Anatomy, physiology and ischemia

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SECOND WORLD CONGRESS OF STROKE, 1992

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TRllNSEScr'HAGEAL E01CCARDICG1APHY rn PATIENTS WITHATRIAL FIBRILLATIOO (!IF) Am ISOWMIC BRAIN DISFASE

G.DRAGONETTI, N.FRATALOCCHI, L.PENNACCHIETTI,A.VESPRINI, G.BELLAGAMBA, *L.FERRINI.GERIATRIC HOSPITAL FERMO (AP), ITALY.*MEDICINE DEPARTMENT MONTEGRANARO HOSPITAL (AP),ITALY

We studied 73 consecutive patients in atrialfibrillation (FA), age 41 to 86 mean age 71,in order to investigate the anatomicalcondition of left atrium and his appendage. 4patients had mitral valve stenosis; 25patients (35%) suffered from ischaemic braindisease: complete stroke, hemiparesis,amaurosis, TIA. We found 4 evident thrombi inleft atrium and 16 thrombi in left atriumappendage, totally 28,5%; 27 patients (37%)presented some fibrousness in left atriumappendage. We didn't found hemodynamicstenosis of epiaortic vessels.

We believe that TEE is very useful tovisualize this pre-embolic condition inorder, first of all, to perform a suitableanticoagulant therapy.

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Dipyridamole-Echocardiography test (DET) in patientswith cerebral ischemia unable to exerciseG. Di Pasquale, A.M.Lusa, S. Urbinati, M. Ruffini, P. Grazi,P. Limoni*, G. Labanti, G.Pinelli. Depts of Cardiology and*Neurosurgery I, Bellaria Hospital, Bologna, Italy

The safety and the effect on the cerebral blood flow of DET inpresence of symptomatic high-grade carotid stenosis (2:70%) wasstudied in 25 pts with stroke (18 M 7 F, mean 52.6 yrs), withoutknown ischemic heart disease (IHD), unable to exercise. Thestudy protocol consisted of a simultaneous 2D-echocardiographyand trans-cranial Doppler (TCD) monitoring from thesymptomatic side after dipyridamole (Dip) i.v. (0.56 mg/kg over4' followed, if DET was normal within 4', by a further dose of0.28 mg/kg over 2'). After DET, aminophylline was given to allpts. DET showed wall motion abnormalities in 9 (36%) (afterhigh dose in 5), with ST segment depression in 2 and angina in1. Dip increased the heart rate (HR) from 70±12 to 83±15 (p<0.001) and slightly reduced the systolic blood pressure (SBP)from 149±23 to 147±20 (ns); the high-dose further increasedHR to 92±15 (p<0.05), and reduced SBP to 144±18 (ns). Minorside effects were observed in 2. TCD detected a reduction of themean middle cerebral artery (MCA) flow velocity respect to theresting value in all pts (p<0.OO5), while pulsatility index wasunchanged. Similar changes were observed in control pts. Inabsence of significant reduction of SBP these data may suggestthat Dip causes a dilation of MCA.In conclusion, DET identifies IHD in over 1/3 of pts with strokeunable to exercise. Dip i.v. reduces the MCA flow velocitywithout symptoms or significant reduction of SBP. Our datasuggest that DET is a safe and reliable tool in the noninvasivescreening for IHD in pts with symptomatic high-grade carotidstenosis.

CONTRIBUTION OF TRANSESOPHAGEAl ECHOCAROIOGRAPHY TO ETIOlOGIC DIAGNOSIS OF ISCHEMIC STROKE. -

F.FEOERICO,C.CONTE,C.CAIATI,N.di T,!!

RI, S.ILICETO, P.lANBERTI.

INSTITUTE OF NEUROLOGY, UNIVERSITY OF BARI, PIAZZA GIULIO CESARE

70124 BARI - ITALIA.

lie included 63 consocutive hospitalized patients vith recent

isthe.it stroke. All pts undervent cranial TC, haeutothe.ital

and Ins truaent al investigation (EKG,RX.Chest,Duphx-Stanner).

On the base of so.e par-aaeter-s (age, onset, risk fat tors for

atherosclerosis, previous car-di ae disease, atrial fibrillation,

size and Iccat i en of TC hsions, Duplex-Stanner findings) ve

supposed a diagnosis of atherothro.boe.bolic stroke (ATE),

..bolit tardiogenit stroke (EC) and stroke of untertain

classifitation (UC).

Distribution of pts vas as following:39 pts (62%) in the ATE

group, 16 pts (24%) in the UC group and g pts (14%) in the EC

group.

Transthoracic ochotardiography (HE) and transesophageal

ochotardiography (TEE) were perfor.ed within two weeks of the

onset of the sy.pto.s. Considering ochoradiographit results we

tan allocate patients as following: 35 pts (55%) in the ATE

group, 20 pts (32%) in the EC group and B ph (13%) in the UC

group. Our results show Tee (26 pts positive) is .ore sensitive

than HE (s pts positive) in the detettion of potential car-di ae

sources of ..boli.. ; furtheroore TEE allows to clarify the

pathogenesis of stroke.

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ISCHEMIA·INDUCED GLUTAMATE RELEASE: EFFECTS OFGM-1 GANGLIOSIDE AND OF ANAESTHESIA.

G. Lombardi, G. Cherici and F. MoroniDepartment of Pharmacology, University of Florence,Viale Morgagni65, 50134 Firenze

The effects of GM1 ganglioside treatment were studied on theischemia-induced release of aspartate (ASP) and glutamate (GLU)in chloralhydrateanaesthetizedgerbilshavingtransversemicrodialysistubing in their hippocampi. Forebrain ischemia was obtained 2 hafter cannula implantationby occluding both commoncarotid arteriesfor 8 min with aneurism clips. Body temperature was controlled at37.4 C. Ischemia increased GLU and ASP concentrations in thedialysate by 3 to 8 fold and the increases lasted 20-30 min. Whenthe animalswere pretreated withGM1-ganglioside (30 mgJkgJdayi.p.for 3 days) the ischemia-induced increases of excitatory amino acidsin the dialysate were significantly reduced (area under the curve incontrols: 14!. 2 and 92 !. 16 for ASP and GLU, respectively, andin GMI treated animals: 8!. 15 and 39 1 33 nmol/ml/h).

In separate experiments increases in the extent and the time courseof the ischemia-induced ASP and GLU concentrations were studiedin hippocampal dialysate of anaesthetized or freely moving gerbilshaving chronically implanted microdialysis tubing in the hippocampus.Ischemia was performed according to Clough-Helfman and Phillis,Free Radical Res. Comms. 15; 171; 1991. In these gerbils, ischemiacaused not only a 3 to 5 fold increase of GLU output similar to thatobserved in anaesthetized animals, but also a second (starting 1 hafter the ischemic episode), long lasting (at least 3 h) increase ofGLU output. The pathogenetic meaning of this second peak in thedevelopment of the delayed neuronal degeneration, is now underinvestigation.

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THE VARIABILITY OF THE VASCULAR TERRITORIES OFTHE MAJOR CEREBRAL ARTERIES OF THE HUMANBRAIN

A. van der Zwan, B.HiIlen, C.A.F. Tulleken.The Netherlands'M. Dujovny, U.S.A. '

In the literature the territorial distribution of the six major cerebralarteries is generally considered as more or less invariant. Inneuroradiological practice, this assumpt ion is also used in thedifferential diagnosis of thrombo-embol ic and watershedischemia. However, recent mathematical model studies and areview of the literature indicated that this assumption isdisputable I,2. These findings demanded for detailed quantitativedata of the territorial distibution of these arteries. We thereforesimultaneously injected the six major cerebral arteries of 25unfixed human brains with differently colored plastic. Theinjection pressure was identical for all arteries and withinphysiological range. The conical and intracerebral territories werestudied by cutting the brains in 6 mm sections . In addition, thevolumes of the separate territories were calculated using graphictablet and Cavalieri's method. From this study the followingconclusions can be drawn. First, the territorial distribution of themajor cerebral arteries are considerably variable not only betweenindividuals but also between the individual right and lefthemisphere. Second, there is a sign ificant correlation betweenthe, also highly variable proximal diameter of the artery, and thevolume of its territory. This indicates that a hemodynamic relationexi sts between the variation of the circle of Willis and thevariation of the hemispheric territorial distribution.References:1. Hillen et al. A mathematical model of the flow in the circle ofWillis . J. Biomech., 19, 1986.2. A. van der Zwan and B. Hillen. A review of the variability ofthe territories of the major cerebral arteries.Stroke, 22, 1991.

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Effect of Hyper-,Normo-and Hypoglycemia onCerebra.l ISchemia in RatsCao Bingzhen,Guo Shusu.Zhao Yuwu.Xue Shenwu

China

We investigated the effeot of nvper- normoand hypoglycemia on Wistar rats unaergo!ng 24hours uf occlusion of both common carotid arteries.In another consecutive experiment, 4rats in each group undergoing 30 minutes ofischemia were used to obServe the ohange ofultrastructure and to measure the mitochondrial change in the neurone quantatetibelY byTexture Analysis.In the 0.2 unit inSUlintreated group the rat mortarity,neurologicaldeficit score and brain water content wereSignificantly than those in normo-and hyperglycemic gro~p.No rats died when preocclusionplasama glucose concentration wqs between 100-170mg/dl(aDout 1/2-2/3 of nbvmal value inrats). 24 hours after ischemia there waS aevident hypoperfusion in hyperglycemic group.Morphology study demonstrated hyperglycemiaexcerbated the mitochondrial damage.We concl~

uded that hyperglycemia may increase ischemicbrain damage. InSU.lin can be used to maintainmild hypoglycemia,which is aSSOciated withdecreased behavioral neurologic deficit andincreased survival following cerebral ischemia .Hyperglycemia or more extreme hypoglycemia(less than 100mg/dl) is detrimental in thismodel.

Striatal cell grarts in ischemic region folio" ing intraluminalocclusion of middle cerebral art ery ameliorate cytoarchitectual,chemical and behavioral damages (Part 2)N.Aihara. K.~lizuka\\ al, K. Kolde.H, Mabe, II. Nagai, II. Nishin02Dept. Neuro surgery and 2physiol ogy, Nagoya City University Med icalSchool, IDepanm ent of Anatomy, Okayarna University. Japan

Few useful treatment for brain damage due to ischemic insult. especially

at ch ronic stage, have been establi shed. We implanted cell suspe nsions of

fetal striatum into ischemic regions cau sed by intraluminal occlusion of

middle cere bral a rtery (MCA) in the rat. and investi gated the

survi val/devel opment of graft ed cells and functional recovery. One hour

intraluminal occlusion of MCA induced ischemic damages in the lateral part of

the striatum, and adjacent cortex.Is chernic animal had deficit' in MOrTh water

maze learning . In these anima" GABA level in the globus pallidus, detected

by microdialysis, "as decreased to about 50Ck of that of controls. Following

the graftin g. the learning was improved ~nd GABA level was increased. In

lhe grafts, muscarinic, 01, D2 and GAB/\A receptors, detected by 3H labeled

Q:\B. SCH23390. spiperone and SR95531 binding respectively, "ere "ell

developed. Data suggest that fetal striatal cell grafting remod els the ischemic

brain and arnellorates damaged function.

ro-esTPA and Ischemia in Rabbits.

Bowes, M. P. and Zivin, J. A.U. S.

Endothelial adherence of leukocytes may impeder ep erfusion following thrombolytic therapy andmay a l s o participate directly in ischemic CNS' i n j ur y , perhaps ~ausing reperfusion injury. Weevaluated the effect of antibodies directedaga i ns t l eukocyte adhesion (Intracellular Adhesion Mol ec ul e- l ; ICMI-l) in a model of CNS ischemiafollowed by thrombolysis with tissue plasminogenactivator (tPA). Ischemia was induced by injecting a suspension of microclots into carotidcirculation of New Zealand white rabbits. The~nimals wer e randomly assigned to one of fourco nditions; control (n=18), anti-ICMI (five ma f t er ischemia; n=13), tPA (30 m after ischemia;n=23) an d the combination of anti-IC~~ and tPA(n=15) • . Ant i - I CMI and tPA each s i gni f i cant l yincreased the amount of clot necessary to produceneurological damaqe (p <.05). Corrbinedtherapy was no more effective than e i the r substance a l one . A similar outcome was obtainedwhen tPA was delayed until 90 m po st ischemia.Thus, although the anti-ICMI antibody effectively reduced neurological da mage, as did tPAalone, no interaction waS detected.

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INFLUENCE OF NEUROTROPIN ON THE WATER CONTENT AND THETRACE ELEHENT DISTRIBUTION IN RECENT CEREBRAL INFARCTS.Jacques L. De Reuck , Geert H. Hebbrecht, \~illy

Haenhaut.Ghent, Belgium.

Neurotropin is a non-protein extract of cutaneoustissue from rabbit, previously inoculated with Vaccina virus, that inhibits t.he release of bradykinin.In a randomized double blind controlled clinicaltrial Neurotropin was found to be effective in treating brain oedema, due to acute ischemic stroke.Postmortem examination of the brain was obtained in5 Neurotropin and 6 placebo treated patients, whodied within 10 days following the stroke. All patients had an infarct in the territory of the middlecerebral artery. Particle-induced X-ray emission wasperformed on samples of the infarcts and their borde~

zones, allowing the measurement of the concentrationof 8 elements (K, Ca, Hn, Fe, Cu, Zn, Se and Rb).Also the reduction factor or wet-to-dry weight ratiowas determined. The most s~riking finding was aless significant increase of the reduction factor inthe white matter of the brain infarcts and their borderzones of the Neurotropin compared to the placebotreated patients. As a whole the Ca concentrationwas significantly less raised and Rb less decreasedin the infarcts of the Neurotropin than of the placebo treated group. Also in the borderzones of theprior group K was less decreased and Zn and Se moreincreased. This postmortem study demonstrates thatNeurotropin reduces significantly brain oedema inthe white matter and probably also ischemic damageafter acute stroke.

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TITLE SECONDARY PREVENl10N OF CEREBRAL JSCBEMICEVDITS wrm MFSOGUCANO: A CONI'ROllED,MULTICENTRlCSI1JDY(S.I.A.M.)

AUTHORS AND COUNTRY

S. Forconi for the "Gruppo Senese CerebropatieVascolari*, University ofSiena, Italy

The aim of this study (S.LA.M., StudioItaliano Asa-Mesoglicano) was to evaluate, inpatients with recent episodes of TIA, RIND, orICtus of atherothrombotic origin, the effect oftreatment with mesoglicano (100 mg/die) oraspirin (300 mgldie) in reducing the risk of newcerebral ischemic events and of vascularmortality.

A total of 1398 patients were included inthe study and the randomization proceduresresulted in two groups of patients, one treatedwith mesoglicano and the other with aspirin,well balanced for number, sex, age, vascularanamnesisand type of events at enrollment. Wepresent the results after one year of follow-up,in which vascular mortality and cerebrovascularevents were equivalent in the two groups oftreatment.

The results are discussed in view of thedifferent mechanisms of action of the drugsstudied.

Neuroprotectlve action of carvedilol, a newvasodllator/(3-adrenerglc blocker In vitro and Invivo.Giora Feuerstein, Paul G. Lysko, Kathryn A. Lysko andTian-Li Vue. SmithKline Beecham Pharmaceuticals,King of Prussia, PA 19406 USA

We have recently shown that carvedilol, a newvasodllator/p-blocker antihypertensive, possessesantioxidant and anti-lipid peroxidation (LPO) propertiesin swine cardiac membrane. Since oxygen radicals areimportant in neuroinjury following ischemia, weexamined carvedilo\'s anti-LPO and neuroprotectiveefficacy in the following models: 1) rat brainhomogenates in vitro, exposed to Fe 2+Nit C (·OHradical generation system): carvedilol dosedependently inhibited LPO (thiobarbituric acid reactivesubstances) with an 1Cso=8p.M (n=8); 2) rat cerebellargranule cells in culture exposed to glutamate or DHFFe 3+/ADP: carvedilol blocked neuronal death dosedependently at ICso= 1.1 p.M and 5p.M, respectively(p<0..001, n=6-8). In both in vitro systems, none of thetraditional (3-blockers such as propranolol, labetalol orceliprofol showed any activity; 3) in gerbils exposed toglobal brain ischemia (6 min) and reperfusion (7 days):carvedilol (3 mg/kg, s.c., BID, pre- and post-ischemia)increased the survival rate of CA1 hippocampal neuronsfrom 37±8 to 78±13 (p<0.01, n=22). Taken together,these studies strongly suggest that this newantihypertensive drug might also be effective inprevention or treatment of stroke.

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Beta-endorphin Progressively Accumulates Near SmallEmbolic Cerebral Infarcts in the Rat

Leszek S. Dujanowicz and Nancy Futrell, U.S.A.

Hacrophages enter cerebral infarcts, beginning at 23 days and becoming abundant by 5 days; triggeringfactors for this macrophage activity are not known.Beta-endorphin is a candidate, as it is present inthe brain and stimulates macrophages. We studied 22Fisher rats, 2-4 months of age, 20 with emboliccerebral infarcts produced by irradiating the rightcarotid artery with a laser (632 nm, 200 mW/sq em,15 minutes) following the injection of a photosensitizing dye, and 2 controls (1 dye only and 1 normalcontrol). There were 114 infarcts studied at I, 2,3, 4, and 5 days (4 animals per group). Betaendorphin was identified with immunoperoxidasestaining using a polyclonal antibody. Beta-endorphinwas present in the brain stem bilaterally in allanimals. Traces of endorphin were seen near 3/25infarcts at 1 day, 7/21 infarcts at 2 days and 10/24infarcts at 3 days. There was a moderate amount ofendorphin near 14/18 infarcts at 4 days, withabundant staining near 26/26 infarcts at 5 days. At5 days there was diffuse, punctate endorphinstaining in the cerebral peduncle, internal capsuleand external capsule ipsilateral to the infarcts.We conclude that beta-endorphin does accumulate nearcerebral infarcts; the time course parallels that ofmacrophage activation. The pattern of accumulationin white matter tracts suggests it may reach theinfarcts via axonal transport from the brain stem.

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Hypothermia Reduces The Extent of Cerebral InfarctionfolloWing Focal Ischemia In Sprague-oawley RatYasunobu Goto, Ken-ichiro H iramatsu, Scott Soleau, NealF. Kassell, Kevin S. Lee.Department of Neurological Surgery, University of Virginia,Charlottesville, VA 22908, USA

Considerable evidence suggests that changes in braintemperature during ischemia affect the extent anddistribution of ischemic injury. We examined the effects ofbrain cooling during reversible, focal brain ischemia on theextent of cerebral infarction in adult, Sprague-Dawley rats.Under halothane anesthesia, the left middle cerebral artery(MCA) and both carotid arteries were temporarily occludedfor three hours. Animals were sacrificed three days later.Brain sections were stained with triphenyltetrazoliumchloride and analyzed using a computerized imageanalysis system. Temporal muscle temperature (TMl) andrectal temperature were mon itored continuously . ThefollOWing three groups with different intra-ischemic TMTswere analyzed : a) control , ' 35 .8-38.2°C, b) mildhypothermia, 33.0-33 .5°C and, c) deep hypothermia, 27.529.2·C. Each group consisted of six animals. The volumesof infarction were 219 .1±21.1, 170 .3±6.8 and 108 .2±5.9

. (mean±SEM) for the control, m ild hypothermia and deephypothermia groups, respectively. These findingsdemonstrate that hypothermia reduces the impact of focalischemia in Sprague-Dawley rats.

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Acute Distention of the Basilar Artery I mpairsEndothelium Dependant Relaxa tion: I mplicationsfor the Pathogenesis of Cerebral Vasospasm

Dan S. Heffez, Chia ~ang, Jonathan Becker,Chicago Institute for NeuroSurgery andNeuroResearch, Chicago, Illinois USA

~e hypothesized that acute systemic hypertensionwhich accompanies ane urysmal s ubarachnoi dhemorrhage may damage the endothelium of cerebralarteries i mpairing vasodilatation and predisposing to cerebral vasospasm.Dog basilar arterieswere perfused in v i tro with heparinized autologous blood to establish an intraluminalpr essure between 180-195 c:m IIg for 15 mi nu t es.Vessel rings, mounted in organ baths, wereconstricted using 10-6M serotonin. Cumulative logdose response curves were determined for acetylcholine, adenosine, papaverine and sodium nitroprusside.Acetylcholine, an endothelium dependantva sodilator, was less effective in experimental(n-12) than control, non-perfused (n-14)ve ssels, (p<O.0005,1-tailcd t-test for unpairedobservations). Papaverine and adenosine , va sodilators that interact with smooth muscl emembrane bound receptors,were also less effectivein experimental vessels(p<O .0005).Sodium nitroprusside , which acts directly at the subcellularlevel to reduce smooth muscle tone, was equallyeffective in both vessel groups . The mechanism ineffect likely involves endothelial injury, andperhaps altered activity of smooth muscle membrane bound receptors. The smooth muscle itselfretains the ability to relax suggesting a molecular mechanism for the treatment of vasospasm.

SELECTIVE VULNERABIUTY OFTHE HINDBRAIN STRUCTURES TOISCHEMIA IN GERBILS"R. Hata, M. Matsumoto, M. Tagaya, T. Ohtsuki, S. Ogawa, H. ueca, N.Handa, "T. Nishimura, T. Kamada, tst Dept. of Internal Medicine and"Biomedical Research Center, Osaka University Medical School,Osaka,Japan

Selectivevulnerabilityofvariousforebrain regionsto Ischemia has beenstudied extensively in the past two decades. But, as for the hindbrain,selective vulnerability has not been elucidated completely. Lately, wehave successfully established hindbrain ischemiamodelingerbilsby theextracranial oedusion of bilateral vertebral artery (1). In the presentstudy, by using this model, differential vulnerability of the hindbrainstructures to ischemia was thoroughly investigated. Adult Mongoliangerbilsof both sexes , weighing 50-BOg, were used. The sutures aroundeach vertebral artery were pulledby59 weightsto oedude the circulation(1). Altera predeterminedischemic period, gerbilswere decapitated andtheirbrainswere processed forthe immunostaining. serial brainsectionswere examined every 200 11m with immunostaining for microtubuleassociated protein 2 (MAP2) , which was thoughtas a sensitive markerprotein for detecting early ischemic damage (2). Immunostaining forMAP2 disclosed the following results.

Brain area Sham Ischemic period5 min 10 min 15 min 30 min

Oculomotor n. (nlll) 0 25 50 100 100Trigeminal motor n. (MoV) 0 0 0 25 100Cerebellar interposed n, (IP) 0 100 100 100 100Lateral vestibular n. (LVe) 0 100 100 100 100Hypoglossal n. (nXII) 0 0 0 0 50

The results were expressed as a percentage of abnonnalities, basedon four gerbils for each time interval. These results demonstrated theprogression of early ischemic lesions in the hindbrain regions and LVeand IP may be most vulnerable to Ischemia.References: (1) J Cereb Blood Flow Metab 11: 5535, 1991

(2) Neuroscience 31: 401-411,1989

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ACUTE CEREBRAL ANEMIA DUE TO STRAIN

Minoru Tomita, Fumio Gotoh, Yasuo Fukuuchi, TakahiroAmano, Norio Tanahashi, and Kortaro TanakaDepartment of Neurology, School of Med icine, Keio Univ.Tokyo, Japan

The effect of strain on the cerebral hemodynamic changes wasinvestigated using our pho toelectric method (Tomita et a1.:Am. J. Physio!. 235:H56-H63, 1978) by which time-to-timechanges in cerebral blood volume (CBV) and cerebral bloodflow (CBP) could be measured in experimental animals. Fivecats were anesthetized with alpha-chloralose and urethane, andventilated artificially. Continuous recording of CBV, asindicated by the red blood cell volume in the cerebral cortex(ectosylvian gyrus) by the photoelectric apparatus was carriedout. CBP was calculated from CBV and meantransit time ofcarbon black injected into the carotid artery. Simultaneousmeasurements were made of the intracranial pressure (ICP)through an epidural catheter. The systemic mean arterial bloodpressure (MABP) was also monitored via a femoral catheter.Strain was j mposed on each cat for approximately 30 s by theValsalva maneuver in which the intrathoracic pressure wasinflated by connecting the tracheal tube to a 10 liter plasticboule with compressed air at 40 mmHg above atmosphericpressure. Iii 2 cases, EEG was monitored. We found with theValsalva maneuver that: 1) MABP was decreased in all casesfrom 144.5±9.7 to 58.6±6.8 mmHg (Mean±S .D.); 2) CBP wasdecreased from 78.4±3.? to 23.2 ±2.1 ml/lOOg.min; 3) ICPwas increased by 33.2±3.5 mmHg; 4) CBV decreased by3.2±1.2 vol% in parallel to the SABP decrease; and 5) EEGtended to slow down. We conclude that strain causes cerebralanemia, not cerebral congestion.

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ROLE OF ENDOTHELIUM-DERIVED RELAXING FACTOR IN CHEMICALCONTROL OF FELINE CEREBRAL CIRCULATIONTet suya l naIuku. Takashi Rata. Funi hi ko Sakai. TadashiKanda. Yoshiakl TazakiKltasato University. Kana gava, Japan

To study the role of endothelium-derived relaxingfactor(EDRF) on the chemical control of feline cerebralcirculation. we examined C02 reactivity of CBF beforeand after the administration of a specific inhibitor ofnitric oxide(NO) synthesis. NG-monomethyl-L-Arginine(LNMIlA). and its Inactive optical isomer. NG-monomethylD-Arginine(D-NMMA). Twenty adul t cats were preparedfor the experiments. Regional CBF in the left thalamusand left deep frontal white matter were measured byhydrogen clearance method and changes in CBF In theright thalamus and right frontal cortex were monitoredby heat clearance method and by the laser Doppler flowmeter continuously. In 15 animals resting CBr was 53.2ml/min/l00g brain In the thalamus and 22.6ml in thewhite matter. During continuous l nt r aca ro t i df l er t) ada i nl s t r at lon of L-NMIlA (0.07# mol/kg/min). resting CBFdecreased to 43.2ml and 18. 8ml respec t ive ly. And thesedecrement In CBF were restored by L-arginine(LA) (0.4#mol/kg/l:lin)(H.OmI,20.5m)). C02 reactivity expressedby %change/mmHg. reduced by L-NM\lA from 5.20 to 3.91(thalamus) and from 2.60 to 1.86(white matted. Additional adminislration of L-A restored C02 reactivity toL 82 and 2.50 respect i ve l r. Ilowever. D-NMMA (in 5animals) changed neither resting CBF values nor C02reactivity (from 2.05 to 2.48;thalamus. from 3.07 to2. 93;white matter>. Our data suggest that EDRr/NO mayplay some role not only in the control of resting CBFbut In the dilatory respo~se to hypercapnea.

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WITHDRAWN

Reactivity of the cerebral microvessels associated withextracranial arteriovenous fistula in lhe rat.

K.IRIKURA, S. MORII, K. TOKIWA, Y. MIYASAKAand K. YADADepartment of Neurosurgery, Kitasato University,School of Medicine, Sagamihara, Kanagawa, JAPAN

The breakthrough phenomenon of the cerebral blood nowautoregulation has been implicated with acute deterioration afterresection of arteriovenous malformation. In order to validatethis hypothesis, the chronic effects of extracranial arteriovenousfistula on morphological changes and reactivity of the cerebralmicrovessels were investigated in the rat.

A right carotid-jugular fistula was created in Sprague-Dawleyrats as described by Morgan et al, Eight to 12 weeks after acarotid-jugular fistula was created, the pial rnicrovessels wereobserved in situ using a closed cranial window technique. Thediameter of the pial arterioles was continuously measured by avideo dimension analyzer. Initially, effects of acute closure ofthe fistula on the diameter of arterioles under various levels ofmean arterial pressure were studied. Secondary, theautoregulatory response of arterioles to hypertension inducedby intravenous norepinephrine was investigated. Themorphology and response of the vessels were compared withthose of control rats.

The carotid-jugular fistula produced marked dilatation andtortuosity in the pial arterioles. The se abnormal arteriolescontracted by fistula closure when the systemic arterialpressure was within normal range. In contrast. theysignificantly dilated during hypertension. [t is concludedthat the cerebrovascular response to hypertension wasimpaired associated with arteriovenous fistula in the rat.Therefore, acute closure of the fistulae can be responsible forcerebral hyperemia.

Determination of serum sarcoplasmic proteins aftercerebrovascular diseaseTakahiro Jimi. Yoshihiro Wakayama. I1iroyuki Okayasu,Seiji Shibuya. 'Kanefusa Kato Div of Neurol. Deptof Med, Showa Univ Fujigaoka Hosp, Yokohama and'Deptof Biochem. Inst of Developmental Res. AichiPrefectural Colony. Aichi. Japan

Purpose: To evaluate the clinical significance ofconsecutive determinations of sarcoplasmic proteinsin acute stroke. we measured serum creatine kinase (CK). myoglobin. aldolase. muscle specific enolase (MSE)carbonic anhydrase'll. (CA:L) and S100& protein (SlOOa,).Subjects and Methods: We took time-sequential samplesof serum from 24 patients (16 men.8 women) with acutestroke within 48 hours. at 1 week. 2 weeks and 4weeks after the ictus. We measured MSE. CA!l andS100a, using a sensitive sandwich enzyme immunoassay.Results : Serum concentration of myoglobin. CAli! andS100a, sampled within 48 hours showed significantrise compared with those of control group (4.5 times.5.2 times. and 12.5 times higher than controls,respectively). CK and MSE values are. on the otherhand, lower than those of control group. S100a,showed higher values during 4 weekS in most strokecases. Concentration of CK. myoglobin. MSE and S1008<demonstrated decline with time. Aldolase. however.peaked 2 weeks after the ictus and then declined.Prognosis of 2 patients with extreme initial increaseof m:(oglobin. CM and S100a,- were fatal. OverallclinIcal severity and changes in concentration ofthese 6 proteins. however. did not show statisticallysignificant correlation .Conclusion: This study indicated the patterns oftransient release of muscle sarcoplasmic proteins arevariable after acute stroke. S100a. showed most sensitive changes among these 6 proteins. Initial extreme increase in myoglobin. CA~ and S100a~ mayreflect generalized severe clinical condition andsuggested poor clinical outcome.

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Istablishment of the stroke-pronerenovascular hypertensive rats

Zena Jins~e"3' HtJM3

R"rU>1. P.I<. Ch:>t....

After the renal arteries were constrictedbilaterally with silver clips ~ the renovascularhypertension was produced in all 55 operatedSprague-Dawley rats .The highest blood pressurein the renovascular hypertensive rats (RHR)remained over 26.7 kPa. Febrinoid necrosis, hyalinosis and formation of microaneurysm wereobsereved in the cerebral arterioles or smallarter ies . The spontaneous incidence of strokewas 66.4%, when the RHR were fed with a standardrat chow diet (23% protein) .The type of strokeincluded cerebral haemorrhage, cerebral infarct ion, subar achno id haemorr ha/ge and mixe d stroke.Comparing with stroke-prone spontaneouslyhypertensive rus (SHRSP), the RHR are free fromgenetic deficiency, easy to be raised and canbe supplied for studies in a large number,therefore the rolR can be used as stroke-pronerolR.

PO-C20

Effect of a new Ca-t+antagonist(SM-6586) on experimental cerebral ischemia*Kashiwagi F, Katayama Y, Igarashi H, Iida S, HuramatsuHand Terashi A. The second department of internalmedicine, Nippon Hedical School, Japan *KouseiHospital

SH-6586(SM) is a derivative of dihydropiridinewith a potent Ca-t+blocking activity and inhibitoryactivity of the Na+/H+ and Na+/Ca-t+ exchangetransport. He have evaluated the effect of SH onsurvival rate, brain edema and metabolites using twodifferent ischemic models in hypertensive rat(SHR).Group I, underwent bilateral carotid artery ligation,received 15 and 50 pg/kg of SH at 30 min prior to andimmediately after ischemia. The survival rate wasobserved until 6 hrs, and the brain water content andmetabolites were determined 3 hrs after ischemia.Group II, underwent }~A occlusion, received 150 pg/kgof SH at 30 min prior and immediately afterocclusion, and 300 pg/kg at 23 hrs after ischemia. Tlrelaxation time and water content were investigatedat 24 hrs of ischemia using MRI(6.34T). Group I:SH-treated group showed higher survival rat~. anddecreased brain water content. The agent alsomaintained the concentrations of ATP at higherlevels, and reduced accumulation of lactate and L/Pratio. Group II: SH-treated group showed reductionof Tl relaxation time as well as suppression of watercontent. These results indicated that SM lessenedmortarity, and ameliorated brain edema and metabolismin ischemia. It is suggested that SH could beeffective in acutely induced cerebral ischemia.

Immunohistochemical study on the expression of cytoskeletat proteinsand proliferation in reactive astrocytes following focal cerebralischemia

HidekiKanai, Hajime Nagai. Hideo Mabe. Naoki ShimazuDepartment of Neurosurgery. Nagoya City University Medical School,Japan

In the repair process of cerebral infarct. reactive gliosis ocurrs, inwhich the increase of intermediate filaments within astrocytes is the.most prominent alteration. Astrocytic reaction following thedegeneration of neurons is also observed in the thalamus andsubstantia nigra which are not primarily involved in ischemia aftermiddle cerebral artery occlusion. little is known as to differences ofthe expression of cytoskeletal proteins in reactive astrocytes betweenin the ischemic and non-ischemic remote areas. To investigate suchdifferences, the present study was designed in rats subjected to 2hour of middle cerebral artery occlusion followed by reperfusion from 1to 30 days. We evaluated GFAP, vimentin,tubulin(a ,psubunit) andMAp·2 in astrocytes immunohistochemically and identifiedproliferating cells by BrdU and proliferating cell nuclear antigenimmunostaining. Proliferating astrocytes with hypertrophied cytoplasmaround the infarct, especially in white mailer stained with GFAP.vimentin, tubulin and MAP·2 antibodies 2 days after the ischemia.Tubulin and MAP-2 immunoreactivities started to disappear after 14days when the number of proliferating astrocytes reduced remarkably.In the ventroposterior nucleus of the thalamus and substantia nigraipsilateral to the infarct. GFAP·positive astrocytes stained withvimentin antibody weakly and did not in tubulin and MAP-2 from 7days after the ischemia. Proliferation of astrocytes was not detected inthese remote areas at any time point investigated. We conclude thatreactive astrocytes proliferating around the infarct express variouscytoskeletal proteins more strongly than those in remote areas whereno mitosis occur.

Po-e21

The sequential change of CBF and pathology in atransient focal ischemic model.Toshiya Katsumata, Rinnosuke Obo, Hiromi Huramatsu,Toshiyuki Soeda, Yasuo Katayama, Akiro Terashi, TheSecond Department of Internal Hedicine Nippon HedicalSchool, Tokyo Japan.

He investigated local cerebral blood flow(CBF)and pathology sequentially following 2hrs oftransient ischemia to determine whether a temporalthreshold of ischemic brain damage exists around 2hrsin focal ischemia. The focal ischemia was induced bymiddle cerebral artery occlusion (Koizumi' s method)in S-D rats. CBF was measured at 2hrs of ischemia andat reperfusion of 30min, 2hrs, 24hrs, 7 days after2hrs ischemia using [l~Cliodoantipyrine quantitativeautoradiographic technique. Brain sections werestained with hematoxylin and eosin. After 2hrsischemia, CBF was more decreased in the parietalcortex(P-cor) and the lateral caudo-putamen(L-CP), 8%and 11% of the control when compared to otherregions. The CBF of frontal cortex(F-cor) which wasborderzone area, decreased to 31% of the control. Atreperfusion of 30min and 2hrs, CBF was various in theP-cor and L-CP. At 24hrs, CBF recovered to57%(P-cor), 88%(L-CP), 80%(F-cor) of the control andspongy changes were observed in the P-cor and L-CP.Although CBF was maintained 46%(P-cor), 76%(L-CP),61%(F-cor) at 7 days, the necrosis was observed inthe P-cor and L-CP. He clarified that P-cor and L-CPwere core areas in the ischemic model and also thetemporal threshold might exist before 2hrs ofischemia.

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PO-C23

S89

Effect of Allopurinol on Cerebral ischemia-- Experimental study using SHR and clinical

efficacy in acute cerebral infarctionH.Kawasaki, S.Tochigi and T.ItohThird Department of Internal Hedicine,St. Harianna University School of Hedicine,Kawasaki ,Japan

Using spontaneous hypertensive rats (SHR), we investigated the effect of Allopurinol (AP), a xanthineoxidase inhibitor, on the progress of cerebral injurysubsequent to the reperfusion of ~lood flow followingcerebral ischemia. The clinical efficacy of AP onacute cerebral infarction (ACI) was also studied withfocus on the change in CT findings.

We previously reported that AP may be effective inpreventing the progress of ischemic cerebral injury,and its inhibition of xanthine oxidase induced freeradical produced upon resumption of blood flow afterischemia is indicated as the mechanism of its action.Based on these results, we conducted intravenous drip,infusion of AP at a dose of 200 mg!day for a week on10 patients with ACI to investigate neurological out'come, cerebral lesions on CT and time course change ofcerebral edema. Though this is__ a preliminary study,the results suggested that AP can suppress the progress of cerebral edema and alleviate deterioration ofneurological symptoms in ACI patients.

The following are suggested as the effects of AP onCT findings: 1) Inhibitory effect on cerebral edema;2) Appearance of fogging effect at an early stage (anincrease in cerebral blood flow due to improvement inenergy failure?); 3) A decrease in stroke volume. Sucheffects of AP are more remarkable in the patients withpenetrating artery infarction than in those with cortical artery infarction,. suggesting that AP might actdifferently depending on the site of infarction.

PO-C24

Orthostatic Hypotension dose not Participate toPeriventricular Hyperintensity in Normal AgedHiromi Koide. Shotai Kobayashi. Nobuo Suyama.Hirokazu Bokura , Kaoru Imaoka, Yuko KawamiraShimane Medical University. Izumo. 693. JAPAN

To investigate whether orthostatic hypotensiondose participate to leukoaraiosis or not. westudied the relationship between changes in BP bypostural change and leukoaraiosis in 100 neurologically normal aged (60-88. mean 71 years).Subjects were divided into 3 groups (none. mild.moderate) according to severity of periventricularhyperintensity (PVH) on O.lST MRI. Cerebral bloodflow (CBF) was measured by 133Xe inhalation method.Supine and standing BP and power spectral analysisof R-R interval (PSA) were examined.Results:Severityof PVH paralleled with aging (p<O.OOOl). Cerebrovascular resistance was higher in moderate PVH groupthan those without (p<0.02). Systolic BP (SBP)related to severity of PVH (p<0.0003). Posturalchanges in diastolic BP was larger in the groupwithout PVH than in mild and moderate group(p<0.002). Incidence of orthostatic hypotension(~SBP<-20) did not differ aIIlOng 3 groups. Midfrequency band of PSA which reflects the sympatheticefferent activation in standing was lower in mildand IIlOderate group than those without (p<0.03).Conclusion: In neurologically normal aged. ortho·static hypotension dose not involve PVH. anddecrease of sympathetic activation with aging mighthave some relation to leukoaraiosis.

ACUTE CEREBRAL ISCHEMIA DECREASF.8 GASTRIC MUCOSALBLOOD FLOW IN SPONTANEOUSLY HYPERTENSIVE RATSK. Kawakubo, S. Ibayashi. M. Iida. S. Sadoshima andM. FujishimaThe Second Department of Internal Nedicine, Facultyof Nedicine. Kyushu University, Fukuoka. Japan

Cerebral ischemia is known to cause highincidence of acute gastric lesions. To clarify themechanism(s). we investigated the effect of acutecerebral ischemia on gastric mucosal blood flow(G:-IBF) in spontaneously hypertensive rats(SHR).Methods: Eleven female SHR (5-7 months old) wereanesthetized with amobarbital and stomach was gentlypulled out through the incision of abdominal wall. Aclip-type contact-electrode was inserted into corpusand G~BF was measured by hydrogen gas clearancemethod. Six rats were subjected to I-hour cerebralischemia induced by bilateral carotid arteryligation (BCL) followed by 1 hour of recirculation.Five rats were served as non-ischemic control. Inour previous study. cerebral blood flow decreased toabout 10% of basal value during 1 hour of BCL withslight hyperemia thereafter.Results: (Mean + Standard Error)--- - MABP (mmlIg) %G~IBF (vs.Basal)

Group N Basal BCL60' Recire BCL60' RecircISCHEMIA 6 156+3 163+3 153+5 54.!.9* 26.!.5*COXTROL 5 151+9 152+9 157+6 102+9 83+6·significantly dIfferent from respective control andbasal value (P<O.Ol, t-test & ANOVA)Conclusion: G~BF markedly decreased during cerebralischemia and to the. further extent afterrecirculation. Gastric lesions in acute stroke mightbe attributed. in part, to gastric mucosal ischemia.

PO-C25

Striatal cell grafts in ischemic region folio" ing intraluminal

occulusion of middle cerebral artery ameliorate cytoarchltectual,

chemical and behavioral damages ( Part I )

K. Koide, H. Nagai, II. Mabe, T. Hashitani", N. Aihara and II. Nishino"

Dept. Neurosurgery and 'Physiology, Nagoya City Univ, Medical School, Japan

Cell survival and behavioral recovery after grafting of fetal striatal cell

suspensions in ischemic rats "as investigated. Ischemia was induced by

intraluminal occlusion (for I h) of the middle cerebral artery under halothane

anesthesia. Fetal striatal cells, marked with DII, were transplanted into the ischemic

striatum. Behavioral function was investigated by passive avoidance task and radial

maze learning. Transplantation reversed the ischemia induced behavioral deficits in

the passive avoidance task, but not significantly in the radial maze learning. Dil

positive cells with dendritic outgrow th could be seen under fluorescent microscopy.

Immunohistochemical study revealed that many choline acetyltransferase (ChAT)

positive cells and GABA positive cells survived, but their neurite ingrowth to the

host brain was poor. No regular striosome-rnatrix compartments were evident

inside the grafts, Thus, partial recoveries in cytochemical and behavioral aspects

were obtained by fetal striatal cell graft in ischemic brain.

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PO-C27

A STUDY ON THE HOURLY CHANGES OF THE LCBF AND THE EEGIN ISCHEMIC RABBIT-BRAIN MADE BY LIGATION OR EMBOLIZATIONO.Komatsu.H.Ketoki.Y.Oike(ReimeikYo Rehabilitaion Hospital,Aomorl.Japan)T.Kanazawa,K.Onodera(Second Internal Medicine,Hirosaki University School of Medicine. Hirosaki. Japan)The hourly changes of the local cerebral blood flow (lCBF)and electroencephalography (EEG) in ischemic brains wereastimatei to study the pathophysiology in ultra-acute stageof the brai~ ischemia. Kethods:Thirty male white Newzealandrabbits were classified into three groups. Ligation group~n=lO):the left carotid artery was ligated. Sephadex G-75group(n=lO):suspensio~ of SG-75 was injected into the leftcarotid artery. Control group(n=lO):sham operation.The lCBFwas measure~ by hydrogen inhalation method and the EEG wasanalyzed by Fourier transformation method. The lCBF and theEEG were recordej simultaneously from both side of theparietal cortex for 3 hours. Results: 1) The histologicalchanges such as swelling of the astrocytes and shrinkage ofthe ~erve cells were observed in the left cortex only ofthe SG-75 group by H-E and electromicrosCopy. 2)In ligationgroup. the lCBF did not change in both side of the cortex,but the total power decreased hourly in the left cortex. InSG-75 group. the lCBf decreased markedly in the left cortex. but the lCEf in the right cortex did not change. Onthe othar ~anc. the total power and mean frequency de:reased not only in the left cortex but also in the rightcortex. Conclusion: It was suspected that the ischelicchange in the left cortex effected on the right helispherethrough some communicating tructs. The lCBF and the EEG didnot show the parallel change in acute ischemic brain. so itwas speculated that the EEG was controlled by not only thelCBF tut another mechanism.

PO-C28

SIMULTANEOUS LASER DOPPLER FLOWMETRY IN CAl SECTORAND CORTEX OF GERBILS SUBJECTED TO FOREBRAIN ISCHEMIA

Toshihiko Kuroiwa*. Petra Bonnekoh and KonstantinAlexander Hossmann

Dept of Experimental Neurology. Hax-PlanckInstitute for Neurological Research, Germany. *Deptof Neuropathol., Tokyo Medical and Dental Univ. Japan

Regional cerebral blood flow (rCBF) in the frontalcortex and hippocampus CAl sector was measuredcontinuously using laser Doppler f10wmetry in gerbilssubjected to 10 min bilateral forebrain ischemia.After induction of ischemia, rCBF in the cortex andCAl sector decreased to 11.8% and 18.0% of thebaseline value, respectively. After recirculation,rCBF returned to the preischemic level at 7.5 min(recovery time), reached the hyperemic peak (122.2%)at 12.4 min (peak latency) and decreased to thepreischemic level at 27.2 min after recirculation.In the CAl sector, rCBF returned to the preischemiclevel already at 2.1 min, reached the hyperemic peak(123.8%) at 5.7 min and decreased to the preischemiclevel at 21.3 min after recirculation. In both thecortex and CAl sector, both recovery time and peaklatency significantly correlated with the depth.~f

ischemic flow. Histologically, cortical neurons werenot injured, whereas only 15.4% of CAl neuronssurvived 1 week after ischemia. Thus CAl neuronswere selectively vulnerable despite the milderdecrease of percentage flow during ischemia andprompter recovery of postischemic flow. Thesefindings show an important role of intrinsic ratherthan hemodynamic factors in the mechanism ofselective vulnerability of CAl neurons aftertransient bilateral forebrain ischemia.

LOWER LEVEL OF ENDOlliELIN-l IN MIGRAINE WIlliAURA

Sitoru Komatsumoto, Taishi Sasaoka, Hidetomo Nakamotor.lls~haru Nara, Fumio Gotoh* Department of Internal Medicine:Ashikaga Red Cross Hospital, Tochigi JAPAN *Department ofNeurology. School of Medicine, Keio University, Tokyo Japan

Endothelin-l (ET-I) has a powerful vasoconstrictive and bloodpressure rising properties by endothelial cells. However. nosystematic evaluation of ET-I in migraine has ever beenattempted. The present study is focused on evaluation of level ofET-I in patients with migraine. Studies of ET-I level were madein 9 patients with migraine with aura (age 17.9±2.3, mean±SD)according to the Classification of Headache of the Ad HocCommittee. All the patients were free of migraine attacks for atleast 7 days prior to the examination. Ten age-matched healthyvolunteers (age 16.4±1.7) were similarly used as control group.None of !hem reve~led evidence of any other disease. such ashypertension, obesity. heart disease. Informed consent wasobtained from each subject. Subjects were kept supine in a quietroom for at least 60 minutes. Blood samples were drawn fromcubital vein by syringe with 23G needle. We have measuredimmunoreactive ET-l by radioimmunoassay in plasma. Theplasma level of ET-l in migraine showed 2.44±1.09 pg/ml. Onthe other hand. the level of ET-I in the control group was4.35±O.86 pg/ml. The level ET-I in migraine was significantlylower when compared with that in the control group (p<0.OO2).u;>wer ~T-.I concentratio~ in plasma was clarified in the patientsWith rmgrame, In conclusion, the lower concentration of ET-I isconsisten.t with pathogenesis of migraine, further supporting thehypothesis that lower ET-I may be closely related to markedcereb.ral vasodilatation following the constriction partly due todeficiency of ET-I to maintain the vasoconstriction.

PO-C29

ACUTE HOHHZi·l0RRllJl.GIIJ S'hWAZ:l-;'::"Unv.LUGICAL .iI.'jD 3IUCHEI';IG.-u' ;:':l'UDi

Kovacevic M.S.,Mrsulja B.B.,Bumba~irevic ~j.

Jovanovic D.,Zidverc J.,Bug~rski C.,DociicA.,Pecic Q.,Beograd,Yugoslavia

The efficacy and safety of dihydroergotoxine(DHETX) mesylate were assessed in a pilot,randomized,placebo-controlled,double blind trial.Treatment of 40 adult p~tients with ischemic stroke 12 hours after stroke onset.Admission (baseline) laboratory serum tests andcerebrospinal fluid (CSF) analyses (glucose,lactate,pyruvute,5-HT,5-HlAA,DO~AC,nV.iI.,GABA)were obtained before the initiation of thetherapy.DHETX was without effect on the level of fibrinogen and hematocrit values, butsignificantly (P?o.o5) reduced serum glucoselevels.CSF glucose level was significantly(PLo.o5) reduced in tho therapy group. Indexof glucose utilization (IGU) in the brain/loo-(glucose CSF/glucose serum) x 100/ wasfound reduced from 68% (contrOl) to 38% before therapy;atter 14 days of the treatmentsIGU was 3~% in placebo but 48p in DEETX group indicating better utilization of brain glucose in the DHETX patients.5-rlT/5-!::IAA ratio was found significantly (p(o.ol) reduced in all patients upon admission suggesting increased 5-HT release.No differences were found in the levels of HVA,DOrAC and GADAwhile the levels of USF pyruvate and lactate indicated increased glycolytic capacityin DHETX patients.

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PO-C31

591

The Effect of Naloxone on rCBF and Infarction Size inFocal Cerebral Ischemia of Rats.

Sang-Bok Lee, ~1. D., Seung-Bong Hong, M. D. ,Jae-Kyu Roh, M.D.• Byung-Woo Yoon, M.D.Seoul, KOREA

Focal cerebral infarctions of Sprague-Dawley rats were madeby middle cerebral artery occlusion(MCAOl. Control group wasgiven sal i ne (4 cc/kg boIus and then 2 cc/kg/hr- i. v, )starting at 30 min before MCAO. Naloxone groups were givenlow-dose Naloxone (I mg/kg bolus and then 0.5 mg/kg/hr i.v.)and high-dose Naloxone (4 mg/kg bolus and then 2 mg/kg/hr'i.v.). After 24 hours, rCBFs were measured by autoradiographyusing C14-iodoantipyrine. Brain slices (2 rom thick) werestained with 2.3,5-triphenyl-tetrazolium chloride and the sizeof infarction was measured by planimeter. There were nosignificant differences of basal BP, pH, PaC02. Pa02 and bloodglucose levels among three groups. Low-dose Naloxone groupshowed no significant rCBF changes, while high-dose Naloxoneimproved rCBFs in two areas [caudate head: 72.7~14.3(saline).

90.0~18.6 ml/IOOg/min (Naloxone). p(0.05 : CA3: 87.0~6.6(saline), II0.3~6.2 ml/lOOg/min (Naloxone). P(0.05]. Onerom-distance serial measurement of rCBFs from paramedian cortexshowed that high-dose Naloxone group maintained slightlyhigher rCBF levels in penumbra areas than those of salinegroup. Infarction sizes were reduced significantly inhigh-dose Naloxone group (6 rom from frontal pole ::) salinegroup: 39.0~I.4 %, high-dose Naloxone: 24.0~3.04 %. P(0.01 :8 rom ::) saline group: 30.7~2.9 %, high-dose Naloxone: 18.5~

2.3 %. P(0.05). Low-dose Naloxone treatment showed nosignificant reduction of infartion size.

In summary, it is suggested that a pretreatment of highdose Naloxone reduces infarction size and may improve rCBF atpenumbra area in focal cerebral ischemia of rats.

PO-C32

An oberservation on arginine vasopressin levels of plasma and

cerebrospinal fluid in patienls with acule ischemic stroke

Liu Xin-Feng, Shi Yin-Mian, Ling Bao-Cheng. Departmenl of

Neurology. Jinling Hospital, Nanjing 210002. P.R. China

The contents of arginine vasopressin (AVP) in cerebro

spinal fluid ([Sf) and plasma were determined b)' radioilllllU!loassy

in 40 patients with ischemic stroke. The results showed that CSF

A\'P was significantly increased, and the change of plasma AVP was

nol significant. E..<;f AVP was much higher in the severe cases than

in the mild cases. The targer volume of the lesion showed by CT

scanning lias, the higher contents of E..<;f AV? Of all the cases,

the contents of (Sf A\'P were the highest in the palienls with

high intracrainal pressure, and there was a posi t ive correlation

between the inlracrainal pressure and (Sf AVP contents. These

suggested that AVP was lnvotved in the palhophysiological process

of ischemic brain edema. The change of (Sf A\'P could ref tei t th~

serious degree of the brain edema in the acute ischemic stroke.

An experlsentat study of the effect arginine vasopressin on acute

ischemic brain edema

Llu Xin-Feng, Shi Yin-Mian, Ling Bao-Cheng. Deparlmenl of

Neurology, Jinting Hospi tal, Nanjing 210002, P.R. China

The purpose of this experiments is to study the role of arginine

vasopressin (AVP) in acute ischemic brain edema and its mechanism.

The results showed that the contents of AVP in ischemic brain

regions significantly increased, and there was a relalionship be

tween the contents of AVP in cortex and Ihe ischemic cortical

edema. Intracerebroventr lcutar injection (ICY) of AVP exacerbated

the ischemic brain edema, and its showed dose-response correla

tion. The catclua entry blocker coutdrr t block this role of AVP

in ischemic brain edema. ICY of AVP antiserum, V. and V.?!. re

ceptor anlagoni~1 respect ivety decreased significantly the ischc

ale cortical edema, while V, receptor antagonisl had no such

effect. In addit ion, the Na'-K" ATPase activity of ischemic cortexwas significantly decreased, the cAMP levet of brain regions, and

cOO of hypcthataaus and slralium were markedly increased after

ICY of AVP: Finat ty we may conclude that AVP was lnvotved in the

pathophys lotogic process of acute brain edema. and its mechanism

mighl be the effecl of AVP on AVP recepter mediated by cAMP, cliMP,

and Ihal in turn inhibi ted the Na'-K" ATPase aclivi ty of brain

cell membrane, then exaggeraled the formation of ischemic brain

edema.

PO-C33

RESIDUAL DISABILITY AND EARLY HEMODYNAMIC ASSESSMENTIN ACUTE CEREBRAL ISCHEMIAM. MARINONI A. GINANNESCHI, D. INZITARI,L. AMADUCCI Department of Neurological Sciences,University of Florence, Italy.

To evaluate disability status in acute ischemia, 22patients (15 males and 7 females, mean age 67.0±9.6years) with hemispheric cerebral deficits werechecked on the Barthel Index (8I) for 120 days afterthe onset of symptoms. CT scan demonstrated ischemichemispheric lesion in all patients. Cerebral hemodynamics was studied by means of Continuous WaveDoppler (CWD) and Transcranial Doppler (TeD) performed within six hours from the beginning of symptoms. CWD demonstrated carotid hemodynamic stenosisor occlusion on the lesion side was considered asabnormal extracranial pattern. Mean flow velocityvalue in the middle cerebral artery on the lesionside out of normal range (>30 and <80 cm/s) wasrated abnormal. BI scores at day 120 were better inpatients with normal than in those with abnormal CWDexamination (Mann-Whitney U test, P=.04). BI scoresat day 120 were better (Mann-Whitney U test, P=.04)in patients with normal TCD values than in thosewith abnormal. Early TCD and CWD examination mayprove useful information about residual disabilityof patients with hemispheric cerebral ischemia.

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PO-C35

15.9±O.9

Control17.1±2.0

No Autonomic Functional AsymmetIy in Patientswith Acute Hemispheric Stroke

Y. Maruki" ••, K. Shimazu", N. 'I'arnura", T.Ohkubo·, H.Sugimoto", T. Yamamoto", Y. Asano", S. Watanabe·, M.Sawada" and K. Uamaguchi'"Department of neurology,Saitama Medical School", and SaitarnaNeuropsychiatric Institution", Saitama, Japan

Autonomic functional asymmetry was demonstrated inexperimental stroke (1991, Hachinski), We tested autonomicnervous functions in 44 patients with . acute hemispheric strokewithin 7 days after the onset and 71 age-matched healthyvolunteers. Hemodynamic functional tests were performed undercontinuous monitorings of blood pressure (BP) and pulse rate.Pupillary diameter was measured before and after 1.25%epinephrine eye instillation.I) Reflex bradycardia in Aschner's eyeball pressure test:

Right stroke Left strokert stimulation 11.2±3.7(1min) 6.8±3.S*It stimulation l2.3±4.0 13.9±5.2

II) Reflex hypertension in cold pressor test:rt stimulation 19.1±4 .1CmmHg) 21.5±2.9*It stimulation 19.1±3.7 25.4±2.9*

III) Fall in BP and reflex tachycardia in postural change:fall in BP 20.7±4.7*(mmHg) 19.7±3.4 l2.6±1.6tachycardia 11.7±1.6(1min) 10.8±1.7 l4.8±1.l

IV) Reflex mydriasis in 1.25% epinephrine instilation test:rtstimulation 1.5±0,4*(mm) 1.1±0.3 0.7±O.1It stimulation 1.S±0.5* 1.3±0.3

values:mean±SE, *:p<0.05 compared with controlNo autonomic functional asymmetries were obtained between

right and left hemispheric lesions, although the pati ents with acutestroke had significant autonomic dysfunctions compared with thecontrols.

PO-C36

Continuous Cerebral Infusion of l-noreplnephrlna Protects againstHippocampal Neuronal DeathafterTranslenl ForebrainIschemia.

K.Nishino. M. KowadaNeurosurgical Service, Akita University Hospital1·1·1 Hondo, Akita, Japan.

It has been suggested that noreplnephrlne(NE) In the CNS plays anInhibitory role In the hippocampal neuronal loss after transient globalIschemia (Blomqvlst ot al., 1985;Nlshlnoet al.,). The 90alof this studyIs to determine whelher Inaeasing lovels of NE protect againstneuronal damage. FIISI, degree of NEdiffusionwas analyzed by NEassay of hemIooronal brainsllces In normal animals subjected to twoweeksInfusionof50mM I·NE.Then. we treated mongoliangerbils(5075gm. male) with sin91e directlntrahlppocampal infusion(lpl of 50mMNE) or Alzet mlnlosmoticpumpfilled with 50mM I·NE shortly before orboth before and after S minutes of forebrain IschemIa A weekposlischemla all animals were transcardiafiy perfused and lhe removedbrains were processed lor 4 pm thick Nissle-slained seelion todetermine the number of survlvlng neurons. The continuous InfusionInaeased levelsof NE S-40fold on lhe Infusion side and 2·4 fold onthe contralateral side (n-5) as compared to normal control (n-S) (t-test;p< O.OOS, p< 0.05, respectively). The same treatment for a weekbeforeand for a week aller the Ischemia also significantly Inaeasedthe numberof survlvlng neurons In CAl and CA3 sUbreglons(n-7} ascomparedtocontrolanlrnals(n-16) (Infusion side,contralateral side, andcontrol; 9.S±2.6" Ul00pm; meantSEM). 5_3t2.8·, 1.110.7 In CAl;12.2±O.9S", 11.3±1.1·, 7.6±3.4 In CA3, Duncan's lest; ., p<O.Ol, •p<O.05}. However, there Is no significant Inaease In the numbers ofsurvival neurons associated wilh the single Infusion. In separateexperiments, hippocampal NElevels weremeasured with HPlC aller Sminutes of forebraln Ischemia plus brief rellow. NE level In thehippocampus significantly deaeased by approximately 40 % after 5minutes 01 IschemIa and an subsequent hour of renow(n-8}. Inconduslon, these data not only support the hypothesis 01 NE·lnduoedneuroprotection but also provide lhe rationale to boost NEIncerebralIschemia.

Effects of Very-Short-Time Ischemic Insult onHilar Somatostatin Neurons in Gerbil Hippocampus

H. Nakamura, T. Matsuyama, M. Tsuchiyama, H.Tachibana, O. Uyama, M. Sugita, Fifth Departmentof Internal Medicine, Hyogo College of Medicine,Japan

It ha s been reported that ea r l y selective lossof s omat o s t a t i n (SS) neurons in dentate hilusprecedes CAl pyramidal neuronal death after 5-mintransient cerebral ischemia in the gerbil (Cereb.Blood Flow Metabol. 11, Suppl e 2, 1991),sugge s tin g that loss of SS neurons induce shyperactivity stimulating the CAl neurons todeath . On the other hand, ve r y-shor t- time i schemicinsul t has been r e ported to induce i schemictolerance on the vulnerable CAl neurons. Thisstudy were undertaken to evaluate the change inthe hilar SS neurons at various intervals aftersingle or repeated 2-min cerebral ischemia, usingimmunocytochemistry. A significant 40% loss ofhilar 5S neurons t oo k place eve n at 1 da y after 2min i schemia, and i n a 60 % loss a t 7 day s afterischemia. At 2 days after secondary i schemia,whi ch was carried out following a I-day delay ofthe first insult, whe n the ischemic tolerance maybe most prominent, almost no SS neurons werede monstrated in the hilus. At 7 days after I -minischemia, no SS neurons wer e lost. The presents t udy c learly demonstates that hilar SS neuronswer e more vulnerable to ischemic insult than CAlpyramid al neurons. Th e ischemic tolerancephenomenon may be induced during the progress ofthe damag e of hilar SS neurons through thesynaptic ch anges to the CAl pyramidal neurons.

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Prostaglandin EI Protects Rat Primary Cultured Neurons againstHypoxic Injury

Hideo Otsuki, Kazuo Yam ada . Marnoru Taneda, Tom HayakawaDepartment of Neurosurgery, Osaka University Medical School.Osaka. Japan

We investigated the effects of prostaglandin E I (PGE I) on thehypoxic injury of fetal rat hippocampal neurons. Primaryhippocampal cell cultures (embryonic day 18) were establi shed andmaintained with the serum-contain ing medium. After 2~ hincubation. cultures wer e rinsed and serum-free. defined N Imedium wa s added. After 72h. PUEI were prepared with fresh N Imedium at the final concentration of 10-5 to 10.11M, and the oldmedium was replaced with an equal volume of POE l-containingones. Con tra] received a fresh N I medium without POE I. Cultureswere devided into two groups: nonnoxia group was in culture foranother-lS h, and hypoxia group was exposed to 24 h of hypoxicstress in which final oxygen tension of the medium was reduced to20 torr. follo wed by continuation of culture for another 24 h. As aquantitative measure of cell death. activity of LDH was estimated inthe culture medium. and the residual activity within the cells wasalso determined in suspensions of freeze -thawed cultures. Incontrol and POE1-lreated cultures. phase cont rast micrographs ofcell cultures showed neuronal damages after hypoxia.The activityof LDH in the culture medium. released by the hypoxic insult . wassignificantly smaller in POEI 10-7 M and 10-~M (p<O_OI).and10-6 M and 10-9 M (p<O.05). compared to the control. In normoxiagroup, no differences were observed between POE l-treatedcultures and control. To determine whether protective effect ofPOEI again st hypoxia was medi ated by gl ias , primary culture wasreplatedtwice. so that large number of cells were glia s. Afterhypoxic stress the LDH act ivity in the medium was not different.We conclude that POE 1 at the concentration of 10-6 to 10-9 Mprotects rat hippocampal neurons against hypoxic insult.

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ASTROGlIAL ANDMICROGLIALREACTIONS IN EARLYSTAGESOF ISCHEMIA: AN IMMUNOCYTOCHEMICAL ANALYSIS IN APRIMATEMODELOF ISCHEMIA.Pardo CA, Monseln L, Mathews V, Barker P and Bryan RN.Neuropathology Laboratory and Division of Neuroradlology,The Johns Hopkins School of Medicine, Baltimore, MD, USA.

The development and magnitude of astrogllal and microglialresponses were evaluated Immunocytochemically In a model ofregional Ischemia In baboons. The model Included two variants:(1) Complete Ischemia (CI) by Irreversible occlusion of themiddle cerebral artery (n=3.mean occlusion time: 15 hours); and(2) Incomplete Ischemia (IIR) by reversible occlusion of MCAfollowed by reperfuslon (N=4j mean occlusion time 4 hours 32rnln; mean reperfuslon time: 2 hours 51 mIn.). Antibodiesagainst astroglial filaments (GFAP) as well as macrophage andmicroglia receptor-proteins (HLA-DR.CD68and ED2)were usedto stUdy the responses of these cells following Ischemia. In bothexperimental approaches, early astrogllal changes wereobserved predominantly In the region surrounding the necroticcore and were characterized by perikaryal hypertrophy,proliferation and extension of the glial processes. In corticalIschemic regions, astrocytes Issued long processes resemblingradial fibers. These changes were most prominent In IIR modeldespite the short duration of the Ischemic Insult as comparedwith the CI model. Few macrophages and activated microglialcells were observed In CI model as compared with numerousactivated microglial cells and migrating macrophages IdentifiedIn pennecrouc regions In the IIR model. These findings suggestthat both astrogllal and microglial cells have an Important roleduring early stages of Ischemic Injury, and reperfuslon mayfacilitate migration and activation of macrophages and microglialcells leading to more severe damage In the Ischemic regions.

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The Effect of New Calcium Antagonist TA3090 (8-Chloro Diltiazem) on The Cerebral Vessels in Experimental Transient focal Ischemia

T SAKAKI, S TSUJIMOTO, Y SASAOKA, S TSUNODA

TA3090 hs been shown in three experimental circumstances to have cerebrovascular protective properties.In vitro investigation was undertaken to determineeffects on pial arterial behaVior, cerebral blood flow (CBf), and its therapeutic value for the brain intransient focal cerebral ischemia. In this experiment400ug/Kg/hour of TA3090 (treated group, n=14) or physiological saline (control, n=10) was administered5 minutes before one-hour middle cerebral artery occlusion in cats. Pial arterial behavior and CBF wereobserved continuously, and then autoradiography forquantitative measurement of CBF was performed 5 hoursafter recirculation of the MCA. Volume of cerebral edema and infarction was estimated in planimetry fromcerebral preparation made for histological examination. Dilatation of pial arteries was significantlysmaller during and after insult in the treated groupthan in the control group (p<O.Ol J. CBf was significantly more increased during ischemia (p<O.Ol), andmore decreased after ischemia (p<0.05 at the end) in~e treated group than in the control group. Autoradiography showed remarkable increase of CBF due to "luxury perfusion" at the cerebral cortex which wasmainlynourished by the MCA in the control (p<O.Ol).Cerebral edema and infarction were much smaller inthe treated group (p<O.Ol). These stud indicate that a pretreatment with TA3090 offers cerebral protection and reduces cerebral pathological areas.

Therapeutic research based on the Bengal Rose modelof focal cerebral ischemia? A MRI supported dischargeW. Reith 2, M. Forsting1, Uta Meydlnq-Larnade 1, A.Dorfler1, R. v. Kurnmerl, W. Hacke 2, K. Sartor l

Depts. of Neuroradiology1 and Neurology2, Heidelberg

Background/Objective: One of the key questions inexperimental stroke research is whether animal modelstruly simulate human disease. The Bengal Rose model isa simple method to induce in situ thrombosis of selectedcerebral cortical vessels. It seems to be an ideal modelfor testing therapeutic agents in focal cerebral ischemia.The aim of our study was to evaluate the usefulness ofthis model in neuroradiologic monitoring of ischemicevents. Methods: A focal ischemic lesion was induced bythe Rose Bengal method in 50 Wistar rats. 6 groups ofanimals underwent MRI examinations at time intervalsbetween 1 and 120 hours after illumination. Gd- DTPAand Polylysin were used as paramagnetic contrastagents. Results: After 1 hour. there was already aremarkable contrast enhancement of the ischemic area inall animals, even after administration of macromolecularagents such as polylysin. This indicates an almostinstantaneous blood-brain barrier (BBB) disruption afterillumination. Conclusion: BBB permeability and thedevelopment of cerebral edema are important events inthe process of ischemic infarction. Our MRI resultssuggest that the way BBB disruption occurs in this animalmodel differs entirely from that in human brain infarcts.We therefore suggest not to use the Bengal Rose modelfor therapeutic research in cerebral ischemia.

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3D MR MICROSCOPY OF CEREBRAL TISSUEDAMAGE IN MOUSE MODEL OF FOCALISCHEMIC STROKESusanta K. Sarkar, Rasesh D. Kapadia, Dorothy J.Knudsen and Frank C. Barone. SmithKline BeechamPharmaceuticals, King of Prussia, PA, USA.

MR microscopy can monitor time-related changes incerebral tissue following focal ischemia and theirresponses to drug therapy. In order to utilize the fullpotential of the technology, we have quantified volume ofinfarction in a mouse focal stroke model using 3D MRmicroscopy in.vivo. This approach to study cerebralischemic events and their consequences in the mousemodel has not yet been evaluated. Mouse focal ischemiaprovides a relatively high throughput, small animal modelthat requires relatively little quantity of drugs for monitoringefficacy. After initial images were taken. each mouseunderwent micro-surgery to occlude the middle cerebralartery. Images were collected after the surgery and thedevelopment of the infarcted region over time was clearlyidentified in reference to other brain structures withexcellent resolution, 80 x 80 x 80 11m. Ischemic damagethat was identified by MRI correlated with histologicalmeasurements. Sham surgery did not produce anychanges in MR images or histologically detectable cerebraltissue damage. We have now extended these studies todiffusion-weighted MR microscopy in order to detect initialchanges in cerebral ischemia. These results along withactual quantitation of infarction profiles and volumes will bediscussed. In conclusion. MR microscopy in mouse focalischemia provides an excellent experimental system tostudy pathology and therapy in stroke.

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Localization of Endogenous Supcroxide DismutaseImmunoreactivity in Gerbil Brain After IschemiaS. Shimizu, T. Matsuyama, H. Michishita, H.Nakamura, O. Uyama, M. Sugita, Fifth Department ofInternal Medicine, Hyogo College of Medicine,Japan

Induction of the supcroxide dismutase (SOD) rnRNAhas been demonstrated in brain following ischemia(Stroke 1992, in press). In the present study,CuZnSOD and MnSOD protein were localized in gerbilhippocampus at intervals after S-minutes'transient ischemia using polyclonal antibodiesspecific for rat CuZnSOD and MnSOD, respectively.

CuZnSOD immunoreactivity was found exclusivelyin the neurons of the CA sectors. The accumulationwas slightly reduced throughout the hippocampus at3 hours, and much more reduced selectively in theCAl pyramidal neurons at 24-hours I recirculationwhen the prolonged expression of CuZnSOD rnRNA wasseen there. The immunoreactivity was recovered at3-7 days only in the spared cells. In contrast,MnSOD accumulation in CAl was minimal in controlanimals and was not changed .dur Lnq the first 24hours after ischemia. At 3 days, however, MnSODimmunoreactivity was found in some glial cellsoccurring in the CAl sector. The microglial cells,which were identified using lectin staining withthe Griffonia simplicifolia B4-isolectin, formedclusters in the CAl pyramidal layer at 7 days.

The present resul ts demonstrate that the SODproteins are not synthesized in CAl vulnerableneurons during the postischemic period, but areinduced in some glial cells which may promote aphagocytic action.

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Delayed Neuronal Oama~c Fo l Iow.ing ~Iiddle CerebralArtery Occlusion in St roke-Pronc SpontaneouslyHyper-t cns ive Rats(SIIRSP)

Tosh ik i Sh i rot an i , NiNako Iwat a , IIideyuki Ki t a ,Katsuji Shima, lIiroo ChigasakiDepartment of Neurosurgery, National Defense

Medical College, Saitama, Japan

We have attempted to identify the delayedneurona l damage in the SllRSP model of middlecerebral artery(~lCA) occlusion, The distrihutionof the neuronal damage was de t erm i ned by using theCa-45 autoradiographic technique, After MeAocclusion, calcium accumulatiun was immediatelyobserved only in the ccr-ebr-a l r-or t ox supplied bythe occl uded ~ICA. Followed by the 3 clays ofocclusion. calcium had accumulated in theipsilateral pyramidal tract, ventral posteriornucleus of the thalamus, and the lateral part ofthe striatum. Remarkahle calcium accumulation wasohserved in the same ar-eas hy 2 weeks. In add i t ion.we have measur-ed changes in t he ext r-ar-eHu larfluid(ECF) concentration of amino acids. dopamine,and dopamine metabolites in the striatum duringischemia using Int r-acercbr-a I mi c rnd i a lys Is .Following NCA occlusion. a 2-foIcl increase indopamine, a 3-fold transient increase in glutamatewere observed in the lateral par-t of striatum.Ther-e was no significant Iuc-rensc in glutamate,dopamine, and dopamine metabolites measured on theinner part of striatum. We speculate that themechanisms of dl.'layed neuronal damage in theipsi lateral pyramirla l t ruct and t ha l amus mal' becaused by secondary neuronal degeneration, butthat in the striatum may be related to massiveglutamate release,

Autoradiographic Analysis of [125IjEndothelin-1

Binding in Ischemic Gerbil Brain

T. Shirai, Y. Fukuuchi, K. Tanaka, S. Takashima,B. Mihara, S. Nogawa, H. Nozaki and E. NagataDepartment of Neurology, Kcio University, Japan

In order to examine the relationship between the distributionof Endothelin (E1)-1 binding sites and the mechanism ofischemic brain damaRe, we conducted autoradiographicbinding analysis with [12SIjET-1 in ischemic gerbil brain. SixMongolian gerbils were divided into two groups: the ischemiagroup (occlusion of the right common carotid artery for 6hours, n=3) and the sham operated group (n=3). At 6 hoursafter the surgery, local cerebral blood flow (ICBF) wasdetermined by the [14C]iodoantipyrine method. The serial brainsections were incubated with 1.16xlO-IO M [12SIjET-1 in thepresence or absence of 8.03xlO-7 M unlabeled ET-1 for 10min at 25°C. The specific binding was calculated bysubtracting the amount of non-specific binding measured withunlabeled ET-1 from the total amount bound. The data for thelCBF (mVI00g/min) in the ischemia group were 30-100 onthe ischemic side, and 80-200 on the contralateral side. Theischemic side exhibited a significant reduction in lCBF(p<0.05-0.0l). The specific binding amount of [12SIjET-l(fmo\fmg, mean±SD) in the ischemia group was 3.13±0.02/3.20±0.02 (ischemic/non-ischemic side) in the frontoparietalcortex, 2.66±0.5812.58±0.79 in the caudate putamen, and3.32±0.45/3.45±0.65 in the hippocampus (CAl). The presentstudy showed no significant changes in the amount of ET-1binding after 6-hour ischemia in the gerbil brain.

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Insulin Growth Factor Type 1 in transientglobal ischemia in gerbils.

S. Izaj, A. Shuaib, S. Hasan, B. Bhaumick.

CANADA.

Insulin when used during transient forebrain ischemiaprotects neurons in the hippocampus in small animals.This effect may be secondary to its glucose-loweringeffects, or alternately, insulin may act as a growthfactor. The use of other insulin-like growthfactors in ischemic damage prevention has not beenwell studied. We used the gerbil model of transientforebrain ischemia (5 minutes) and tested two dosesof insulin growth factor (type 1) (IGF-l) via Alzapumps continuously for 7 days after the insult. Thedegree of neumnal damage in the hippocampus,striatum, thalamus, and cerebral cortex was similarbetween two doses of IGF-l (n=12) and controls (n=6).

IGF-l does not appear to protect the brains from theeffects of forebrain ischemia in gerbils. It ispossible the protective effects of insulin may besecondary to its glucose-lowering effects oralternately neural protective effects of insulin donot appear to be present in IGF-I.

SECOND WORLD CONGRESS OF STROKE, 1992 595

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Oxygen-free radicals and antioxidant statusduring acute ischemic stroke.

Shuaib, A, "Mosowich, R, Kalra, J, ~mntha, C andPrasad, M.

CANADA.

Oxygen-free radicals (OFR) have been i mplicated inthe pathogenesis of experimental cerebral ischemia.OFR may exert their cytotoxic effects by lipidperoxidation, resulting in production of malondialdehyde (~IDA). Their role in human acute stroke hasnot been well studied. We collected blood frompatients with acute stroke within 48 hours andcompared it to age-matched controls free of acutebrain injury. Superoxide dismutase (SOD), catalase(CA), and glutathione peroxide (GHS-Px) activitieswere measured in the serum and erythrocytes. OFRproducing activity of PMN in leukocytes was .measured by chemiluminescence. In al l , 30 patLentsand 2S controls were studied. There was a significant increase in activity of GSH-Px and MDA(P(O.OS). SOD and CA were not s~gnificantly

increased in acute stroke, although there was atrend towards higher values in acute stroke. Theincrease in GIlS-Px and ~IDA would indicate thatthere i s an increase in OFR activity in acutestroke. Further studies within the first 4-6h~urs after an acute stroke need to be done tobetter understand the effects of free radicals insuch patients.

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TEMPORAL PROFILE OF NEUROWGICAL SYMPTOMS ANDffiSTOWGICAL DAMAGES IN GERBILS WITH TRANSIENTUNILATERAL BRAIN ISCHEMIAM. Tagaya, M. Matsumoto, K. Kitagawa, T. Ohtsuki, R. Hata, S. Ogawa,H. Ueda, N. Handa and T. KamadaFirst Dept. of Med., Osaka Univ, Medical School, Osaka, Japan

Introduction; In gerbils with unilateral brain ischemia, neurological manifestations were observed in a wide range. In this study, we demonstrated thecharacteristic temporal profile of neurological symptoms and histologicalfindlngs during both ischemia and reperfusion to establish a system ofclassification in gerbils with 30 min of unilateral ischemia according to thedegree and severity of neurological signs.Methods: Cerebral ischemia was produced by occluding the right commoncarotid for 30 min in 120 gerbils with light ether anesthesia. Animals weredivided into classes with no. mild, moderate. and severe symptoms using thecumulative neurologic score (1). We observed the survival rate. neurologicalsymptoms and histological findings up to 24 hour after braiJ.! ischemia. ~ forneurological symptoms, the dynamic change of motor function was examinedin particular using an inclined plane method useful for detecting a subtle degreeof paresis quantitatively (2). Histological evaluation '7'as made br HE and anirrununohistochemical staining for microtubule-associated protem 2 (3).Results : All animals with no. mild or moderate symptom survived. but abouta half of the severe group died throughout the procedure. Symptoms in themoderate group disappeared in 2 hours and violent ones such as seizurecontinued for 24 hours in the severe group. As for motor function, hemiparesisin the mild, moderate and severe groups already developed at the ischemicperiod of 2 min. Thereafter mild animals recovered before reperfusion but therecovery of moderate and severe ODCS was slow. The result of histologicalexamination was in accordance with the severity of ischemic symptoms.Conclusion : The present study characterized the temporal profile ofneurological symptoms and histological damages in gerbils w~th 30 min ofunilateral cerebral ischemia and suggested the usefulness of this model as amimic of human stroke.References : 1. M.Matsumoto el al. Stroke 19:490,1988 2. N.Handa et al, LifeSci 42:1825,1988 3. K.Kitagawa et aI. Neuroscience 31:401,1989

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TilE EFFECTS OF STRAIN DIFFERENCEON INFARCT SIZE FOLLOWINGHISTOLOGICAL QUANTIFICATION OFFOCAL ISCHEMIA IN THE RAT.J.T. Simmonds and J.A. Mover. CNS Division,Wyeth-Ayerst Research, Princeton, NJ 08543-8000

It has been previously shown that differences in rat strainproduce cerebral lesions which vary in mean infarctvolume as well as lesion reproducibility (Duverger et al.,1988). The effects of strain difference in histologicallyverified focal ischemia in Sprague-Dawley, Fisher-344 andSHR rats was examined as a means of establishing apharmacological model for evaluating known and novelEAAA (excitatory amino acid antagonists). Male rats(29Q..310g) were sacrified 24 hours post middle cerebralartery (MCA) occlusion. Infarcted volume was detl;nninedby extensive area measurements on every tenth sectionthrough the lesion-by image analysis. Sectioning wasdone at 20 micron increments through the infarcted areaand volume was multiplied by the tissue density. Themean infarct volume produced by the Sprague-Dawley,Fisher-344 and SHR rat strains (n =4 per group) was33.2 mm3, 64.8 mm3, and 98 .0 mm3, respectively. Thelesions produced in the Fisher-344 strain were larger andless vanable in size than those in the Sprague-Dawleystrain . The SHR strain produced the largest lesions andthe interanimal variability was minimal. Each strain'smean infarcted volume was statistically different from theother when tested by analysis of variance with repeatedmeasures. These results demonstrate how strain differenceplays a major role in infarct size and reproducibilityfollowing MCA occlusion.

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PARACRYSTALLINr: ARRAY IN Vr:SSr:L WALL IN r:Xpr:RIt-reNrALfOCAL CEREBRAL ISCHAE.'lIA

DR. DEE:PA R. THr:OOORE & C.D.BHANUMATHY, INDIA

r:M studt of insula and caudate nucleus from thebrains of monkeys with right MCA occluded fordifferent time periods ranging fro~ ~ hour to twoweeks, revealed a very few unusual dtructures apartfrom the well described changes. Among triem thepresence of paracrystalline arrays in tne vesselwall only at ~ hour and four hours post-occlusionperhaps-is a clue to the nature of the "signal instroke".

eould the cnange in shear stress on occlusionof ~l:A have released s erotonin, which polymerisedthe actin or activated the immune system, and the

. I . 1 ,,~ ?paracrystalline array as on y an umunog omn.

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SECOND WORLD CONGRESSOF STROKE, 1992

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Effect of brain-derived neurotrophic factor (BDNF)on neuronal death after cerebral ischemiaTetsuya Tsukahara, Yasuhiro Yonekawa, KimitoTanaka,Dept, of Cerebrovascular Surgery, National CardiovascularCenter, Suita, Osaka 565, Japan

We investigated the effect of brain-derived neurotrophic factor(BDNF) on the neuronal death which occurs after transientforebrain ischemia. Recombinant culture cells (CHOcell) withBDNF secreting ability were made by the transfection ofplasmid vector including BDNF cDNA. The activity ofrecombinant BDNF in the culture medium was first confirmedin vitro by observing the neurite growth of PCl2 cells and chickdorsal root ganglion cells. Delayed neuronal death wasinduced in the rat hippocampal CAl region by transientocclusion of bilateral carotid arteries (8 min) and lowering thesystemic blood pressure. The effect of BDNF on theneuronal death after ischemia was then examined by continuousintraventricular infusion of 200 p I of normal or concentratedrecombinant cell culture medium containing BDNF. Theculture medium of the recombinant cells including BDNF antisense eDNA was infused into the same ischemic brains, whichwere analyzed as a conrol group. Cell death of the CAlregion 7 days after the ischemia is summarized as follows:

Cell death; % of total CAl neuronsCulture medium Concentrated medium (x30)

each, n=6 BDNF anti-sense BDNF anti-senseMean ± SO 22±8 27±8 l±l*(p<O.OI) 22±5These results suggest that BDNF at sufficient dose haspreventive effect on the delayed hippocampal neuronal deathobserved after transient forebrain ischemia.

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CHRONIC ISRADIPINE ADMINISTRATIONPRODUCES DRAMATIC REDUCTION IN RATFOCAL ISCHEMIC DAMAGERaymond F. White, Giora Feuerstein and Frank C. Barone.SmithKline Beecham Pharmaceuticals, King of Prussia, PA,USA.

Isradipine (ISR;PN 200-110) antagonizes voltageassociated calcium influx into vascular smooth muscle andnerve cells. This action might increase cerebral blood flowand decrease toxic calcium influx into neurons, thusreducing tissue damage in focal ischemia. In the presentstudy, ISR (2.5 or 5.0 mg/kg, s.c.) or vehicle wasadministered chronically for 6 days before or 1 hr aftermiddle cerebral artery occlusion (MCAO) in spontaneouslyhypertensive rats. Contralateral forelimb (FND) andhindlimb (HND) neurological deficits were graded andbrains were analyzed for hemispheric infarct (%; HEM INF)and infarct volume (INF VOL; mm 3) 24 hr later. Results(*p<0.05 vs Vehicle, N=11-24/group; ANOVA) for chronictreatment was:GROUP HEMINF INFVOL fiill l:::!..t:{Q.VEHICLE 16±1 107±19 1.S±O .3 O.8±o .2ISR-2.5mglkg 7±1* 43±9· 0.3±O.2· O.O±O.O·ISR-5.0mglkg 3±1· 19±5· 0.5±O.3· 0.3±0.2·Reductions in infarction and neurological deficits wereproduced by acute, post-MCAO ISR administration.However, chronic -treatment produced a greater reductionin infarction (81±S%) than post-treatment (36±10%) .Chronic administration of ISR dramatically protects againstfocal ischemic injury, suggesting that the use of selecteddihydropyridines for hypertension can also provideneuroprotection in stroke.

Ischemic Neuronal Damage -A Morphological Study onEvolution of Damage in Hyperglycemia and Normoglycemia-Usuda K., Inamura Ie., Katayama Y. and Terashi A.Second Department of Internal Hedicine, NipponHedical School, Tokyo, Japan

The evolution of neuronal damage in variousregions during ischemia and in early recovery ",asinvestigated morphologically, using hyperglycemic andnormoglycemic Wistar rats.

Hyper glycemia (20-25 ~mol/ol plasoa) ",as achieved",ith infusion of glucose (Lv.) prior to ischemia.Forebain ischemia ",as induced by bilateral co mmoncarotid artery occlusion and hypotension.Normoglycemic rats ",ere fasted prior to ischemia.Ischemic changes of cells "'ere quantified by a fivepoint scale in the parietal cortex (Par),caudoputamen (CPu), globus pallidus (GP), hippocampus(HP) and substantia nigra (SN) during ischemia andthrough 120 minutes after recirculation.

I The hyperglycemic group (1) Par. CPu and liP;severely damaged neurons ",ere seen at 60-90 minutesafter recirculation, then there ",as some improvementat 120 minutes after recirculation. (2) GP; there",as little neuronal damages. (3) SN; illlI:lediatelyafter recirculation damaged neurons ",ere observed.and more damage ",as observed at 90 minutes postrecirculation. There "'as some improvement at 120minutes post recirculation. II The normoglycemicgroup No prominent neuronal damage was observed inany region.

Hyperglycemia exacerbated ischemic neuronaldamage. The evolution of neuronal damage ",as similarin the CPu. HP and Par regions. However, the SN ",asmore vulnerable and the GP ",as tolerant to ischemicinsult.

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THE EFFECT OF NIMODIPINE ON ACUTECEREBRAL VASOSPASM IN THE DOG

Robert N. Willette, Marcus P. Mitchell, Giora Z. Feuerstein,and Charles F. Sauermelch, Smith Kline BeechamPharmaceuticals, King of Prussia, PA, USA

The effects of nimodipine on acute cerebralvasospasm were examined angiographically in theanesthetized/ventilated dog. In this model, a singleintracisternal (ic) injection of autologous arterial blood (4ml), subsequent to the withdrawal of an equal volume ofcerebrospinal fluid, caused a consistent and progressivereduction in the cross sectional area of the basilar arteryand the adjacent anterior spinal artery in untreatedanimals (n=5) . The vasospasm was maximal in botharteries within one hour (30-40% of control), persisted forat least two hours and was not accompanied by asignificant increase in intracranial pressure. Similarresults were obtained in the vehicle treated group (n=6)which received a 30 minute intravenous (iv) infusion of5%-10% polyethylene glycol (0.1 mllmin) 30 minfollowing the ic administration of blood. In a separatetreatment group (n=5), the administration of nimodipine(10 Ilglkg, iv bolus followed by a 1·3 Ilglkg/min iv infusionfor 30 min) resulted in a 20% reduction in mean arterialpressure, but had no significant effect on the vasospasmobserved in the basilar and anterior spinal arteries .These results suggest that nimodipine does not affectacute cerebral vasospasm at moderate hypotensivedoses.

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111 and 31P-llIagnetie Resonance Spectroscopy ofCerebral Infarction in Rats

Yamamuro M, Igarashi.II,Yuasa T*,Katayama Y,Terashi AThe Seccind Department of Internal Medicine NipponMedical School, Department of KeurologyBrain Research Institute Niigata University*, JAPAN

Magnetic resonance spectroscopy (~IRS) al l ows thenoninvasive study of metabolism in vivo. In order tofurther understand the biochemical changes during anischemia of middle cerebral artery HICA) occlusion.The left MCA was occluded in male spontaneouslyhypertensive rats (SUR) weighing 200-250g using themethod of Tamura et al. Spectra lI"ere obtained fromthe infarcted hemisphere by placing the surfacecoils over the left side of the carvarium. 3lP and11l-~lRS were performed at 3 hours.24 hours and 7 daysafter MCA occlusion. A JOEL KMR system GX270(6.34T)was used. 3lP spectra were obtained using single onepulse sequence. III-MRS was performed using the spinecho sequence. Following ischemia the increase ininorganic phosphate (Pi) and the decrease in ATP andPCr were evident. After 7 days of ischemia thesechanges became obscure. The pili decreased after3hours ischemia and recovered to almost controllevels at 24 hrs post ischemia. Alkalosis wasapparent 7 days after ischemia. Although the lactatelevel was high 24 hrs post ischemia, the pili wasalmost normal. The K-aeetyl-aspartate/Creatine ratiodecreased significantly after ischemia.

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THE RAT FOREBRAIN IS INJURED MORE SEVERELYIN GLOBAL PARTIAL ISCHEMIA THAN IN GLOBALTOTAL ISCHEMIA

John Zinkel, MD, Fernando G. Diaz, MD, PhD

USA

In the awake rat, the forebrain can be isolatedfrom systemic circulation, subjected to global totalischemia, or to global partial ischemia (byextracorporeal perfusion), and recovered long term.Crucial to this ischemia technique is careful surgicalpreparation of the rat with full recovery before use.

At surgery, forebrain perfusion is reduced by 7permanent ligations essentially to that by the leftinternal carotid. Ligations are of the basilar and leftexternal carotid distally; the right internal carotid,both pterygopalatine and both occipital arteries,proximally. Femoral and left external carotid cathetersand loops for reversible bilateral common carotidocclusion are placed percutaneously. The next day,catheters are hooked to a perfusion pump. The pump isrun for 15 min with flow at O.2cc/min from leg tobrain. For controls (n-12), common carotids are leftpatent. For partial ischemia (n-12), carotid loops aretightened after 5 min pump perfusion for 10 min andreleased. For total ischemia (n-12), the pump isstopped at 5 min, and carotid loops are tightened for10 min, and released. Rats are recovered, and 72 hrslater, sacrificed by in vivo perfusion fixation, andtheir brains sliced at 6 um, and stained with Hand E.

At recovery, rats were active, and nonfocal. Atischemia, rats were decerebrate with EEG's flat by 30sees., and respiration at 55-60fmin. At histology,::ontrols brains were normal. Ischemic brains showedlaminar necrosis. As by Pulsinelli ('82), neocorticalinj ury in total ischemia was grade 2 or 3, and inpartial ischemia, grade 3 or 4. CAl hippocampal injuryin total ischemia was grade 3, and in partial ischemia,$rade 4. Implications will be discussed.

Antiatherogenic Action of Calcium Channel Blockade onVascular Smooth Muscle Cell

Rita Alam, Satoshi Kataoka, Frank M. Yatsu, Houston,Texas, USA.

Clentiazem (TA-3090, 10'51\1), a calcium channel blocker,significantly inhibits 3H-Thymidine incorporation (53%) invascular smooth muscle cell (VSMC). Platelet derivedgrowth factor (PDGF:40mg/ml) increases DNA synthesisin VSMC 39% higher than control, whereas TA-3090reduces PDGF-induced 3H-Thymidine incorporation by42%. TA-3090 does not affect inositol phosphates (IPsIP!, IPz, IP3, and IP~) production and does not significantlyreduce PDGF-induced IPs production. TA-3090 increasesLDL receptor activity by 3.4 times but does notsignificantly affect cholesterol synthesis (HC-acetateincorporation). Our results indicate that anantiatherogenic effect of TA-3090 is associated withreduced cell proliferation, determined by DNA synthesis,and up regulation of LDL receptor, without affecting thePDGF-induced signal transducing pathway. Currentstudies on the effects of TA-3090 on the proto-oncogeneC-fos and c-jun and the AP-l genomic transcription factormay clarify the molecular biological mechanisms.

PO-C57

Immunohistochemical charachteristics of humanatherosclerotic carotid plaques.B.Gallicchio, F.Maiuri,S.Montagnani*, E.Di Vaia*,A. Bernardo,L. L.Serra,Inst. of Neurosurgery and Ins t , of Anatomy*2nd School of Medicine Naples - Italy

We submitted samples from about twentyatherosclerotic plaques from hu~an carotid arteriesto an immunohistochemical pannel of monoclonal andpoliclonal antibodies. Our purpose was to clarifysome variations in cell population normally presentin arterial ual.l: and to define characteristics ofinfiltrating cells involved in the development ofthe disease. Recently it was evidentiated by manyAuthors the role that smooth muscle cells could playin atherosclerosis either with proliferation or withsome unespected changes of their immunologicalmarkere i In fact,a great numberof smooth musclecells in atherosclerotic vasal wall show expressionof the MCH class II (Ia) antigens in addition tonormal smooth muscle markers as desmin and smoothmyosin. Our, results confirm these data and define aprevalence of T-lynphocytes and related cell in theplaque infiltrate. It is hypotized that not onlymacrophages but also smooth muscle cells may play arole Of antigen presentation in the development ofat.heroealeroe io disease and are corresponeabl:e ofmorpho-functional aspects of this pathology.

Date post:	03-Jan-2017
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