Reorganization of Movement Representations
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J~URNALOFNEUROPHYSIOLOGY RAPID PUBLICATION Vol. 75, No. 5, May 1996. Printed in U.S.A. Reorganization of Movement Representations in Primary Motor Cortex Following Focal Ischemic Infarcts in Adult Squirrel Monkeys RANDOLPH J. NUDO AND GARRETT W. MILLIMEN Department of iVeurohiology and Anatomy, University of Texas Health Science Center at Houston, Houston, Texas 77030 SUMMARY AND CONCLUSIONS 1. Intracortical microstimulation (ICMS ) techniques were used to derive detailed maps of distal forelimb movement representa- tions in primary motor cortex (area 4) of adult squirrel monkeys before and a few months after a focal ischemic infarct. 2. Infarcts caused a marked but transient deficit in use of the contralateral hand, as evidenced by increased use of the ipsilat- era1 hand, and reduced performance on a task requiring skilled digit use. 3. Infarcts resulted in a widespread reduction in the area1 extent of digit representations adjacent to the lesion, and apparent in- creases in adjacent proximal representations. 4. We conclude that substantial functional reorganization occurs in primary motor cortex of adult primates following a focal isch- emit infarct, but at least in the absence of postinfarct training, the movements formerly represented in the infarcted zone do not reappear in adjacent cortical regions. INTRODUCTION Despite over a century of study, the precise neurophysio- logical and neuroanatomic mechanisms for functional motor recovery after cortical injury are poorly understood (e.g., Bach-y-Rita 1980). Early studies which employed surface stimulation techniques suggested that the hand representa- tion in primary motor cortex of adult primates undergoes substantial reorganization following small lesions, and that cortical reorganization is correlated with functional recovery (Glees and Cole 1950). These studies have not been re- peated since the introduction of intracortical microstimula- tion (ICMS ) techniques. To examine lesion-induced plasticity in primary motor cortex (area 4) in more detail, we have employed ICMS techniques to derive detailed maps of distal forelimb repre- sentations before and a few months after a focal ischemic infarct. In sharp contrast to results obtained by Glees and Cole ( 1950), movements represented in the infarcted zone did not reappear in the cortical sector surrounding the infarct. METHODS Standard ICMS mapping techniques were employed in nine nor- mal, adult squirrel monkeys (Saimiri boliviensis) to derive maps of distal forelimb representations. After the derivation of baseline maps, one group of animals underwent ischemic infarct procedures (~2 = 5). A second group of animals, serving as controls, underwent ICMS procedures, but no infarct (n = 4). Surgical and electrophysiological techniques Under sterile conditions and after induction of halothane/nitrous oxide anesthesia, area 4 contralateral to the animal’s preferred hand was exposed. Then, under ketamine anesthesia ( 15 mg l kg -I l h -’ ) , standard ICMS mapping procedures were conducted (e.g., Gould et al. 1986; Nudo et al. 1992; Strick and Preston 1982). Briefly, an electrolyte-filled glass micropipette was advanced perpendicular to the cortical surface to a depth of 1,700- 1,800 pm at several hundred sites within the distal forelimb (digit, wrist, forearm) representation. Interpenetration distances were -250 pm throughout the distal fore- limb zone, and 500- 1,000 pm in the surrounding, proximal zone. The ICMS train burst consisted of 13 200~ps cathodal pulses deliv- ered at 350 Hz from an electrically isolated stimulation circuit. Site- specific movements evoked at threshold and at a fixed suprathreshold level (20 PA) were recorded (maximum = 30 PA). With the use of a computer algorithm, representational maps were drawn by out- lining cortical sectors whose stimulation evoked specific movements (see Nudo et al. 1992 for further details). After mapping was completed, the anesthetic regimen was switched to halothane/nitrous oxide (infarct group). Small cortical lesions were made by using microforceps connected to a bipolar electrocoagulator. Blood vessels were permanently occluded as they entered the cortical surface. The boundary between the adja- cent, intact cortex was easily discernible within a few minutes because the infarcted tissue became blanched throughout its extent, while the adjacent tissue and its vascular supply remained intact. The extent of the damaged cortex was then measured directly from comparison of digital photographs taken before and several minutes after the infarct. This boundary was later verified by examining the following: 1) digital photographs taken immediately before the postinfarct mapping procedure, 2) regions unresponsive to ICMS in postinfarct maps, and 3) postmortem histological assessment. These procedures verified that immediate postinfarct estimates of lesion size were reliable. ICMS procedures were repeated 3 -5 mo after the ischemic in- farct. After completion of these experiments, each animal was injected with a lethal dose of pentobarbital, perfused with fixative, and the brain removed for histological examination. The area 3a/ 4 border was defined, and extent of the lesion verified. These procedures are in accordance with National Institutes of Health guidelines and were approved by the institutional animal care and use committee. Behavioral techniques To assess spontaneous reorganization in motor maps, no specific behavioral training procedures were employed either before or after the infarct. Although extensive behavioral testing could have introduced training effects, limited testing of motor performance was conducted in two monkeys before and after the infarct, to determine the extent of the motor deficit and track recovery. Test- 2144 0022-3077/96 $5.00 Copyright 0 1996 The American Physiological Society
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J~URNALOFNEUROPHYSIOLOGY RAPIDPUBLICATION Vol.75,No.5,May1996.PrintedinU.S.A. ReorganizationofMovementRepresentationsinPrimaryMotorCortex FollowingFocalIschemicInfarctsinAdultSquirrelMonkeys RANDOLPHJ.NUDOANDGARRETTW.MILLIMEN DepartmentofiVeurohiologyand Anatomy,UniversityofTexas HealthScience CenteratHouston, Houston,Texas 77030 SUMMARYAND CONCLUSIONS 1.Intracorticalmicrostimulation(ICMS)techniqueswereused toderivedetailedmapsofdistalforelimbmovementrepresenta- tionsinprimarymotorcortex(area4)ofadultsquirrelmonkeys beforeandafewmonthsafterafocalischemicinfarct. 2.Infarctscausedamarkedbuttransientdeficitinuseofthe contralateralhand,asevidencedbyincreaseduseoftheipsilat- era1hand,andreducedperformanceonataskrequiringskilled digituse. 3.Infarctsresultedinawidespreadreductioninthearea1extent ofdigitrepresentationsadjacenttothelesion,andapparentin- creasesinadjacentproximalrepresentations. 4.Weconcludethatsubstantialfunctionalreorganizationoccurs inprimarymotorcortexofadultprimatesfollowingafocalisch- emitinfarct,butatleastintheabsenceofpostinfarcttraining, themovementsformerlyrepresentedintheinfarctedzonedonot reappearinadjacentcorticalregions. INTRODUCTION Despite overa centuryofstudy,the precise neurophysio- logicaland neuroanatomic mechanismsforfunctionalmotor recoveryaftercorticalinjuryarepoorlyunderstood(e.g., Bach-y-Rita1980).Earlystudies whichemployedsurface stimulationtechniques suggested thatthehandrepresenta- tioninprimarymotorcortexofadultprimatesundergoes substantial reorganizationfollowingsmall lesions, and that corticalreorganizationis correlated withfunctionalrecovery (GleesandCole1950).Thesestudies havenotbeenre- peated since theintroductionofintracorticalmicrostimula- tion(ICMS)techniques. Toexaminelesion-inducedplasticityinprimarymotor cortex(area4)inmoredetail,wehaveemployedICMS techniques toderivedetailed maps ofdistal forelimbrepre- sentations beforeanda fewmonthsaftera focalischemic infarct.Insharp contrasttoresultsobtainedbyGlees and Cole( 1950),movements represented intheinfarctedzone did not reappear in the corticalsector surrounding the infarct. METHODS StandardICMSmappingtechniqueswereemployedinninenor- mal,adultsquirrelmonkeys(Saimiriboliviensis)toderivemaps ofdistalforelimbrepresentations.Afterthederivationofbaseline maps,onegroupofanimalsunderwentischemicinfarctprocedures (~2 =5).Asecondgroupofanimals,servingas controls,underwent ICMSprocedures,butnoinfarct(n=4). Surgicalandelectrophysiologicaltechniques Understerileconditionsandafterinductionofhalothane/nitrous oxideanesthesia,area4contralateraltotheanimalspreferredhand wasexposed.Then,underketamineanesthesia( 15mgl kg-Il h -) , standardICMSmappingprocedureswereconducted(e.g.,Gouldet al.1986;Nudoetal.1992;StrickandPreston1982).Briefly,an electrolyte-filledglassmicropipettewasadvancedperpendicularto thecorticalsurfacetoa depthof1,700-1,800pmatseveralhundred siteswithinthedistalforelimb(digit,wrist,forearm)representation. Interpenetrationdistanceswere-250pmthroughoutthedistalfore- limbzone,and500-1,000pminthesurrounding,proximalzone. TheICMStrainburstconsistedof13200~pscathodalpulsesdeliv- eredat350Hzfromanelectricallyisolatedstimulationcircuit.Site- specificmovementsevokedat thresholdandata fixedsuprathreshold level(20PA)wererecorded(maximum=30PA).Withtheuse ofacomputeralgorithm,representationalmapsweredrawnbyout- liningcorticalsectorswhosestimulationevokedspecificmovements (seeNudoetal.1992forfurtherdetails). Aftermappingwascompleted,theanestheticregimenwas switchedtohalothane/nitrousoxide(infarctgroup).Smallcortical lesionsweremadebyusingmicroforcepsconnectedtoabipolar electrocoagulator.Bloodvesselswerepermanentlyoccludedas theyenteredthecorticalsurface.Theboundarybetweentheadja- cent,intactcortexwaseasilydiscerniblewithinafewminutes becausetheinfarctedtissuebecameblanchedthroughoutitsextent, whiletheadjacenttissueanditsvascularsupplyremainedintact. Theextentofthedamagedcortexwasthenmeasureddirectlyfrom comparisonofdigitalphotographstakenbeforeandseveralminutes aftertheinfarct.Thisboundarywaslaterverifiedbyexamining thefollowing:1)digitalphotographstakenimmediatelybeforethe postinfarctmappingprocedure,2)regionsunresponsivetoICMS inpostinfarctmaps,and3)postmortemhistologicalassessment. Theseproceduresverifiedthatimmediatepostinfarctestimatesof lesionsizewerereliable. ICMSprocedureswererepeated3 -5moaftertheischemicin- farct.Aftercompletionoftheseexperiments,eachanimalwas injectedwithalethaldoseofpentobarbital,perfusedwithfixative, andthebrainremovedforhistologicalexamination.Thearea3a/ 4borderwasdefined,andextentofthelesionverified. TheseproceduresareinaccordancewithNationalInstitutesof Healthguidelinesandwereapprovedbytheinstitutionalanimal careandusecommittee. Behavioraltechniques Toassessspontaneousreorganizationinmotormaps,nospecific behavioraltrainingprocedureswereemployedeitherbeforeor aftertheinfarct.Althoughextensivebehavioraltestingcouldhave introducedtrainingeffects,limitedtestingofmotorperformance wasconductedintwomonkeysbeforeandaftertheinfarct,to determinetheextentofthemotordeficitandtrackrecovery.Test- 21440022-3077/96$5.00Copyright01996TheAmericanPhysiologicalSociety REORGANIZATIONINMOTORCORTEX2145 A O-e-- CASE#I300 I - I - I - I -20 40 60 80 A14 MOTORMAP DAYSPOST-INFARCTMoTofRMAP &INFARCT C W0z-O z05 0.8- i ne 0.6- 5)-00.4- E o-2-- 4well1 ---I)-well4 MOTORMAP DAYSPOST-INFARCTMOTORMAP &INFARCT D f z a 0 IL 3 e u 0 I- O L CASE#I449 4 well1 I - I - I - I20 40 60 80 +--12 I c MOTORMAP DAYSPOST-INFARCTMOTOR,c,, &INFARCT n4well1 +well4 -.- -200 20 40 60 80 -1204DAYSPOST-INFARCT MOTORMAP MOTORiAp &INFARCT FIG.1.Effectsofischemicinfarctonlateralityandmotorperformance.AandB:percentretrievalswiththehand contralateraltotheinfarctareshownforeachwell.A:lateralitywasnormalforeachwellby60days.Monkeypersistedin usingthecontralateralhandonwell4trialsthroughouttherecoveryperiod.B:lateralitywasnormalforwellsl-3by40 days.CandD:motorperformanceonbothwellswasnearnormallevelsbyday60inCandbyday40inD.Motor performanceisdefinedas(totalnumberfingerflexions/totalnumberretrievals)-I.Maximalperformance=1.0.Forclarity, motorperformanceonlyonwells1(25mmdiam)and4(9.5-mmdiam)areshown. ingwasconductedbyusingaPlexiglasboardcontainingfood wellsofdifferentdiameters(Nudoetal.1992).Afoodpelletwas pairedhand,theyrequiredmorefingerflexionsperre- placedrandomlyintooneofthewells,andtheanimalwasallowed trieval,especiallyonthesmallest well(Fig.3,Cand 0). toretrieveitwiththedigitsofeitherhand.Tentrialsperwell Thisdeficitwas marked inthe firsttwowkafterthe lesion, wereconductedoneachtestingdayandvideotaped.Byanalyzing butwithin2 mo,motorperformancereturnedto preopera- thevideotapesframe-byframe,wenotedforeachtrialthetotal tivelevels.Videotapesofnormalcagebehaviorofthe numberoffingerflexionsandthehandusedtoretrievethepellet threeothermonkeysindicatednodiscernibledeficitsin successfullv. climbingorfeedingwiththecontralateralhand beforethe J finalmappingprocedure. RESULTS Eflectsof focalischemic infarctsonmotorbehavior Eflectsoffocalischemic infarctson adjacentmovement representations LimitedtestingontheKliiverboardintwomonkeys indicatedthatthe infarctresulted indeficitsinskilleduse ofthecontralateralhand(Fig.1).Duringthepostinfarct period,monkeysgenerallyused the contralateralhand less thantheydidbeforetheinfarcttoretrievepellets.How- ever,one monkeypersisted inuse ofthe contralateralhand onthesmallest well(Fig.1A).Althoughmonkeyswere able toretrievefoodpelletsfromeach wellwiththeim- Baseline maps ofmovementrepresentations were similar tothose previouslyreported(e.g.,Gouldetal.1986; Nudo etal.1992;Fig.2C).Thedistalforelimbrepresentation (digit,wrist/forearm)wascontiguousandbounded medi- ally,rostrally,and laterallybyproximalmovement represen- tations(elbowandshoulder),andcaudallybyalargely nonresponsive zone(area3a).Responsive sites inarea 3a wereeliminatedfromstatisticalanalysis. 2146R.J.NUDOANDG.W.MILLIKEN APRE-INFARCT B 3MONTHSPOST-INFARCT ,A#area 250pm area4,A ,,.AGrea3a DISTALDIGITWRIST/ FORELIMBFOREARM FIG.2.Remodelingofmotorrepresentationsinarea4followingafocalischemicinfarct,AandB:photographsofcortical vasculature.Cand19:Intracorticalmicrostimulation(TCMS)-derivedmapsofdistalforelimbmovements.Infarctdestroyed 21%ofdigitand7%ofwrist/forearmrepresentationwithinarea4.Whitedotsindicatemicroelectrodcpenetrations.Area 3a/4borderindicatedbygraydiagonalline,E:arealchangesinmovementrepresentationsurroundinginfarct.Undamaged digitareaadjacenttothelesiondecreasedby40%.L:cresylviolet-stainedcoronalsectionthroughtheinfarctedzone.False sulcus(i);zoneoflowneuronaldensityandhighgliadensityboundedby(...).Scalebar=Imm.Distalforelimb consistsofdigitandwrist/forearm.Proximalconsistsofelbowandshoulder. REORGANIZATIONINMOTORCORTEX2147 CONTROLSPONTANEOUS RECOVERY ndistalforelimb 0 wrist/forearm FIG. 3.Changeindistalforelimbrepresentationsincontrolgroup(n= 4)andspontaneousrecoverygroup(n=4).(*),statisticallysignificant differencesbetweenpre-andpostinfarctmapsbyusingposthocpairedt- tests(P
