Expressions of P-selectin- and HSP72-like immunoreactivities in rat brain after transient middle cerebral artery occlusion

The role of an adhesion molecule such as P-selectin may be important in the pathogenesis of stroke. However, temporal, spatial and cellular profiles of the expression of such a protein have not been fully studied. Change of immunoreactive P-selectin was examined in rat brain after transient middle cerebral artery (MCA) occlusion in comparison with that of 72 kDa heat shock protein (HSP72) which is a well known marker of cell injury. Western blot analyses were performed to ensure the selective detection of immunoreactive P-selectin and HSP72 proteins with each antibody using brain samples before and after ischemia. Temporal, spatial and cellular changes of immunohistochemical expressions of P-selectin and HSP72 were evaluated with rat brain sections at 2 and 8 h, and 1, 3 and 7 days of reperfusion after 1 h of MCA occlusion (MCAO). Hematoxylin-eosin (HE) staining was performed to evaluate brain cell damage at 3 and 7 days of reperfusion. Western blot showed a single band at molecular weights of 140 and 72 kDa for P-selectin and HSP72, respectively, only after ischemia. No significant band was observed without primary antibody. P-selectin-like immunoreactivity was not normally present in rat brain sections. However, it was expressed mainly in the post-capillary venules of the cerebral cortex and caudate in the MCA territory with a peak at 8 h to 1 day. The expression was diminished by 3 days of reperfusion. An immunoreactive HSP72 was scarcely present in the cerebral cortex and caudate of the sham control brain. However, the protein was induced in neurons of the MCA territory. The HSP72 induction was gradually intensified from 8 h with peaks at 1 day in the cortex and at 3 days in the caudate. The immunoreactivity decreased by 7 days. Histopathological study with HE staining showed no evident cell damage at 3 and 7 days of reperfusion. The present results indicate that temporal, spatial and cellular differences were present in the expressions of immunoreactive P-selectin and HSP72 proteins. P-selectin was expressed from an earlier stage of reperfusion in post-capillary venules, and the expression became maximum at the same time both in the cerebral cortex and caudate. In contrast, HSP72 induction began later in neurons and reached maximum at a different time between the cortex and caudate.     

The effect of dopamine depletion on the H2O2 production in the rat striatum following transient middle cerebral artery occlusion

The changes in the extracellular concentrations of rat striatal H2O2, dopamine (DA) and its metabolites during middle cerebral artery (MCA) occlusion and reperfusion were simultaneously examined by microdialysis, and the relationship between the ischemia-induced release of DA and the generation of H2O2 was estimated by assessing the effect of the lesion of the substantia nigra (SN). In the rats without SN lesions, a significant increase in the striatal H2O2 level was observed during the ischemia and reperfusion phases. In the rats with SN lesions, the ischemia-induced H2O2 production was not attenuated. These results suggest that DA is not an important source of H2O2 in cerebral ischemia and reperfusion.     

Incomplete infarct and delayed neuronal death after transient middle cerebral artery occlusion in rats

BACKGROUND AND PURPOSE: The clinical syndrome of transient ischemic attacks is accompanied in a significant percentage of patients by brain lesions or neuroimaging abnormalities whose structural counterparts have not been defined. The objective of this study was to analyze, in an experimental model of short-term (< 25 minutes) focal ischemia and long-term (< or = 28 days) reperfusion, the extent and nature of the structural abnormalities affecting neurons and glia located within the territory of the transiently occluded artery. METHODS: Adult Wistar rats (n = 121) had the origin of one middle cerebral artery (MCA) occluded with a nylon monofilament for periods of 10 to 25 minutes. Experiments of transient MCA occlusion were terminated at variable periods ranging from 1 day to 4 weeks. Control experiments consisted of (1) MCA occlusion without reperfusion (n = 7) lasting 7 to 14 days and (2) sham operations (n = 2) followed by 1- to 4-day survival. After in situ fixation, brain specimens were serially sectioned and subjected to detailed morphometric evaluations utilizing light and electron microscopes. The statistical method used to evaluate the results was based on ANOVA followed by Bonferroni's corrected t test and Student's t test comparisons. RESULTS: Brain lesions were not detectable in the sham-operated controls. All brains with permanent MCA occlusion (7 to 14 days) had large infarctions with abundant macrophage infiltration and early cavitation. Forty-five (37%) of the experiments involving transient MCA occlusion had no detectable brain lesions after 4 weeks. Selective neuronal necrosis was found in 76 of 121 rats (63%) with transient MCA occlusion. Neuronal necrosis always involved the striatum, and in 29% of the brains with ischemic injury, necrosis also included a short segment of the cortex. In the striatum, the length of the arterial occlusion was the main determinant of the number of necrotic neurons (20 minutes [22.6 +/- 19] is worse than 10 minutes [4.9 +/- 7]) (P < .0001). In the cortex, the length of reperfusion determined the number of necrotic neurons appearing in layer 3. Experiments with reperfusion of 4 to 7 days' duration yielded more necrotic neurons per microscopic field (2.02 +/- 3) than those lasting fewer days (0.04 +/- 0.1) (P < .05). The histological features of these lesions underwent continuous change until the end of the fourth week, at which time necrotic neurons were still visible both in the striatum and in the cortex. CONCLUSIONS: Arterial occlusions of short duration (< 25 minutes) produced, in 76 of 121 experiments (63%), brain lesions characterized by selective neuronal necrosis and various glial responses (or incomplete infarction). This lesion is entirely different from the pannecrosis/cavitation typical of an infarction that appears 3 to 4 days after a prolonged arterial occlusion. Delayed neuronal necrosis, secondary to a transient arterial occlusion or increasing numbers of necrotic neurons in experiments with variable periods of reperfusion, was a response observed only at a predictable segment of the frontoparietal cortex.     

Cathepsin B and middle cerebral artery occlusion in the rat

Lysosomal proteases, although tightly regulated under physiological conditions, are known to contribute to cell injury after various forms of tissue ischemia have occurred. Because cathepsin B is a prominent lysosomal protease found in brain parenchyma, the authors hypothesized that it may contribute to neuronal cell death after focal cerebral ischemia. The authors measured the expression and spatial distribution of cathepsin B within the ischemic brain in 43 animals by means of immunohistochemical analysis in a rat model of transient middle cerebral artery (MCA) occlusion. Cathepsin B activity was also measured within specific ischemic brain regions by using an in vitro assay (22 animals). In addition, the authors tested the therapeutic effect of preischemic intraventricular administration of stefin A, a cysteine protease inhibitor, on the volume of cerebral infarction after transient MCA occlusion (15 animals). Increased cathepsin B immunoreactivity was detected exclusively within the ischemic neurons after 2 hours of reperfusion following a 2-hour MCA occlusion. Cathepsin B immunolocalization in the ischemic region decreased by 24 hours of reperfusion, but then increased by 48 hours of reperfusion because the infarct was infiltrated by inflammatory cells. Increased immunolocalization of cathepsin B in the inflammatory cells located in the necrotic infarct core continued through 7 days of reperfusion. Cathepsin B enzymatic activity was significantly increased in the ischemic tissue at 2, 8, and 48 hours, but not at 24 hours of reperfusion after 2 hours of MCA occlusion. Continuous intraventricular infusion of stefin A, before 2 hours of MCA occlusion (15 animals), significantly reduced infarct volume compared with control animals (12 animals): the percentage of hemispheric infarct volume was 20+/-3.9 compared with 33+/-3.5 (standard error of the mean; p = 0.025). These data indicate that neuronal cathepsin B undergoes increased expression and activation within 2 hours of reperfusion after a 2-hour MCA occlusion and may be a mechanism contributing to neuronal cell death. Intraventricular infusion of stefin A, an inhibitor of cathepsin B, significantly reduces cerebral infarct volume in rats.     

Assessment of sensorimotor neglect after occlusion of the middle cerebral artery in the rat

Evaluating the efficacy of neuroprotective drugs in rat models of focal cerebral ischemia has involved histological and behavioral batteries to examine treatment outcome. However, the behavioral tests used to date provide little insight into the nature of the neurological impairments. To provide an analysis of a possible "neglect" syndrome after occlusion of the middle cerebral artery, M. I. Posner's (1980) visual attentional paradigm was adapted for use in the rat. A paw-reaching task and a test of somatosensory "neglect" also were used to assess forelimb sensorimotor function. The lesion group displayed unilateral deficits; however, there was no evidence of attentional dysfunction. Results are consistent with the conclusion that the behavioral deficits identified arise from a somatosensory deficit rather than hemineglect due to dysfunctional spatial attention.     

Regional cerebral blood flow during and after 2 hours of middle cerebral artery occlusion in the rat

In this study we explored if the secondary bioenergetic failure, which occurs a few hours after recirculation, following transient middle cerebral artery occlusion (MCAO) in rats, is caused by a compromised reflow. We induced 2 hours of MCAO and measured CBF at the end of the ischemia, as well as 15 minutes, 1, 2, and 4 hours after the start of recirculation, using autoradiographic or tissue sampling 14C-iodoantipyrine techniques. After 2 hours of MCAO, the autoradiographically measured CBF in the ischemic core areas was reduced to 3 to 5% of contralateral values. The reduction in CBF was less in neighboring, penumbral areas. After recirculation, flow already normalized in core tissues after 15 minutes, and remained close to normal for the 4 hours recirculation period studied. However, in penumbral tissues, recovery CBF values were usually below normal. The results show that tissues that are heavily compromised by the 2-hour period of ischemia and are destined to incur infarction, show a "relative hyperemia" during recirculation. In fact, some areas of the previously densely ischemic tissue showed overt hyperperfusion. This finding raises the question whether the relative or absolute hyperemia reflects events that are pathogenetically important. Because drugs that clearly ameliorate the final damage incurred fail to alter the relative hyperperfusion of previously ischemic tissues, it is concluded that vascular events in the reperfusion period do not play a major role in causing the final damage.     

Specific induction of protein kinase C delta subspecies after transient middle cerebral artery occlusion in the rat brain: inhibition by MK-801

Protein kinase C (PKC) consists of a family of closely related Ca2+/phospholipid-dependent phosphotransferase isozymes, most of which are present in the brain and are differentially activated by second messengers. Calcium-dependent PKC activity may cause neuronal degeneration after ischemic insult. PKC is also involved in trophic-factor signaling, indicating that activity of some PKC subspecies may be beneficial to the injured brain. Therefore, we screened long-term changes in the expression of multiple PKC subspecies after focal brain ischemia. Middle cerebral artery occlusion was produced by using an intraluminal suture for 30 min of 90 min. In in situ hybridization experiments, mRNA levels of PKC alpha, -beta, -gamma, -delta, -epsilon, and -zeta were decreased in the infarct core 4 hr after ischemia and were lost completely 12 hr after ischemia. In areas surrounding the core, PKC delta mRNA was specifically induced 4, 12, and 24 hr after ischemia in the cortex. At 3 and 7 d, the core and a rim around it showed increased mRNA levels of PKC delta. No other subspecies were induced. At 2 d, immunoblotting demonstrated increased levels of PKC delta protein in the perifocal tissue, and immunocytochemistry revealed an increased number of PKC delta-positive neurons in the perifocal cortex. In the core, PKC delta-positive macrophages and endothelial cells were seen. Pretreatment with MK-801, an NMDA antagonist, inhibited cortical PKC delta mRNA induction. The data show that focal brain ischemia induces PKC delta mRNA and protein but not other PKC subspecies through the activation of NMDA receptors and that the upregulation lasts for several days in neurons of the perifocal zone.     

Long-term functional outcome following transient middle cerebral artery occlusion in the rat: Correlation between brain damage and behavioral impairment.

The assessment of both histological and functional long-term outcomes after cerebral ischemia is increasingly recommended for preclinical studies. Whereas correlations between behavioral impairments and primary ischemic lesion are documented, little is known about their relationships with remote nonischemic regions that undergo secondary degeneration, such as the thalamus. Anesthetized rats were subjected to mild (30 min) or severe (60 min) occlusion of the middle cerebral artery. Two months after ischemia, sensorimotor behavior was assessed according to the neurological score, limb-placing, adhesive-removal, and staircase tests; the final histological lesion was measured after this assessment. Cortical damage was correlated to all transient and long-lasting sensorimotor deficits, whereas striatal lesion was more consistently reflected by the forelimb-placing reflexes and adhesive-removal motor deficits. By contrast, the thalamic atrophy was not correlated to early neurological impairment, but rather to the late sensory deficit at the adhesive-removal test and to the skilled forepaw reaching alteration at the staircase test. This suggests that thalamus contributes, albeit moderately, to the ischemia-induced long-lasting sensorimotor deficits, some of which represent relevant targets for therapeutic interventions. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Structural alterations in synaptosomal membrane-associated proteins and lipids by transient middle cerebral artery occlusion in the cat

We have previously reported that ischemia reperfusion injury results from free radical generation following transient global ischemia, and that this radical induced damage is evident in the synaptosomal membrane of the gerbil. [Hall et al, (1995) Neuroscience 64: 81-89]. In the present study we have extended these observations to transient focal ischemia in the cat. We prepared synaptosomal membranes from frontal, parietal-temporal, and occipital regions of the cat cerebral cortex with reperfusion times of 1 and 3 hours following 1 hour right middle cerebral artery occlusion. The membranes were selectively labeled with protein and lipid specific paramagnetic spin labels and analyzed using electron paramagnetic resonance spectrometry. There were significant motional changes of both the protein and lipid specific spin labels in the parietal-temporal and occipital regions with 1 hour reperfusion; but, both parameters returned to control values by 3 hours reperfusion. No significant changes were observed in the normally perfused frontal pole at either reperfusion time. These results support the argument that free radicals play a critical role in cell damage at early reperfusion times following ischemia.     

Monofilament intraluminal middle cerebral artery occlusion in the mouse

The rat middle cerebral artery (MCA) occlusion model with an intraluminal filament is well characterized with a two hour period of occlusion in widespread use. The recent availability of transgenic animals has led to an interest in adapting the MCA model in the mouse. To date the model has not been well characterized in the mouse. We performed the present study to compare different durations of MCA occlusion and to validate new functional assessments in this model. The MCA occlusion model (5-0 filament) was used. Swiss-Webster mice, 24-44 g, were randomly assigned to four groups: one hour of occlusion; two hours of occlusion; three hours of occlusion; or permanent occlusion. At 48 hours post-ischemia, the animals were rated on three neurologic function scales, and then the brains were removed for lesion size determination. Overall, there was a significant difference in lesion volume (p < 0.001) between the groups. In the permanent group of mice, the average lesion volume was 78.41 +/- 17.47 mm (n = 12); two and three hours of ischemia produced 51.29 +/- 29.82 mm3 (n = 11) and 54.85 mm3 (n = 13), respectively, significantly different than the one hour group 14.84 +/- 31.34 mm3 (n = 11). All three functional scoring systems found significant overall differences between the four groups with our detailed General and Focal scores producing more robust between group treatment differences and showing correlation coefficients of r = 0.766 and r = 0.788, respectively to infarct volume. The MCA filament occlusion model can be successfully adapted in the mouse with either two or three hour occlusions producing reliable infarcts. New functional scoring systems unique to the mouse appear to add additional information.     

Spatial stability of extracellular potassium ion and blood flow distribution in rat cerebral cortex after permanent middle cerebral artery occlusion

Extracellular potassium ion activity ([K+]o) increases precipitously during brain ischemia when blood flow falls below threshold values less than approximately 15 mL/100 g/min. This flow threshold for increase of [K+]o occurs also in focal ischemia producing gradient from ischemic core to adjacent normally perfused brain. In this study we investigated the spatial and temporal stability of extracellular potassium ion and blood flow gradients after permanent middle cerebral artery occlusion (MCAO) in rats. [K+]o and regional CBF were measured, respectively, with K+-sensitive and polarographic hydrogen-sensitive microelectrodes at different cortical locations in the middle cerebral artery distribution region. Spatial assessment of [K+]o and regional CBF was conducted at 30, 90, and 180 minutes after MCAO. [K+]o in the more lateral cortex (core) increased from near 3 mmol/L before MCAO to greater than 50 mmol/L and was associated with flow values less than 25% of pre-ischemic levels. Measurements medial to the core (penumbra) indicated progressively decreasing levels of [K+]o and improvement of CBF. There was a tendency for [K+]o in penumbral zones to decrease toward normal levels with time, but there was little dissipation of [K+]o in core regions. In contrast, the spatial CBF profile remained remarkably constant for the entire recording period. Thus, unlike infarction which has been reported to expand with time after focal ischemia, the spatial [K+]o disturbance tends to contract primarily due to decreasing [K+]o with time in the penumbra. Thus, steady state levels of [K+]o after focal ischemia may not be a valuable predictor of cell viability.     

Collateral hemodynamics after middle cerebral artery occlusion in Wistar and Fischer-344 rats

We investigated whether the difference in infarction volume after occlusion of a long proximal segment of the middle cerebral artery between Wistar and Fischer-344 rats, is caused by differences in collateral blood flow rate through leptomeningeal anastomoses. In view of the retrograde direction of collateral blood flow into the middle cerebral artery territory, we developed parasagittal laser-Doppler flowmetry. Using this method two laser-Doppler probes are placed on the cerebral cortex: probe 1 is placed near the anastomoses between the middle- and anterior cerebral artery, probe 2 is placed 2 mm further away from these anastomoses than probe 1. We found in both rat strains a comparable relation between the areas under the curve of the signal measured by both laser-Doppler probes for 2 h after middle cerebral artery occlusion. This relation is considered to be a measurement of the collateral blood flow rate into the middle cerebral artery territory through leptomeningeal anastomoses after middle cerebral artery occlusion. We conclude that collateral blood flow for the two strains were essentially similar for the initial 2 h after MCA occlusion. Although these collateral blood flows could have been different at a later time, it is unlikely that the interstrain difference in cerebral infarction volume between Wistar and Fischer-344 rats after proximal middle cerebral artery occlusion is caused by an apparent interstrain difference in the magnitude of collateral blood flow rate through leptomeningeal anastomoses. The parasagittal laser-Doppler flowmetry technique we developed for these experiments is currently successfully used in our laboratory to evaluate the efficacy of hemodynamically active pharmacotherapeutical agents in raising the collateral blood flow rate into the middle cerebral artery territory after middle cerebral artery occlusion.     

Electrophysiological studies of rat substantia nigra neurons in an in vitro slice preparation after middle cerebral artery occlusion

We studied sequential changes in electrophysiological profiles of the ipsilateral substantia nigra neurons in an in vitro slice preparation obtained from the middle cerebral artery-occluded rats. Histological examination revealed marked atrophy and neurodegeneration in the ipsilateral substantia nigra pars reticulata at 14 days after middle cerebral artery occlusion. Compared with the control group, there was no significant change in electrical membrane properties and synaptic responses of substantia nigra pars reticulata neurons examined at one to two weeks after middle cerebral artery occlusion. On the other hand, there was a significant increase in the input resistance and spontaneous firing rate of substantia nigra pars compacta neurons at 13-16 days after middle cerebral artery occlusion. Furthermore, inhibitory postsynaptic potentials evoked by stimulation of the subthalamus in substantia nigra pars compacta neurons was suppressed at five to eight days after middle cerebral artery occlusion. At the same time excitatory postsynaptic potentials evoked by the subthalamic stimulation was increased. Bath application of bicuculline methiodide (50 microM), a GABA(A) receptor antagonist, significantly increased the firing rate of substantia nigra pars compacta neurons from intact rats. These results strongly suggest that changes in electrophysiological responses observed in substantia nigra pars compacta neurons is caused by degeneration of GABAergic afferents from the substantia nigra pars reticulata following middle cerebral artery occlusion. While previous studies indirectly suggested that hyperexcitation due to deafferentation from the neostriatum may be a major underlying mechanism in delayed degeneration of substantia nigra pars reticulata neurons after middle cerebral artery occlusion, the present electrophysiological experiments provide evidence of hyperexcitation in substantia nigra pars compacta neurons but not in pars reticulata neurons at the chronic phase of striatal infarction.     

Enhanced protein synthesis in the ipsilateral substantia nigra following middle cerebral artery occlusion in the rat

The purpose of this randomised single blinded study was to determine the optimal size of laryngeal mask airway in the normal adult population, to test the validity of the current selection criteria and to determine if any externally measured anatomical variable correlated with optimal size. In each of 30 apnoeic anaesthetised adults weighting less than 100 kg, size 3, 4 and 5 laryngeal mask airways were inserted in random order by a skilled user and the cuff inflated to a standard pressure (60 cm H2O). Optimal size was based on four criteria in order of priority: number of attempts at placement, oropharyngeal leak pressure, fiberoptic score and percentage of vocal cords seen. The size 5 laryngeal mask airway was optimal in 19/30 and the size 4 in 11/30. In no patient was the size 3 the optimal fit. Oropharyngeal leak pressure was significantly higher for each progressively large size and the fiberoptic view was significantly better for the size 4 and size 5. There was no significant predictive value in any externally measured anatomical variable, but height was the most useful. The best current selection strategy was to choose a size 5 for males and size 4 for females. Potentially useful new strategies may be to use the size 5 in all adults, or a size 5 > or = 165 cm in height and size 4 for < 165 cm. We conclude that predicting the optimal size of laryngeal mask airway for individual adult patients is complex. The best size selection strategies involve use of the size 4 and 5 laryngeal mask airways in adults.     

Increased endothelin-1 in the rabbit model of middle cerebral artery occlusion

Human neutrophils (5 x 10(4) incubated on fibronectin precoated wells released 2.83 +/- .25 nmoles of superoxide (0(2)-) (x +/- 1 SEM, n = 15) in response to 5.9 nM (100 ng/ml) Tumor Necrosis Factor Alpha (TNF). On the contrary, the 0(2)- production induced by interleukin-8 (IL-8) (doses ranging from 0.1 nM to 1 microM) was comparable to that of "resting" cells (< .6 nmoles/5 x 10(4) cells). IL-8 (100 nM) did not affect the TNF-dependent 0(2)- production when added with TNF at the beginning of the assay, but reduced it by approximately 80% when added with TNF on neutrophils previously incubated for 1 hour on fibronectin. As compared with IL-8, N-formyl-methionyl-leucyl-phenylalanine (FMLP, 100 nM) failed to suppress the TNF-triggering of the oxidative burst in neutrophils plated on fibronectin. The data suggest that the interaction of neutrophils with fibronectin uncovers the capacity of IL-8 to limit the cell response to TNF, without affecting the response to the combination of FMLP and TNF. Thus, although the chemotactic factors IL-8 and FMLP share the capacity of triggering the oxidative burst of neutrophils incubated in suspension, only IL-8 has the potential to down-regulate the responsiveness of fibronectin-adherent cells to TNF.     

Diffusion, perfusion, and T2 magnetic resonance imaging of anti-intercellular adhesion molecule 1 antibody treatment of transient middle cerebral artery occlusion in rat

The effect of anti-intercellular adhesion molecule-1 (anti-ICAM-1) antibody treatment of transient (2 h) middle cerebral artery (MCA) occlusion in the rat was measured using diffusion (DWI)-, T2 (T2I)- and perfusion (PWI)-weighted magnetic resonance imaging. Rats were treated upon reperfusion with an anti-ICAM-1 monoclonal antibody (n=11) or a control antibody (n=7). DWI, T2I and PWI were performed before, during, and after induction of focal cerebral ischemia from 1 h to 7 days. In both groups, the apparent diffusion coefficient of water (ADCw) and cerebral blood flow (CBF) values in the ischemic region significantly declined from the preischemic ADCw values (p<0. 05). The post ischemic increase in T2 of the control group was significantly higher at 48 h than in the anti-ICAM-1 treated group (p<0.05). CBF was not significantly different between the two groups. The temporal profiles of MRI cluster analysis, which combines ADCw and T2 maps into a single image, was significantly different between groups. These data suggest that the neuroprotective effect of anti-ICAM-1 antibody treatment is reflected in reductions of T2 and lesion growth during reperfusion and may not be associated with increased cerebral perfusion.     

Changes in local cerebral blood flow, local EEG, and flow in the distal stump of the middle cerebral artery in cats with occlusion of the middle cerebral artery

The local EEG, the local cerebral blood flow (lCBF), and the flow in the distal stump of the occluded middle cerebral artery was simultaneously recorded in 28 acute experiments in cats. Nembutal anaesthesia was used eleven times, and Halothane anaesthesia 17 times. The recordings were made via platinum electrodes: 12 in the ischaemic hemisphere, and 2-3 in the opposite non-ischaemic hemisphere. The flow in the occluded middle cerebral artery was recorded via a platinum electrode introduced into this artery via the transorbital approach. The changes in lEEG, lCBF, and middle cerebral artery flow were studied during normotension, hypertension, and hypotension. A beneficial effect of hypertension was noted in the acute phase of brain ischaemia. Hypertension counteracted also the "diaschisis" in the non-ischaemic part of the ischaemic hemisphere and in the opposite non-ischaemic hemisphere. A correlation between lEEG changes and lCBF changes was noted. In addition, an interesting discrepancy was observed between the rapid H2 clearance in the middle cerebral artery stump and the much slower H2 clearance in the ischaemic brain area. Significant differences between experiments under Halothane and experiments under Nembutal anaesthesia were noted. In the acute phase those changes are probably the result of the different levels of blood pressure in those two groups.     

An integrated analysis of the progression of cell responses induced by permanent focal middle cerebral artery occlusion in the rat

Defining the chronology and severity of cell damage in an evolving lesion after ischemia is important for understanding the underlying mechanisms in the development of therapeutic intervention. In the present study, we used a combination of histological and immunocytochemical methods to evaluate cell responses from 30 min to 48 h after permanent occlusion of the middle cerebral artery (MCAO) in the rat. Specific immunocytochemical markers clearly revealed acute early responses in neurons (neurofilament protein 200), astrocytes (glial fibrillary acidic protein), and microglia/macrophages (OX-42 and ED-1) such as enlarged, convoluted neuronal processes, and disintegration of glia. Progressive topographic changes in the developing lesion, pinpointed by immunolabeling, indicated the severity and extension of the cell damage. Proliferation and hypertrophy of astrocytes and microglia around the infarct, and contralaterally, occurred 24-48 h after MCAO and coincided with mass necrosis and infiltration of neutrophils and macrophages into the core. These observations corroborate the suggestion that the inflammatory process is involved in the progression of the infarct.     

Early assessment of motor dysfunctions aids in successful occlusion of the middle cerebral artery

Occlusion of the rodent middle cerebral artery by embolism, using an intraluminal filament, produces behavioral alterations which resemble many symptoms associated with stroke. This model has been used to examine treatment interventions for the disease, however, variable success rate in completely blocking the middle cerebral artery may present inconclusive interpretation of the data. To detect successful occlusion of the middle cerebral artery, we demonstrate here sensitive and reliable behavioral parameters including the elevated body swing test, the postural tail-hang test, the spontaneous rotational test, and the forelimb akinesia test. These assays provide a criterion for identifying animals with incomplete occlusion which could promote host-related spontaneous recovery and might confound true effects of experimental therapies on ischemia-induced dysfunctions. From a practical standpoint, the early reliable identification of partial cerebral ischemia aids in immediate and efficient adjustments of the surgical procedure to create a complete and stable ischemia stroke animal model.     

Testosterone increases and estradiol decreases middle cerebral artery occlusion lesion size in male rats

This study was undertaken to determine the effects of estrogen and testosterone on cerebral ischemic lesion size induced by middle cerebral artery (MCA) occlusion in male rats. Rats were gonadectomized and treated with testosterone, estrogen, or testosterone plus estrogen filled Silastic pellets. The animals were divided into 6 groups: intact, intact + estrogen (E2), castrate, castrate + testosterone (T), castrate + E2, and castrate + T + E2. One week after treatment, cerebral ischemia was induced by MCA occlusion for 40 min, followed by reperfusion. After 24 h, rats were sacrificed and slices were then stained to assess lesion size. The presence of testosterone increased and the removal of testosterone decreased lesion size. A strong positive correlation (r2 = 0.922) between plasma testosterone concentrations and ischemic lesion size was observed. Estradiol treatment reduced ischemic area. In summary, the present study provides evidence that testosterone exacerbates and estrogens ameliorate ischemic brain damage in an animal model of cerebral ischemia.