HYPERTENSION AND CEREBRAL ISCHEMIA/Hayakawa et al. 267

17. Ogata J, Fujishima M, Morotomi Y, Omae T: Cerebral infarc-tion following bilateral carotid artery ligation in normotensiveand spontaneously hypertensive rats: a pathological study.Stroke 7: 54-60, 1976

18. Jones JV, Fitch W, MacKenzie ET, Strandgaard S, HarperAM: Lower limit of cerebral blood flow autoregulation in ex-perimental renovascular hypertension in the baboon. Circ Res39: 555-557, 1976

19. Strandgaard S: Autoregulation of cerebral blood flow in hyper-

tensive patients: the modifying influence of prolonged anti-hypertensive treatment in the tolerance to acute, drug-inducedhypertension. Circulation 53: 720-727, 1976

20. Ooshima A, Fuller G, Cardinale G, Spector S, Udenfriend S:Collagen biosynthesis in blood vessels of brain and other tissuesof the hypertensive rat. Science 190: 898-900, 1975

21. Laurent JP, Molinari GF, Oakley JC: Primate model ofcerebral hematoma. J Neuropathol Exp Neurol 35: 560-568,1976

A New Model of Bilateral Hemispheric Ischemiain the Unanesthetized Rat

W I L L I A M A. P U L S I N E L L I , M . D . , P H . D . A N D J A M E S B. B R I E R L E Y , M . D .

SUMMARY A new model of transient, bilateral hemispheric ischemia in the unanesthetized rat is described.During ether anesthesia the rat's vertebral arteries were electrocauterized through the alar foramina of thefirst cervical vertebra and reversible clasps placed loosely around the common carotid arteries. Twenty-four hrlater, the awake rats were restrained and the carotid clasps tightened to produce 4-vessel occlusion. Thecarotid clasps were removed after 10,20 or 30 min of 4-vessel occlusion and the animals killed by perfusion fix-ation 72 hr later. Rats which convulsed during the ischemic or post-ischemic period were excluded from furtherstudy. All rats subjected to 20 or 30 min of 4-vessel occlusion demonstrated ischemic neuronal damage. The HIand paramedian hippocampus, striatum and layers 3, 5 and 6 of the posterior neocortex were the regions mostfrequently damaged. The advantages of this model are the ease of preparation of large numbers of animals, ahigh rate of predictable ischemic neuronal damage, a low incidence of seizures and the absence of anesthesia.

Stroke Vol 10, No 3, 1979

THE STUDY of the pathophysiology of cerebralischemia is limited by the lack of a small animalmodel uncomplicated by anesthesia, systemic hypox-ia, hypotension or generalized seizures. Cerebralhypoxia-ischemia in the rat represents one well-studied preparation where cerebral energymetabolism1"4 and histopathology57 are concerned.The rat is sufficiently large to permit easy monitoringof physiological variables (body temperature, elec-troencephalogram, arterial pressure and arterial gas-es) yet small enough to be used in the number requisitefor statistical analysis without excessive costs. Anefficient collateral circulation in the rat prevents eitherunilateral or bilateral carotid artery ligation from con-sistently altering cerebral metabolism8 or producingischemic brain damage.9 Present methods to producecerebral ischemia in this animal require systemic in-sults of hypoxia3-7> 10 and/or hypotension,8' n

anesthesia either at the initiation of ischemia or

From the Cornell University Medical College, 1300 York Ave.,New York, NY 10021 (Dr. Pulsinelli) and the Medical ResearchCouncil Laboratories, Carshalton, Surrey, U.K. (Dr. Brierley).

This investigation was supported in part by U.S. Public HealthService Grant No. NS-03346. Dr. Pulsinelli is a recipient of aNINCDS Research Service Award No. NS-05833 and a Teacher-Scientist Award of the Andrew W. Mellon Foundation.

A preliminary report of this model was presented at the AmericanAcademy of Neurology (Neurology-Minneap-28: 379, 1978).

Reprints: Dr. Pulsinelli, Department of Neurology, CornellUniversity Medical College, 1300 York Ave., New York, NY10021.

throughout the ischemic interval,3'8> n~13 and, finally,the ischemia is frequently permanent12- " rather thantransient.

Occlusion of all 4 major arteries supplying the ratbrain might decrease cerebral blood flow sufficientlyto produce ischemic changes. We report here amethod for producing reversible common carotidartery occlusion, which, combined with permanent in-terruption of the vertebral arteries in the awake, free-running rat, results in bilateral hemispheric ischemiawith a high incidence of predictable brain damage andwithout the complications of previous models in smallanimals.

Methods

Male Wistar rats weighing between 250 and 300 gmwere allowed free access to Purina Laboratory chowand tap water in day-night regulated quarters at 24°C. They were anesthetized with ether and both com-mon carotid arteries isolated via a ventral, midlinecervical incision. An atraumatic arterial clasp5 wasloosely placed around each common carotid arterywithout interrupting carotid blood flow and the inci-sion closed with a single suture. A second incision, 1cm in length, was made behind the occipital bonedirectly overlying the first two cervical vertebrae. Theparaspinal muscles were separated from the midlineand with the use of an operating microscope, the rightand left alar foramina of the first cervical vertebraewere exposed. As described by Green15 the rat'svertebral arteries travel within the vertebral canal and

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268 STROKE VOL 10, No 3, MAY-JUNE 1979

FIGURE 1. Dorsal view of rat skull plus first and secondcervical vertebrae, a. alar foramina of first cervicalvertebrae, v. vertebral arteries as they pass rostrally throughvertebral canal and beneath alar foramina.

pass beneath the alar foramen before entering into theposterior fossa (fig. 1). A 0.5 mm diameter elec-trocautery needle (Bowie Monopolar Electrocautery,Cincinnati, Ohio) was inserted through each alarforamen and both vertebral arteries electrocauterizedand permanently occluded. The electrocautery wasgrounded through the animal's foreleg withoperational settings that produced minimal, localmuscle contraction.

While the rats were still anesthetized, stainless steelscrews were mounted on the calvarium to record theelectroencephalogram (EEG) and a tail artery cannulainserted to record the blood pressure, Poa, Pco2 andpH. The cannula was secured under the animal's skinand kept patent overnight by injecting a dilute solutionof heparin (2 units/ml) into the cannula tip which wasthen sealed. The rats were allowed to recover fromanesthesia for 24 hours at which time they were in-distinguishable from normal animals by clinical andEEG criteria. The awake rats were then hand-held in asimple restraint, the ventral neck suture removed andboth carotid clasps were tightened to produce 4-vesselocclusion. Body temperature was monitored and/ormaintained at 37° with a rectal thermistor (YellowSprings Inst., Yellow Springs, Ohio) coupled to aheating lamp. Carotid clasps were removed following10, 20 or 30 min of 4-vessel occlusion and restorationof carotid blood flow was verified by direct observa-tion. The animals were observed, clinically, for theirlevel of consciousness, the presence or absence of acorneal reflex, their ability to walk and to climb.

At 72 hr of postischemic survival the rats wereanesthetized and their brains perfusion-fixed with

FAM (40% formaldehyde: glacial acetic acid:methanol, 1:1:8) via the ascending aorta after briefly(30 sec) washing out the cephalic circulation (ab-dominal aorta clamped) with heparinized,physiological saline. The brains were left in situ for1-4 hours at 4°C before removal and then stored inFAM. Paraffin sections (7 and 12 /xm) were stainedwith cresyl violet, Luxol fast blue and cresyl violet,phosphotungstic acid and hematoxylin, and withhematoxylin and eosin. The sections were examinedwith the light microscope and ischemic neuronaldamage graded on a scale of 0 = normal, 1 = a fewneurons damaged, 2 = many neurons damaged,3 = majority of neurons damaged.

Results

With this method of 4-vessel occlusion, 98 of 127rats (77%) became unresponsive and lost their rightingreflex within 15-30 sec after bilateral carotid arteryligation; 19 (15%) of the rats became only lethargicand the remaining 10 (8%) died within 2-3 min fromrespiratory failure. In contrast, bilateral carotid arteryligation in 10 rats with intact vertebral arteriesresulted in no change in their level of consciousness.The 4-vessel occluded rats that became only lethargicwere excluded from further studies. Following com-pletion of this study the incidence of hemisphericischemia in this model, i.e. the rapid loss of con-sciousness and the inability to right upon carotidartery ligation, decreased below the 77% level quotedabove. Although all rats were purchased from thesame supplier, inbreeding may have improved thecollateral supply to the brain in the latter animals.Wistar rats from several suppliers were tested beforefinding a strain which resulted in a similar 70-80%success rate observed earlier. Payan et al." reportedsimilar variability in the incidence of brain damagebetween several strains of rats subjected to permanentbilateral carotid artery ligation. Differences incollateral blood supply to the brain were most likelyresponsible for this latter observation. These collateralvessels probably originate from cervical branches ofthe subclavian arteries.

Four-vessel occlusion resulted in hyperventilationwith a mild elevation of arterial Po,, a moderatehypocapnia and respiratory alkalosis (fig. 2). Themean arterial blood pressure rose 20-30 mm Hg im-mediately after bilateral carotid artery ligation,presumably as a consequence of reduced carotidbaroreceptor discharge. Arterial blood gas values,hydrogen ion concentration, and blood pressurereturned to control values within 15 min of carotidclasp removal. Mean rectal temperature decreased1°C during 4-vessel occlusion and returned to controlvalues after 4 hours. The EEG became isoelectricwithin 2-3 min of 4-vessel occlusion (fig. 3) in thoseanimals that became unresponsive and lost theirrighting reflex. The EEG remained isoelectricthroughout the ischemic period. Intermittent slowwaves appeared after 15 min of reperfusion but theEEG remained abnormally slow for at least 4 to 6 hr.

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NEW MODEL OF BILATERAL HEMISPHERIC ISCHEMIA/Pulsinelli et al. 269

BPmm Hg 140<n=10)

•ao

Temp -C T l II(n-6) 37J-T T y - - ^ i

I I I0 15 30• 1

c o n t r o 1 clamp

30

unclamp

45 60 ' 120Time (min)

180 240

FigjURE 2. Physiological variables in the rat prior to, dur-ing (shaded) and following 4-vessel occlusion. Vertical barsrepresent mean ±1 SE of arterial Po2, PcoipH and BP.

Clinically, the degree of neurological deficit wasproportional to the duration of ischemia. The animalscontinued to breathe spontaneously and never losttheir corneal reflex during the ischemic orpostischemic period suggesting that the brainstem wasunaffected. Animals subjected to 10 min and 20 min of4-vessel occlusion developed no permanent neurologicdeficit other than bilateral ptosis which resulted fromcarotid artery manipulation. Animals subjected to 30min of 4-vessel occlusion remained unresponsive for

EEC

Control

Ischemia0-30 min

25MV|

1 second

Post-ischemia:15 min -

up to 2 hr after release of the carotid clasp; spon-taneous movements were negligible for up to 6 hr and,although they could walk by 12 hr, the majority neverregained normal motor activity. The percentage ofanimals that convulsed during a 72 hr survival periodfollowing 10 min or 20 min of 4-vessel occlusion was0% and 8% respectively. Following 30 min of ischemia20% of the rats convulsed by 24 hr and 40% by 72 hr ofsurvival. All rats which convulsed during the post-ischemic period were excluded from further study.

An estimate of cerebral perfusion was made by in-jecting a blue dye into the ascending aorta at 120-130mm Hg. This was done in control rats, in rats witheither the carotid or vertebral arteries occluded and inrats with 4-vessel occlusion. Well-perfused areas of thebrain stained dark blue while ischemic areas appearedpale. When only the carotid or vertebral arteries wereoccluded, perfusion of all brain structures was wellmaintained (fig. 4A). Four-vessel occlusion resulted ina marked decrease in perfusion of the cerebralhemispheres but did not produce complete ischemia,i.e. a colorless brain. Perfusion of the brain stem in 4-vessel occluded rats was preserved but was less than incontrols (fig. 4B). Perfusion of the cerebellum andolfactory lobes was variable but usually preservedeven when the cerebral hemispheres were ischemic(fig. 5). Blood flow to the brainstem and cerebellum in4-vessel occluded rats was probably due to filling viathe anterior spinal artery. The source of blood flow tothe olfactory lobes in this model remains unknown.

Histological examination of the brains from 8 con-trol (vertebral arteries electrocauterized and claspsplaced loosely around both carotids) rats showed nopathological alterations. Neuropathological damagein brains from ischemic rats was characterized byischemic cell change with and without incrustationsand homogenizing changes in neurons as describedpreviously by Brown and Brierley5'16>" in otherischemic models. All animals exposed to 20 or 30 minof 4-vessel occlusion demonstrated ischemic neuronal

FIGURE 3. Electroencephalograms represen-

tative of rats which became unresponsive andlost their righting reflex within 15-30 seconds of4-vessel occlusion. Post-ischemia = aftercarotid clasp removal.

180 min

240 min

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270 STROKE VOL 10, No 3, MAY-JUNE 1979

BFIGURE 4. Dorsal (A j and ventral (B) view of brains from 4 rats perfused with a blue dye viathe ascending aorta, a. control rat, b. bilateral carotid artery ligation, c. bilateral vertebralartery occlusion, d. bilateral carotid and vertebral artery occlusion. Note that the cerebellum(Ad) is partially perfused and that there is dye in the basilar artery and circle of Willis of thisbrain (Bd).

damage; the grade of damage varied among regionsand was proportional to the duration of ischemia. Thetable demonstrates the distribution and incidence ofgrade 2-3 damage in rats subjected to 10, 20 and 30min of 4-vessel occlusion. The HI and paramedian(PM) hippocampus were the most vulnerable struc-tures. Ten minutes of 4-vessel occlusion resulted ingrade 2-3 damage to 40% of these structures while theremaining regions were less frequently damaged.Following 20 min of ischemia 85%-9O% of the HI andPM areas of the hippocampus demonstrated grade

2-3 damage. When 4-vessel occlusion was increased to30 min there was an increase in the incidence of grade2-3 damage in the striatum (93%) and thalamus(61%), the latter being focal in nature. Neocorticaldamage was limited to layers 3, and/or 5 and 6, theposterior cortex being more frequently involved thanthe anterior cortex. The cerebellum was infrequentlydamaged while the brainstem appeared histologicallynormal in all animals except one which had grade 3damage of the right substantia nigra. The integrity ofthe cerebellum and brainstem was attributed to perfu-sion via the anterior spinal artery.

FIGURE 5. Dorsal view of brains of 3 rats perfused with ablue dye during 4-vessel occlusion. Note that the ischemia isnot complete. The cerebellum and olfactory lobes arevariably perfused while the cerebral hemispheres appearpale.

TABLEDamage

Percentage of Hemispheres with Grade SS lachemic

Region

Ant. Cortex(Layers 3, 5, 6)

Poat. Cortex(Layers 3, 5, 6)

Striatum

HI Hippocampus

H3-5 Hippocampus

PM Hippocampus

Thalamus

Cerebellum

Brainstem

Operatedcontrols*

0

0

00

0

0

000

7210 MintHohemia

10

15

0

40

25

40

fl

0

0

Hour survival20 Miniischomia

37

47

35

85

55

90

0

10

0

30 Miniisohemia

57

64

93

100

79

93

61

36

7

* 8 rats X Qeft and right) — 16 hemisphere*UO rmta X (left and right) - 20 hemispheres|14 rats X (left and right) - 28 hemispheres

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Comment

The study of pathophysiological mechanisms ofischemic brain damage in small animals has beencomplicated by factors such as anesthetics, systemichypoxia, hypotension and generalized seizures. Theadvantage of the Mongolian gerbil is that ischemicbrain damage can be produced by carotid occlusionper se as a consequence of a functionally incompletecircle of Willis.18'19 Its disadvantage is that only30-40% of gerbils treated this way will develop clinicalsigns of hemispheric ischemia.18'19 Moreover, the ger-bil is genetically predisposed to convulsions inducedby mild stimuli20-21 and following unilateral carotidartery ligation 75% of clinically affected animalsdeveloped generalized seizures.19-22 Since seizures in-crease cerebral energy metabolism23 they may jeopar-dize neurons in the ischemic and postischemic periods.In man, seizures are a rare complication (<10%) ofischemic stroke24-26 and therefore exclusion of con-vulsing animals in pathophysiological studies ofischemic brain damage is justified.

Various methods are available for producingcerebral hypoxia-ischemia in small animals. However,most animals have a well-developed circle of Willisand, therefore, unilateral or bilateral carotid arteryligation must be combined with hypoxia and/orhypotension to consistently derange cerebralmetabolism and produce ischemic damage. Systemichypoxia and hypotension may damage organs (heart,kidney) other than brain and thereby alterphysiological mechanisms of postischemic survival.Moreover, the above methods, as well as cerebral em-bolization12 and middle cerebral artery occlusion,13

employ anesthetics which may depress cerebral energymetabolism,26-27 modify ischemic brain damage28 andhinder attempts to ameliorate post-ischemicencephalopathy. The method described herein of 4-vessel occlusion in the rat is devoid of systemic hypox-ia, hypotension or anesthetic drugs; there is asufficiently low incidence of seizures to permit exclu-sion of convulsing animals; the procedure is surgicallysimple so that large numbers of animals can beassessed statistically; there is a high incidence ofpredictable ischemic neuronal damage in certainregions of the brain. Regions most vulnerable toischemic damage in this model were the HI, H3-5,and paramedian zones of the hippocampus, layers 3, 5and 6 of the posterior neocortex, and the striatum.This pattern of selective vulnerability is similar to thatfound in other animal models of hypoxia-ischemia.29

The method of initial, permanent occlusion of thevertebral arteries permits control of cerebral circula-tion by the occlusion of the common carotid arteriesalone or with the addition of cervical branches of thesubclavian arteries. The surgical approach and elec-trocauterization of the vertebral arteries through thealar foramen or groove of the first cervical vertebrae issimple, atraumatic, and may be accomplished beforecarotid artery occlusion. This 2-stage procedureshould be applicable to other animals having a com-plete circle of Willis providing collateral circulation is

adequate to maintain vital brain stem centers. Occlu-sion of the vertebral arteries does not compromise car-diorespiratory centers in the dog30-31 or monkey.32 Itremains to be determined whether the brain stem ofadditional animals will tolerate permanent vertebralartery occlusion. If this were so, permanent occlusionof the vertebral arteries under anesthesia could befollowed by carotid artery occlusion using techniques(pneumatic cervical cuff, arterial snare) that do notrequire anesthesia.

References

1. Clendenon N, Allen N, Komatsu T et al: Biochemicalalterations in the anoxic-ischemic lesion of rat brain. ArchNeurol 25: 432-448, 1971

2. Siesjo B, Nilsson L: The influence of arterial hypoxemia onlabile phosphates and on extracellular lactate and pyruvate con-centrations in rat brain. Scand J Clin Lab Invest 27: 83-96,1971

3. Salford L, Plum F, Siesjo B: Graded hypoxia-oligemia in ratbrain. Arch Neurol 29: 227-233, 1973

4. Salford L, Siesjo B: The influence of arterial hypoxia and uni-lateral carotid artery occlusion upon regional blood flow andmetabolism in rat brain. Acta Physiol Scand 92: 130-141, 1974

5. Brown A, Brierley J: The nature, distribution and earliest stagesof anoxic-ischemic nerve cell damage in the rat brain as definedby the optical microscope. Brit J Exp Path 49: 87-106, 1968

6. McGee-Russell S, Brown A, Brierley J: A combined light andelectron microscope study of early anoxic-ischemic cell changein rat brain. Brain Res 20: 193-200, 1970

7. Salford L, Plum F, Brierley J: Graded hypoxia-oligemia in ratbrain: neuropathological alterations and their implications.Arch Neurol 29: 234-238, 1973

8. Eklof B, Siesjo B: The effect of bilateral carotid ligation uponthe blood flow and energy state of the rat brain. Acta PhysiolScand 86: 155-165, 1972

9. Payan H, Levine S, Strevel R: Effects of cerebral ischemia invarious strains of rats. Proc Soc Exp Biol Med 120: 208-209,1965

10. Levine S: Anoxic-ischemic encephalopathy in rats. Am J Pathol36: 1-17, 1960

11. Nordstrom C, Rehncrona S: Postischemic cerebral blood flowand oxygen utilization rate in rats anesthetized with nitrous ox-ide or phenobarbital. Acta Physiol Scand 101: 230-240, 1977

12. Kogure K, Busto R, Scheinberg P: Energy metabolites andwater content in rat brain during the early stage of developmentof cerebral infarction. Brain 97: 103-114, 1974

13. Ljunggren B, Ratcheson R, Siesj5 B: Cerebral metabolic statefollowing complete compression ischemia. Brain Res 73:291-307, 1974

14. Robinson R, Bloom F: Pharmacological treatment of abehavior disorder following experimental stroke. In ScheinbergP (ed) Cerebrovascular Disease, Tenth Princeton Conference.New York, Raven Press, pp 199-206, 1976

15. Greene E: Anatomy of the rat. Transactions Am PhilosophicalSoc, 1935

16. Brown A, Brierley J: Anoxic-ischaemic cell change in rat brainlight microscopic and fine-structural observations. J Neurol Sci16: 59-84, 1972

17. Brown A, Brierley J: The earliest alterations in rat neurons andastrocytes after anoxia-ischemia. Acta Neuropathol 23: 9-22,1973

18. Levine S, Sohn D: Cerebral ischemia in infant and adult gerbils.Relation to incomplete circle of Willis. Arch Pathol 87:315-317, 1969

19. Levy D, Brierley J, Plum F: Ischemic damage in the gerbil inthe absence of "no-reflow." J Neuro Neurosurg Psych 38:1197-1205, 1975

20. Loskota W, Lomax P: The Mongolian gerbil as a model for thestudy of epilepsies. Electroenceph Clin Neurophysiol 381:597-604, 1975

21. Seaman R, Seaman S, Sun A: Neurochemical correlates in

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272 STROKE VOL 10, No 3, MAY-JUNE 1979

seizure prone gerbils. Society for Neuroscience, Abstr 7th AnnMeeting, Vol III: p 145, 1977

22. Yanigihara T: Experimental stroke in gerbils: correlation ofclinical, pathological and electroencephalographic findings andprotein synthesis. Stroke 9: 155-159, 1978

23. Howse D, Caronna J, Duffy T, Plum F: Cerebral energymetabolism, pH and blood flow during seizures in the cat. Am JPhysiol 227: 1444-1456, 1974

24. Richardson E, Dodge P: Epilepsy in cerebrovascular disease,Epilepsia 3 (3rd series): 49-74, 1954

25. Louis S, McDowell F: Epileptic seizures in nonembolic cerebralinfarction. Arch Neurol 17: 414-418, 1967

26. Gatfield P, Lowry O, Schultz D, Passonneau J: Regionalenergy reserves in the mouse brain and changes with ischemiaand anesthesia. J Neurochem 13: 185-195, 1966

27.

28

29

Michenfelder J, Theye A: Cerebral protection by thiopentalduring hypoxia. Anesthesiology 39: 510-517, 1973Smith A: Barbiturate protection in cerebral hypoxia.Anesthesiology 47: 285-293, 1977Brierley JB: Cerebral hypoxia. In: Blackwood W, Corsellis AN(eds) Greenfield's Neuropathology. London, Edward ArnoldPublishers Ltd., pp 43-85, 1976

30. Kabat H, Dennis C: Decerebration in the dog by complete tem-porary anemia of the brain. Proc Soc Exptl Biol Med 38:864-865, 1938

31. Boyd R, Connally J: Total cerebral ischemia in the dog. ArchSurg 84: 434-438, 1962

32. Donald D, White T: Temporary bilateral occlusion of the com-mon carotid and vertebral arteries in the monkey at normalbody temperature. Neurology (Minneap) 11: 836-838, 1961

Survival of Rabbits After Prolonged Cerebral Ischemia

RONALD J. KOLATA, DVM

S U M M A R Y Cerebral ischemia was produced by a combination of vascular occlusion and mild systemichypotension in 2 groups of rabbits. Arterial blood pressure, arterial pH, arterial blood gases, blood glucose andPCV were monitored and recorded before, during and for 3 hours after reperfusion. Return of EEC activity,vasomotor control, spontaneous ventilation and corneal reflex were also recorded. At 4, 8 , 1 2 , 24 and 48 hoursafter reperfusion, the rabbits' neurologic status was assessed according to an arbitrary scale based on motorfunction. The 2 groups differed in return of reflexes and motor function. Eighty percent of the rabbits ischemicfor 20 minutes and 75% of the rabbits ischemic for 30 minutes survived. The graduated response of motor func-tion to cerebral ischemia is attributed to the ventilatory and circulatory support given the rabbits for the first 3hours after reperfusion. The graduated response of motor function to ischemia supports the suggestion thatmotor function can be used as an index of neurologic damage.

Stroke Vol 10, No 3, 1979

A VARIETY OF METHODS to control the cerebralcirculation of rabbits has been used to study theeffects of cerebral ischemia (CI) in these animals. Themethods vary in their complexity. Simple clamping ofthe aorta just distal to the coronary sinus can causeCI.1 If a left ventricular outflow reservoir is also used,fatal cardiac dilatation is prevented and reversiblecerebral ischemia is achieved.2 Most other methods ofachieving CI involve arterial ligation and one or moreother maneuvers. One method involves permanentbasilar artery ligation, temporary bilateral carotidocclusion, and inflation of a cervical pressure cuff tocause CI.4 A simplified method employs bilateral tem-porary carotid artery occlusion and drug-inducedprofound systemic hypotension.5 A similar and moresimplified method uses a cervical pressure cuff inflatedto 1.5 atm pressure to cause CI.6 A method which doesnot employ arterial occlusion but achieves CI by usingdrug-induced profound hypotension, tilting the rabbithead up and ventilating it with a mixture of 96% N2and 4% O2 has been described.7 These methods havebeen classified and commented on.8 Some have to be

From the University of Pennsylvania School of VeterinaryMedicine.

This study was supported by U.S. Navy Contract N62269-76-C-0206.

Dr. Kolata's present address is the College of VeterinaryMedicine, University of Georgia, Athens, GA 30602.

criticized because they require extensive or traumaticsurgery to gain control of the cerebral circulation.Some of them, also, cause hypoxemia and ischemia oforgans in addition to the brain, which could result inartifactual responses during recovery.

The method described herein overcomes many ofthese criticisms as it requires minor surgical interven-tion, allows venous return from the brain, maintainsadequate blood oxygenation, and preserves adequatesystemic blood pressure.

Methods

Surgical Preparation

Each rabbit is given 0.2 mg of atropine kg/bodyweight. Anesthesia is induced with a gas mixture ofhalothane and oxygen. After intubation, anesthesia ismaintained with the same mixture. The rabbit isprepared and draped for aseptic surgery. A 3 cm inci-sion is made in the lateral surface of the neck begin-ning at the caudal edge of the transverse process of the1st cervical vertebra. The subcutaneous tissues are in-cised, and the splenius muscle is identified andreflected dorsally to expose the intertransversariuscervicus dorsalis muscle. The intertransversarius cer-vicus dorsalis and the intertransversarius intermediusmuscles are separated to expose the vertebral artery asit passes out of the transverse foramen of the secondcervical vertebra. The artery is identified and ligated

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W A Pulsinelli and J B BrierleyA new model of bilateral hemispheric ischemia in the unanesthetized rat.

Print ISSN: 0039-2499. Online ISSN: 1524-4628 Copyright © 1979 American Heart Association, Inc. All rights reserved.

is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231Stroke doi: 10.1161/01.STR.10.3.267

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