Lesions In Rat Brain Research Articles

The chemokine SDF‐1/CXCL12 contributes to the ‘homing’ of umbilical cord blood cells to a hypoxic‐ischemic lesion in the rat brain

Previous studies have shown that transplanted human umbilical cord blood (hUCB)-derived mononuclear cells exert therapeutic effects in various animal models of CNS impairments, including those of perinatal hypoxic-ischemic brain injury. However, the mechanisms of how transplanted cells exert their beneficial effects on the damaged tissue are still unclear. As detection of hUCB cells at the lesion site coincides with the therapeutic effects observed in our model, we investigated the role of the chemokine stromal derived factor (SDF)-1 (CXCL12) as a possible candidate for chemotaxis-mediated 'homing' of transplanted hUCB cells to a hypoxic-ischemic lesion in the perinatal rat brain. Following the hypoxic-ischemic insult expression of SDF-1 significantly increased in lesioned brain hemispheres and was mainly associated with astrocytes. Transplanted hUCB cells expressing the SDF-1 receptor CXCR4 migrated to the lesion site within one day. Inhibition of SDF-1 by application of neutralizing antibodies in vivo resulted in a significantly reduced number of hUCB cells at the lesioned area. The increase in glial SDF-1 expression shortly after induction of the lesion and hUCB cells expressing the corresponding receptor makes SDF-1 a potential chemotactic factor for hUCB cell migration. The reduction of hUCB cells present at the lesion site upon functional inhibition of SDF-1 strengthens the view that the SDF-1/CXCR4 axis is of major importance for cell 'homing'.

Fractalkine and CX3CR1 are involved in the migration of intravenously grafted human bone marrow stromal cells toward ischemic brain lesion in rats

Recent research has shown that transplanted bone marrow stromal cells (MSCs) migrate to the injured regions and exert their therapeutic effects in cases of intracranial trauma, stroke, inflammation and degenerative disease. The specific mechanisms involved in their migration to lesions are still to be fully elucidated. In the present study, a rat model of transient middle cerebral artery occlusion (MCAO) was established. At 24 h after reperfusion, human bone marrow stromal cells (hMSCs) were transplanted by intravenous injection to explore the effects of fractalkine/CX3CR1 on the migration of transplanted MSCs to lesions. In vitro study using real-time PCR and western blot revealed that CX3CR1, the only known receptor of fractalkine, was expressed in cultured hMSCs. The expression of fractalkine in the ischemic brain was significantly increased. The directional migration of transplanted hMSCs to the damaged region was observed through detection of green fluorescence protein (GFP). The results indicated the cells were mainly distributed in the ischemic boundary zone with high fractalkine expression. In a further study, lentivirus-mediated RNA interference of CX3CR1 expression was employed. The results of these experiments indicated that CX3CR1 knock-down dramatically decreased the migration of hMSCs to the ischemic brain. The present study suggests that fractalkine and its specific receptor CX3CR1 are involved in the directional migration of transplanted MSCs to the ischemic damaged brain region.

Design and fabrication of a polyimide-based microelectrode array: Application in neural recording and repeatable electrolytic lesion in rat brain

The design and testing of a new microelectrode array, the NCTU (National Chiao Tung University) probe, was presented. Evaluation results showed it has good biocompatibility, high signal-to-noise ratio ( SNR: the root mean square of background noise to the average peak-to-peak amplitude of spikes) during chronic neural recordings, and high reusability for electrolytic lesions. The probe was a flexible, polyimide-based microelectrode array with a long shaft (14.9 mm in length) and 16 electrodes (5 μm-thick and 16 μm in radius); its performance in chronic in vivo recordings was examined in rodents. To improve the precision of implantation, a metallic, impact-resistant layer was sandwiched between the polyimide layers to strengthen the probe. The three-dimensional (3D) structure of electrodes fabricated by electroplating produced rough textures that increased the effective surface area. The in vitro impedance of electrodes on the NCTU probe was 2.4 ± 0.52 MΩ at 1 kHz. In addition, post-surgical neural recordings of implanted NCTU probes were conducted for up to 40 days in awake, normally behaving rats. The electrodes on the NCTU probe functioned well and had a high SNR (range: 4–5) with reliable in vivo impedance (<0.7 MΩ). The electrodes were also robust enough to functionally record events, even after the anodal current (30 μA, 10 s) was repeatedly applied for 60 times. With good biocompatibility, high and stable SNR for chronic recording, and high tolerance for electrolytic lesion, the NCTU probe would serve as a useful device in future neuroscience research.

Systemic Inflammatory Response Reactivates Immune-Mediated Lesions in Rat Brain

The potential association between microbial infection and reactivation of a multiple sclerosis (MS) lesion is an important issue that remains unresolved, primarily because of the absence of suitable animal models and imaging techniques. Here, we have evaluated this question in an empirical manner using immunohistochemistry and magnetic resonance imaging (MRI), before and after the induction of a systemic inflammatory response in two distinct models of MS. In a pattern-II-type focal myelin oligodendrocyte glycoprotein-experimental autoimmune encephalomyelitis model, systemic endotoxin injection caused an increase in regional cerebral blood volume (rCBV) around the lesion site after 6 h, together with a reduction in the magnetization transfer ratio of the lesioned corpus callosum. These changes were followed by an increase in the diffusion of tissue water within the lesion 24 h after endotoxin challenge and new leukocyte recruitment as revealed both immunohistochemically and by MRI tracking of ultrasmall superparamagnetic iron oxide-labeled macrophages. Importantly, we detected in vivo expression of E- and P-selectin in quiescent lesions by MRI-detectable glyconanoparticles conjugated to sialyl Lewis(X). This finding may explain, at least in part, the ability of quiescent MS lesions to rapidly reinitiate the cell recruitment processes. In a pattern-I-type delayed-type hypersensitivity response model, a similar effect of endotoxin challenge on rCBV was observed, together with delayed breakdown of the blood-brain barrier, showing that systemic infection can alter the pathogenesis of MS-like lesions regardless of lesion etiology. These findings will have important implications for the management and monitoring of individuals with MS.

Transient Widespread Blood—Brain Barrier Alterations after Cerebral Photothrombosis as Revealed by Gadofluorine M-Enhanced Magnetic Resonance Imaging

Magnetic resonance imaging (MRI) is a powerful tool to assess brain lesions, but currently available contrast agents are limited in the assessment of cellular and functional alterations. By use of the novel MRI contrast agent gadofluorine M (Gf) we report on imaging of transient and widespread changes of blood-brain barrier (BBB) properties as a consequence of focal photothrombotic brain lesions in rats. After i.v. application, Gf led to bright contrast in the lesions, but also the entire ipsilateral cortex on T1-weighted MRI. In contrast, enhancement after application of gadolinium diethylenetriamine-pentaacetic acid (Gd-DTPA), a common clinical indicator of BBB leakage was restricted to the lesions. Remote Gf enhancement was restricted in time to the first 24 h after photothrombosis and corresponded to a transient breakdown of the BBB as revealed by extravasation of the dye Evans blue. In conclusion, our study shows that Gf can visualize subtle disturbances of the BBB in three dimensions not detectable by Gd-DTPA. Upon entry into the central nervous system Gf most likely is locally trapped by interactions with extracellular matrix proteins. The unique properties of Gf hold promise as a more sensitive contrast agent for monitoring BBB disturbances in neurologic disorders, which appear more widespread than anticipated previously.

Epileptogenesis after cortical photothrombotic brain lesion in rats

We investigated epileptogenesis after cortical photothrombotic stroke induced with Rose Bengal dye in adult Sprague–Dawley rats. To detect spontaneous seizures, video-electroencephalograms were recorded at 2, 4, 6, 8, and 10 months for 7–14 days (24 h/day). At the end, spatial and emotional learning and memory were assessed using the Morris water-maze and fear-conditioning test, respectively, and the brains were processed for histologic analysis. Seizures were detected in 18% of rats that received photothrombosis. The average seizure frequency was 0.39 seizures per recording day and mean seizure duration was 117 s. Over 60% of seizures occurred during the dark hours. Rats with photothrombotic lesions were impaired in the water-maze (P<0.05) but not in the fear-conditioning test as compared with controls. Histology revealed that lesion depth varied from cortical layers I to VI in photothrombotic rats with epilepsy. Epileptic rats had light mossy fiber sprouting in the inner molecular layer of the dentate gyrus both ipsilateral and contralateral to the lesion. This study extends the current understanding of epileptogenesis and functional impairment after cortical lesions induced by photothrombosis. Our observations support the hypothesis that photothrombotic stroke in rats is a useful animal model for investigating the mechanisms of post-stroke epileptogenesis.

Robert William Kerwin

When Rob Kerwin arrived at the Maudsley to begin his psychiatric training his bosses were secretly in awe of him. For he was in the vanguard of a new breed...

Treatment of hyponatraemia by urea decreases risks of brain complications in rats. Brain osmolyte contents analysis

Brain damage (myelinolysis) develops in hyponatraemia after a large increase in serum sodium regardless of the currently available methods of correction. However, a preliminary study suggests that treatment of hyponatraemia with urea limits the risks of brain lesions in rats. Benefits of sustained high blood levels of urea and mechanisms of protection remain hypothetical. In the first part of the study, hyponatraemic rats received repeated (i.p.) doses of urea (2 g/kg b.w./6 h) leading to sustained blood levels (urea+/-230 mg/dl). Neurologic outcome was compared to correction of hyponatraemia by water diuresis. In the second part of the study, we analysed the adaptative response of the brain to correction of hyponatraemia with either urea or water diuresis, by measurement of cerebral osmolyte contents. Despite a large correction of the serum sodium (mean Delta SNa=32 mEq/l/24 h), mortality rate (13%) and neurological symptoms were low in the urea-treated group contrary to control groups treated by water diuresis (mortality: 87%). This shows with stronger evidence the protective effect of urea against brain myelinolysis. In the second part, analysis of brain composition shows that, in the urea groups, 24 h after correction, intracerebral hyperionization (NaCl) was avoided and brain water contents remained normal. No significant changes of the major brain organic osmolyte composition were observed after urea administration except reaccumulation of betaine. No difference in brain composition was noted regarding concomitant plasma urea levels (>or<150 mg/dl). In rats treated by water diuresis, the brain remained also significantly depleted in organic osmolytes 24 h after correction, but contrary to administration of urea, this treatment was associated with a high mortality rate. These comparative results suggest a specific brain-protective effect of urea itself against myelinolysis.

Expression of heparanase in nestin-positive reactive astrocytes in ischemic lesions of rat brain after transient middle cerebral artery occlusion

Heparanase is an enzyme that cleaves heparan sulfate proteoglycans, an important component of the extracellular matrix to generate heparan sulfate fragments, leading to the remodeling of the extracellular matrix and the basement membrane particularly during cancer metastasis. A growing body of evidence suggests that heparanase serves multiple functions in normal tissues including the central nervous system. In this study, we showed that heparanase is expressed in reactive astrocytes in the peri-infarct lesion of a rat brain whose middle cerebral artery was transiently occluded for 90 min. RT-PCR and Western blot analyses revealed that heparanase expression was markedly upregulated during the subacute phase of ischemia (from 3 to 7 days post-reperfusion (dpr)). As revealed by immunohistochemical study, heparanase was localized in astrocytes located in the peri-infarct region. Heparanase + astrocytes expressed nestin that is known as a marker of reactive astrocytes. Infiltrated neutrophils were weakly heparanase +. After 7 dpr, the expression level of heparanase + astrocytes considerably decreased. Therefore, the maximum expression of heparanase by astrocytes may correlate with the time of migration of reactive astrocytes toward the ischemic core, which may result in astrogliosis. These findings suggest a novel role of heparanase in the pathophysiology of brain ischemia.

Angiogenesis and improved cerebral blood flow in the ischemic boundary area detected by MRI after administration of sildenafil to rats with embolic stroke

To dynamically investigate the long-term response of an ischemic lesion in rat brain to the administration of sildenafil, male Wistar rats subjected to embolic stroke were treated with sildenafil ( n = 11) or saline ( n = 10) at a dose of 10 mg/kg administered subcutaneously 24-h after stroke and daily for an additional 6 days. Magnetic resonance images were acquired and functional performance was measured in all animals at 1 day, 2 days and weekly for 6 weeks post-stroke. All rats were sacrificed 6 weeks after stroke and endothelial barrier antigen immunostaining was employed for morphological analysis and quantification of cerebral vessels. Map-ISODATA was computed from T 1, T 2 and T 1sat maps. ISODATA derived tissue signatures characterize the degree of ischemic injury. Based on the map-ISODATA calculated at 6 weeks, the ischemic lesion for each animal was divided into two specific regions, the ischemic boundary and ischemic core. The temporal profiles of cerebral blood flow (CBF) and tissue signature were retrospectively tracked in these two regions and were compared with histological evaluation and functional outcome. After 1 week of sildenafil treatment, the ischemic lesion exhibited two significantly different regions, with higher CBF level and correspondingly, lower tissue signature value in the boundary region than in the core region. Sildenafil treatment did not significantly reduce the lesion size, but did enhance angiogenesis. Functional performance was significantly increased after sildenafil treatment compared with the control group. Administration of sildenafil to rats with embolic stroke enhances angiogenesis and selectively increases the CBF level in the ischemic boundary, and improves neurological functional recovery compared to saline-treated rats.

The histopathological and electron-microscopic examination of the stereotactic pulsed radiofrequency and conventional radiofrequency thermocoagulation lesions in rat brain

Objective: Neurodestructive procedures have been used for treating intractable pain for a long time. Pulsed radiofrequency (RF) is a newly defined energy type. Pulsed RF may be used in the treatment of patients with some pain syndromes in whom the pain could not be controlled by the alternative techniques. The objective of the present study was to examine the histological and electron microscopical changes in rat brain after pulsed RF application.Methods: Forty-five male rats were used in these experiments. Lesions were applied stereotactically to the target areas of the rat brains. Two different RF energy type were used as representative models of pulsed-RF and conventional-RF procedures. The rats were kept alive for 21 days and then killed. The effect of pulsed RF lesions on cerebral tissue ultrastructure was studied.Results: In the pulsed RF group, intracytoplasmic edema, clarity of the mitochondrial cristas and opening in the cell membrane pores were observed on the electron microscopic examination. In the conventional RF group, these findings were more prominent. In the pulsed RF group, the ratio of the effected neurons was 5.5% on light microscopic examination. In the conventional RF group, the ratio of the effected neurons was 14.26% and central necrosis was observed additionally.Discussion: Pulsed RF caused ultrastructural changes in the neurons. The pulsed RF may possibly cause a depression on the cell membrane potential by opening the cell membrane pores and resulting in the ion entrance into the cell cytoplasm and intracytoplasmic edema. However, it seems that all these changes were reversible.

Temporal profile of connexin 43 expression after photothrombotic lesion in rat brain

Following focal ischemic injury, several mechanisms lead to secondary expansion of the affected area and therefore increase the initial damage. We thoroughly investigated the expression of astrocytic connexin 43 (Cx43) after photothrombosis in rat brain. The temporal profile of Cx43 mRNA as well as protein expression was studied in remote, structurally uninjured cortical and hippocampal areas. The hippocampal formation revealed an increased number of Cx43 mRNA positive astrocytes and an up-regulated protein expression exclusively in the ipsilateral stratum oriens. We assume a participation of this region in glia scar formation. While Cx43 mRNA positive cells were transiently increased, immunoreactivity was reduced in the somatosensory cortex of injured hemispheres. The observed decrease of Cx43 protein in the post-ischemic cerebral cortex implies an impairment of gap junctional intercellular communication which might be detrimental to the brain.

Ischemic Cerebral Tissue Response to Subventricular Zone Cell Transplantation Measured by Iterative Self-Organizing Data Analysis Technique Algorithm

To investigate the changes of the ischemic lesion in rat brain after subventricular zone (SVZ) cell transplantation and the influence of the grafted cells on the appearance of angiogenesis, SVZ cells, superparamagnetically labeled, were intracisternally transplanted into the rat brain 48 h after onset of embolic stroke. A complete set of magnetic resonance (MR) images was acquired for all animals with (n=8) and without (n=3) cell grafting at approximately 24 h, 72 h, and weekly for 6 weeks after stroke. Transplanted cells were tracked by high-resolution three-dimensional gradient-echo images and the interaction between the cells and ischemic lesion was detected by ISODATA (Iterative Self-Organizing Data Analysis Technique Algorithm) calculated from T(1), T(2) and T(1sat) maps. Tissue status from ISODATA was characterized by a specific signature, which represents the deviation from normal tissue in the feature space. Transplanted SVZ cells selectively migrated towards the ischemic side of the rat brain and approached the lesion boundary within 1-week after grafting. Cell treated rats exhibited a significant reduction of average lesion size compared with control rats (P<0.05). A significant reduction of tissue signature (P<0.001) induced by cell transplantation was localized to the position of grafted cells, and these sites exhibited stably restored cerebral blood flow (CBF) (approximately 85% of normal CBF). Angiogenesis was present in sites either immediately adjacent to or surrounded by the grafted cells. Our data indicate that map-ISODATA accurately and dynamically characterizes the ischemic lesion and its response to cell therapy.

Effect of early decrease in the lesion size on late brain tissue loss, synaptophysin expression and functionality after a focal brain lesion in rats

The purpose of the present study is to determine the effects of early decrease in the lesion size on late brain tissue loss, synaptogenesis and functionality after a focal brain lesion in rats. The lesion was induced either to the cortex using the photothrombotic ischemic stroke or to the striatum using the malonate poisoning model. The cortical and striatal lesions amounted to 66–80 mm 3 at day 1 post-lesion and were reduced by 50% after the acute administration of dipyridyl (a liposoluble iron chelator) and aminoguanidine (an inhibitor of the inducible nitric oxide synthase), respectively. Loss of histologically intact tissue and synaptophysin expression as an indicator of synaptogenesis were examined at day 35 post-lesion. Both types of lesion resulted in synaptophysin upregulation in contralateral and ipsilateral cortical areas. On the contrary, brain tissue loss was greater after the striatal (−17%) than the cortical lesion (−5%). Synaptophysin expression and tissue loss were not different between drug- and vehicle-treated rats. Moreover, a set of standard neurological tests revealed a difference in deficit between the both types of lesion, yet only in the acute post-lesion stage. However, it did not distinguish between vehicle- and drug-treated rats whatever the lesion location. Our results indicate that late histological endpoints measurements are not recommended to probe the potential neuroprotective properties of a drug administered within the acute post-lesion stage. They also suggest that inhibition of cytotoxic mechanisms involved in lesion growth is of no clinical interest when it cannot lead to a long-term histological protection and/or increased synaptogenesis.

Quantification of acetogenins in Annona muricata linked to atypical parkinsonism in guadeloupe

Atypical parkinsonism in Guadeloupe has been associated with the consumption of fruit and infusions or decoctions prepared from leaves of Annona muricata L. (Annonaceae), which contains annonaceous acetogenins, lipophilic inhibitors of complex I of the mitochondrial respiratory chain. We have determined the concentrations of annonacin, the major acetogenin in A. muricata, in extracts of fruit and leaves by matrix-assisted laser desorption-ionization mass spectrometry. An average fruit is estimated to contain about 15 mg of annonacin, a can of commercial nectar 36 mg, and a cup of infusion or decoction 140 microg. As an indication of its potential toxicity, an adult who consumes one fruit or can of nectar a day is estimated to ingest over 1 year the amount of annonacin that induced brain lesions in rats receiving purified annonacin by intravenous infusion.

TPA as an ‘interface’ between blood and brain

Over the last decade, tPA has been implicated in a variety of brain functions. It is secreted from growth cones or extending neurites, it modulates neurite outgrowth and promotes neuronal migration. tPA has also been involved in physiological glutamatergic-dependent processes such as synaptic plasticity and long term potentiation. In addition, t-PA has been shown to play an important role in the pathogenesis of seizure, multiple sclerosis, trauma and ischemic brain injury. In a previous study, we have shown that the proteolytic activity of t-PA enhances the N-methyl-D-aspartate (NMDA) receptor-mediated signalling (Nicole et al., 2001). Here, we provide evidences that tPA is able to cross the intact blood brain barrier both in vitro and in vivo and that this mechanism is independent of its proteolytic activity and mediated by an interaction with a LDL-related receptor, LRP. Overall, these data suggest that blood-derived tPA could influence neuronal functioning. Accordingly, intravenous injection of tPA potentiates the NMDA-induced brain lesion in rats even in the absence of blood brain barrier leakage. As a model demonstrating that preventing the NMDA-dependent effect of tPA could improve the efficiency of thrombolysis in the treatment of stroke, we evidence that the blood-derived tPA failed to enhance NMDA-dependent signalling in a model of mice previously vaccinated with the amino-terminal domain (ATD) of NR1 subunit of the NMDA receptor. Thus, we propose a new concept suggesting that tPA is one of the key element of the interface between the blood and the brain parenchyma.

MRI Reveals That Early Changes in Cerebral Blood Volume Precede Blood–Brain Barrier Breakdown and Overt Pathology in MS-like Lesions in Rat Brain

Magnetic resonance imaging (MRI) is an established clinical tool for diagnosing multiple sclerosis (MS), the archetypal central nervous system neuroinflammatory disease. In this study, we have used a model of delayed-type hypersensitivity in the rat brain, which bears many of the hallmarks of an MS lesion, to investigate the development of MRI-detectable changes before the appearance of conventional indices of lesion development. In addition, we have correlated the MRI-detectable changes with the developing histopathology. Significant increases in regional cerebral blood volume (rCBV) preceded overt changes in blood-brain barrier (BBB) permeability, T2 relaxation and the diffusion properties of tissue water. Thus, changes in rCBV might be a more sensitive indicator of lesion onset than the conventional indices used clinically in MS patients, such as contrast enhancement. In addition, we show that BBB breakdown, and consequent edema formation, are more closely correlated with astrogliosis than any other histopathologic changes, while regions of T1 and T2 hypointensity appear to reflect hypercellularity.

Determination of relaxation characteristics during preacute stage of lysophosphatidyl choline-induced demyelinating lesion in rat brain: an animal model of multiple sclerosis

Relaxation time measurements were carried out during the preacute stage of lesion progression in an animal model of demyelination created in the internal capsule (ic) area of the rat brain using lysophosphatidyl choline (LPC). T1 and T2 were determined both before and after 36 h of lesion creation. Histology carried out on the rats after MR measurements showed focal demyelinating lesion and surrounding edema with prominent infiltration of inflammatory cells. Both T1 and T2 were statistically higher for the lesion compared to that determined before lesion creation. Percentage increase in T2 was found to be higher by approximately 45% compared to before lesion creation while T1 showed about 25% increase. Increase in T1 and T2 may be attributed to the early acute inflammatory response due to LPC. The beginning of the inflammatory response following LPC injection may also be a contributing factor. The study demonstrates that the quantitative estimate of MR relaxation provides useful information on the pathological events occurring during the early phase of the progression of demyelination.

GABA Neurons Survive Focal Ischemic Injury

Focal cerebral lesions in rat brain induced by photothrombosis lead to an impaired inhibitory neurotransmission. A reduced gamma-aminobutyric acid (GABA)-mediated inhibition has been revealed by electrophysiological recordings associated with a diminished immunostaining of GABA handling proteins. Changes were found in ipsi- as well as in contralateral brain areas. Inhibition is mediated by interneurons using GABA as neurotransmitter. These cells use GAD (glutamate decarboxylase) to synthesize GABA. To analyze the vulnerability of GABAergic neurons in rats with a lesioned hindlimb area, cells expressing GAD65/67 mRNA were labeled using in situ hybridization. Positive somata were counted 7 and 30 days after focal ischemia in different cortical (hindlimb cortex, frontal cortex, primary and secondary somatosensory cortex) and hippocampal subsectors (pyramidal cell layer, stratum oriens and stratum radiatum/lacunosum-moleculare). The lesioned hemispheres were compared with the intact brain sides and with control brains. GABAergic interneurons survived the injury for up to 30 days in all investigated brain regions. Therefore it is unlikely that a loss of GABAergic neurons contributes to the reduced inhibition.

Astrocytic Connexin 43 mRNA is Upregulated in the Vicinity of the Photothrombotic Lesion in Rat Brain

Intercellular coupling via gap junction channels is vital for astrocytic function. The role of gap junctional communication under ischemic conditions is discussed controversely. In our model of focal cerebral ischemia, we studied the expression of connexin 43, one of the major gap junction proteins in astrocytes. Connexin 43 mRNA is elevated in cells surrounding the lesion site which was shown by RNA/RNA in situ hybridization. The hybridization signal in pericarya around the lesion was much higher than in the remaining brain areas. This upregultion was further investigated by semiquantitative RT-PCR. Biopsies were taken from the lesion site as well as from the corresponding area in the intact hemisphere and from sham-operated animals. In each analyzed animal (n=6) with a focal cerebral ischemia the amount of specific PCR product was higher in the sample from the operated site than in the intact. Sham-operated animals (n=5) showed no variation between sides. Real-time RT-PCR analysis confirmed the results obtained in semiquantitative RT-PCR studies. Our results affirm that gap junctions composed of connexin 43 play a role after ischemia. Whether this upregulation of connexin 43 mRNA after an ischemic insult contributes to neuroprotection or propagation of the cell needs to be determined.