

5-HT:serotonin;5-HIAA:5-hydroxyindoleaceticacid;P:postnatalperiod;Trp:tryptophan;Tyr:tyrosine.

cells in the dentate gyrus but not in the CA2–CA3 region of thehippocampalformation. Accordingtotheseauthors,undernutrition may induce cell death related to the availability of neurotrophicfactors.Apoptoticprocessesarehighlysensitiveto nutritionalstatusandimmaturegranuledentatecellsmaybemore

vulnerabletoundernutrition thanmaturepyramidalcells at21 daysofage.

Growthrestrictionbyunilateralligationoftheuterinearteryon embryonicday17wasassociatedwithdiffusewhitematterdamage inratsaswellasmicrogliaactivationandastrogliosis.Therewasa

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Table8

Undernutrition:effectsonbrainneurochemistry.

Dietarymanipulation Ageoftesting(days) Test Drug Results References

Undernutrition

Type Timing

Undernutrition Prenatal P1–P10 Brainandblood5-HT

PlasmadosageofTrp

5-HT Trp(plasmatic)

Hernandezetal.(1989)

Undernutrition P0–P21 Suckling

P30 THactivity DAreceptorsin

striatum

Wigginsetal.(1984)

Undernutrition P25–P125 P67

P81 P84

HPLC Radialmaze

Hal DA,∼5-HT

∼Learning Retention

Alamyetal.(2005)

Chronicstarvation P25–P95 P95 Activity Hal Suppressionofreduced

retention

MasurandRibiero(1981) DA:dopamine;E:embryonicperiod;Hal:haloperidol;5-HT:serotonin;P0:birth;P:postnatalperiod;Trp:tryptophan.

Table9

Malnutrition:oxidativestatus.

(days)

Malnutrition

Type Timing

Protein Prenatal Lactation

Biochemicalassays Lipid

Peroxidationincerebellumand cerebralcortex

Feolietal.(2006)

P75

SOD CAT

Lipoperoxidation

SOD CAT

Lipoperoxidationin hippocampus

Bonattoetal.(2005)

Protein Prenataland P0–P21

P60 CAT CAT Tatlietal.(2007)

CAT:catalase;P:postnatalperiod;SOD:superoxidedismutase.

lossinimmatureoligodendrocytesonpostnatalday7followedbya delayinmyelinization.Inthecaseofmoderategrowthrestriction, restorationofmyelinizationwasobservedonpostnatalday14with increasesinthedensityofoligodendrocytesandintheproliferation ofimmatureoligodendrocytes(Olivieretal.,2007).

Perinatalundernutrition resulting fromthereduction of both proteinandfattyacidintakethroughoutpregnancy,lactationand fromweaningtoday70,ledtoadecreaseinvolumeofthehip- pocampusandthecerebellum(Souzaetal.,2008).

Perinatalundernutritionindamsfedhalfthedailyfoodintake ofcontrolsduringgestationandlactationresultedinareduction inthetotalnumberofneuronsindentategyrus(granulecells)of offspring(Bedi,1991).Matosetal.(2011)showedthatthisrestric- tionreducescellproliferationin thehippocampusatadulthood butdoesnotimpairtheneuronaldifferentiationprocessesofthe

newgenerated cells.Inthis modelofundernutrition,Katzetal.

(1982)describedadecreasedwidthofhippocampusinratdeprived duringaperiodincludinggestation.Moreover,perinatal50%food restrictionproduceddeleteriouseffectsonthepopulationofnitric oxidesynthaseneuronsinCA1,CA3anddentategyrusregionofthe hippocampusinadultoffspring.Changesinthedensityofthese neuronsmay partlyexplain learning and memorydisturbances observedinundernourishedrats(Zhangetal.,2010a).

3.2. Postnatalmalnutritionorundernutrition

Dietarytryptophan(aserotoninprecursor)restrictioninrats frompostnatal day30 to 60 inducedreduced arborisationand altereddendriticspinedensityinCA1pyramidalneuronsandgran- ulecellsofthehippocampuswithastrocyteactivation.Serotonin

Table10

Undernutrition:oxidativestatus.

Dietarymanipulation Ageoftesting(days) Test Results References

Malnutrition

Type Timing

Caloricrestriction Pregnancy P0 Malondialdehyde

levelsinbrain Polyunsaturatedlipids levels

Fattyacidsinbrain andplasmaindams andnotinoffspring Reducedantioxidant capacity

Agaleetal.(2010)

Proteinandfattyacidrestriction PregnancyandP0–P21 P70 Lipidextractionand chromatography

Lowerlevelsinlinoleic andlinolenicin hippocampusand linoleicacidin cerebellum

Souzaetal.(2008)

P0:birth;P:postnatalperiod.

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maymodulateglialmorphologythroughalteredhomeostaticor metabolicinteractionsbetweenneuralcells,producing apatho- physiologicalconditionforneuraltransmission(Zhangetal.,2009).

Protein deprivationinadultcaused a 30%loss ofsynapses in mossyﬁbersanddendritesofhippocampus.Thetotalnumberof granulecellsandCA3pyramidalneuronswasalsoreducedrelative tocontrols(LukoyanovandAndrade,2000;Paula-Barbosaetal., 1989).AprogressivelossofgranulecellsandpyramidalCA3neu- ronswasfound,withreducedthicknessofCA3pyramidallayer inratssubjectedtoproteinmalnutritionfor6,12or18months (Paula-Barbosaetal.,1989).Thedecreaseinthetotalnumberof neuronswasobservedeveninnutritionallyrehabilitatedanimals (2monthsperiodofrehabilitation)(LukoyanovandAndrade,2000).

Amarkeddecreaseinthetotalnumberofsynapseswasdetected inproteinmalnourishedratsbutnotinnutritionallyrehabilitated animals,whichsuggestedthattheshifttoadequatedietreversed thesynapticchange(LukoyanovandAndrade,2000).

Toourknowledge,therearenoreportsconcerningtheeffect ofpostnatalundernutritiononcellnumberandsynaptogenesisin adult.Thisaspectmaybeinterestingtoinvestigate.

In conclusion, prenatal and postnatal protein malnutrition inducedsimilargranulecelllossinhippocampusbutonlypostna- talproteinmalnutrition(duringlactationorintheadult)affected pyramidalcells.Differenttimecoursesingenesisofpyramidaland granulecellsmayexplainthisdifferenceintheextentofcellloss.

Rehabilitationreversedsynapticchangeregardlessoftheperiodof malnutrition.

Both malnutrition and undernutrition during gestation and lactationinduced granulecellloss.Differencesbetweenstudies concerningtheincidenceofgranulecelllossmaybeexplainedby thedegreeofcelldeathfollowingmalnutritionorundernutrition and/orbyaninitialdelayingranulecellgenerationinmalnourished animalswhichiscompensatedbyalaterincreaseinneurogene- sis(Debassioetal.,1996).Suchacompensatorymechanismmay notextendintoolderage.Sinceonlyafewstudieshaveaddressed undernutritionduringtheprenatalorpostnatalperiods,itwasdif- ﬁculttocomparetheeffectsofmalnutritionandundernutritionin termsoftheoccurrenceordurationoftheirrespectiveeffects.

4. Behavioraleffectsofmalnutritionorundernutrition (Tables5and6)

Malnutritionearlyinlifeaffectedthemorphologyandneuro- chemistryofthehippocampalformation.Thisstructureisknown tobeinvolvedinfunctionssuchaslearningandmemory,especially thatofspatialcharacteristics(Morris,1984).Thusmoreattention hasbeendevotedtospatiallearningthanotherlearningparadigms, usingtestssuchradialmaze(Jordanetal.,1981),spatialalterna- tion(Goodlettetal.,1988)andMorrismaze(CastroandRudy,1987;

Fukudaetal.,2002;Lausetal.,2011;LukoyanovandAndrade,2000;

Souzaetal.,2008;Tonkissetal.,1994).

Inthecaseofprenatalproteinmalnutrition,theanalysisofprox- imalanddistalcuelearninginMorriswatermazeshowedthatthe developmentofthesetwotypesofnavigationcuesappearstobe unaffectedbymalnutrition.Prenatallymalnourishedandcontrol ratsweresimilarintheirrateofacquisitionbothatweaningand inadulthood(Tonkissetal.,1994).However,Bronzinoetal.(1991) reportedanenhancedinhibitioninthehippocampalformationof ratssubjected topairpulse paradigm.They alsoindicated that itwasdifﬁculttoinduceandtomaintainlong-termpotentiation inmalnourishedrats(Bronzinoetal.,1997,1994).Theseauthors, (Bronzinoetal.,1999,1996),suggestedthatproteinmalnutrition

altersmodulationofdentategyruscellexcitabilityinfreelymov- ingratsduringtheearlystageofdevelopment(15and30days)but notinadult(90days)followingstimulationofthemedialperforant path.Themostsigniﬁcanteffectsofproteinmalnutritionoccurred attheearlierstagesofdevelopment.At15daysofage,thepaired- pulseindex(PPI)valuesusingthepopulationspikeamplitude(PSA) measureshowedsigniﬁcantlygreaterlevelsofearlypaired-pulse depression(PPD)inmalnourishedanimalsrelativetocontrolani- mals.However,at30daysofage,themalnourishedgroupexhibited greaterfacilitationatinterpulseintervals(IPIs) between50and 70msduringquietwaking(QW)andslow-wavesleep(SWS)states, butnotrapideyemovementsleepstate(REM),whencomparedto thecontrolgroup.Thus,itappearedthatprenatalproteinmalnutri- tionaffectedearlyinhibitoryprocessesinthe15-day-oldanimals andalteredthedevelopmentoffacilitatoryprocessesinthejuve- nile(30-day-oldanimals).AstheanimalcontinuedtomaturePPI valuesintheadultmalnourishedanimalbecamesimilartothose ofthecontrolgroupinadulthood.

Prenatal protein malnutrition was alsoreported to increase responsestoreward(Tonkissetal.,1990a)whenratsweretested in adulthood, to impair visualdiscrimination learning (Tonkiss etal.,1991)andtheacquisitionofadifferentialreinforcementof lowratesofrespondingtask(Tonkissetal.,1990b),allsuggesting increasedimpulsiveness(Almeidaetal.,1996b).

Prenatalmalnutritionaffectedsocialinteractionsofjuvenilerats (AlmeidaandDeAraujo,2001;Almeidaetal.,1996c).Itdecreased bothplayfulpinandnonplayfulsocialbehaviorsincludinganogen- ital sniff, walkover and allogroom. It also increased non-social rearing.Thisalteredearlysocialbehaviorinratstestedat45daysof ageafterprenatalproteinmalnutritionmayplayanimportantrole indeterminingsomeofthelaterbehavioraldifferencesdescribed inadultanimals(Almeidaetal.,1996c).

Ithasbeendemonstratedmorerecentlythatproteinmalnour- ishedanimalsexhibited behaviormore consistentwithalower anxietyand/orahigherimpulsivenessinmodelsofanxiety(ele- vatedmaze)involvingaversivestimulithatwereneitherpainful nor artiﬁcial. In suchmodels, malnourished ratsshowed more transitionsinthelight-darktransitiontest(BrioniandOrsingher, 1998),moreopenarmsexplorationintheelevatedplusmazetest (Almeidaetal.,1996b)andlowerinhibitoryavoidancelatencyin theelevatedT-mazetest(Almeidaetal.,1996b;Hernandesetal., 2005).Malnourishedratsexhibitedloweranxiety(Hernandesetal., 2005).

Inthecaseofprenatalundernutrition,maternaldeprivation(MD) differentially affected physiological variables in offspring. After MDfor12h(MD12)frompostnatalday1today7,pupsshowed increasedbasalandstress-inducedcorticosteronelevelsinplasma.

Atweaning,theircircadianrhythmswerephase-reversedinthe absenceoflight(Yamazakietal.,2005).Theseauthorsshowedthat theeffectsofMDonthecircadianclockdependedonthelengthof MD.ThelongerMD,thegreaterwasthephase-shiftofthecircadian rhythminduced.

Smart et al. (1975) showed that a mildly painful stimulus (foot-shock)producedgreaterreactivityinratssubjectedtounder- nutrition prenatally and immediately postnatally suggesting that undernourishedanimalsweremorereactivetounpleasantoraver- sivestimuliirrespectiveoftheageatwhichthenutritionalregime wasinitiated.

Malnourished rats exhibited lower anxiety, (Franc¸olin-Silva et al., 2006; Hernandes et al., 2005), the extent of which was related to the duration of postnatal exposure todeﬁcient diet.

Theseresultswereinlinewithresultsobtainedwithdiazepam treatment(Franc¸olin-Silvaetal.,2006).Theauthorssuggestedthat short-termmalnutritionmayaffectneuraland/orneurochemical systems believed to underlie behavioral expression in anxio- genicexperimentalsituations.Moreover,anumberofhormonal