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Background:The onset of progressive cerebellar ataxia in early childhood is considered a key feature of ataxiatelangiectasia (A-T), accompanied by ocular apraxia, telangiectasias, immunodeficiency, cancer susceptibility and hypersensitivity to ionizing radiation.Methods:We describe the clinical features and course of three Mennonite children who were diagnosed with A-T following the completion of therapy for lymphoid malignancies.Results:Prior to cancer therapy, all had non-progressive atypical neurological abnormalities, with onset by age 30 months, including dysarthria, dyskinesia, hypotonia and/or dystonia, without telangiectasias. Cerebellar ataxia was noted in only one of the children and was mild until his death at age eight years. None had severe infections. All three children were “cured” of their lymphoid malignancies, but experienced severe adverse effects from the treatments administered. The two children who received cranial irradiation developed supratentorial primitive neuroectodermal tumors of the brain, an association not previously described, with fatal outcomes.Conclusions:The range of neurological presentations of A-T is broad. Ataxia and telangiectasias may be minimal or absent and the course seemingly non-progressive. The diagnosis of A-T should be considered in all children with neuromotor dysfunction or peripheral neuropathy, particularly those who develop lymphoid malignancies. The consequences of missing the diagnosis may be dire. Radiation therapy and radiomimetic drugs should be avoided in individuals with A-T.

The germinal matrix of human brain gives rise to oligodendrocytes and astrocytes after mid-gestation. Hemorrhage in the germinal matrix of premature infants is associated with suppressed cell proliferation. We hypothesize that soluble blood constituents have an adverse effect on the proliferation of cultured rat subventricular zone (SVZ) cells and the proliferation, migration, and differentiation of oligodendrocyte progenitor cells (OPC). Using caspase 3 activation and lactate dehydrogenase release assays, rat plasma, serum, thrombin, and kallikrein killed SVZ cells when grown in the presence (but not absence) of platelet derived growth factor. Plasma and serum killed OPC at 1 : 1 to 1 : 100 dilutions. Using a bromodeoxyuridine incorporation assay OPC proliferation was reduced by plasma, serum, thrombin and plasmin. Blood proteins also suppressed OPC migration in a concentration dependent manner. However, differentiation of OPC into myelin basic protein expressing cells was suppressed only by thrombin. We conclude that soluble blood components, particularly thrombin, have an adverse effect on maturing SVZ cells and OPC derived from newborn rat brain.

Cryptic intracerebral hemorrhage as an etiological factor in fetal hydrocephalus has been postulated but not described at autopsy. Four fetuses with overt hydrocephalus diagnosed by in utero ultrasound examination were examined at autopsy at 19-22 weeks gestation. Although a hemorrhagic etiology was not evident on ultrasound, hemosiderin-containing macrophages and associated reactive changes were found to obstruct the otherwise well-formed cerebral aqueduct in all four. Coagulopathy due to thrombocytopenia was implicated in one case. Anomalies involving other parts of the body were identified in two cases, although a direct link to the hydrocephalus was not obvious. The abnormality was isolated in one case. In three cases, possible sites of hemorrhage in the ventricles were identified. This abnormality represents a significant proportion of the fetuses examined for hydrocephalus in our referral center. We discuss the importance of careful autopsy examination in the diagnosis of cryptic intracerebral hemorrhage and the implications for counseling.

This study was undertaken to determine if dialysis of damaged brain surface can reduce cerebrospinal fluid (CSF) pressure and progressive brain edema. The authors secondarily determined if local brain cooling was simultaneously possible.Telemetric pressure transmitters were implanted into the lumbar subarachnoid space of 58 young adult male rats. Cryogenic brain injury was created and 2 hours later decompressive craniectomy was performed. An osmotic cell with a semipermeable dialysis membrane placed on the damaged brain surface was perfused with dextran 15% solution for 2 or 4 hours. Water content was determined in the cerebral hemispheres using the wet-dry weight method. Evans blue-albumin spread was measured morphometrically. Brain temperature was measured bilaterally.The CSF pressure increased after cryogenic injury and decreased after craniotomy. Two hours of brain dialysis significantly reduced CSF pressure in comparison with craniotomy alone and sham dialysis. Injured brain had higher water content, but this was not affected by dialysis. Spread of Evans blue-albumin, however, was significantly reduced by the treatment. Cooling of the dialysis solution caused significant local brain cooling.Surface dialysis of cryogenically injured rat brain controls CSF pressure and reduces intraparenchymal spread of edema fluid in the acute period after injury. The authors postulate that edema fluid moves into the osmotic cell rather than spreading through the uninjured brain. Long-term experiments will be needed to prove that this combination therapy is effective.

Eosinophilic intracytoplasmic inclusions have been rarely described in Purkinje neurons of children with a variety of neurological conditions. Here we document these inclusions in five children from 3 to 14 years of age. One child had 7q deletion syndrome and a second had profound motor and cognitive delay ("cerebral palsy") of unknown origin, while three others were neurologically normal prior to death. These inclusions stain with the PAS method, are not strongly ubiquitinated, and are located in the lumen of endoplasmic reticulum. Their appearance in a wide range of disorders and in neurologically normal children suggests that they are a nonspecific protein trafficking anomaly, possibly aggravated under degenerative conditions.

Background The cerebral cortex may be compressed in hydrocephalus and some experiments suggest that movement of extracellular substances through the cortex is impaired. We hypothesized that the extracellular compartment is reduced in size and that the composition of the extracellular compartment changes in rat brains with kaolin-induced hydrocephalus. Methods We studied neonatal (newborn) onset hydrocephalus for 1 or 3 weeks, juvenile (3 weeks) onset hydrocephalus for 3–4 weeks or 9 months, and young adult (10 weeks) onset hydrocephalus for 2 weeks, after kaolin injection. Freeze substitution electron microscopy was used to measure the size of the extracellular compartment. Western blotting and immunohistochemistry with quantitative image densitometry was used to study the extracellular matrix constituents, phosphacan, neurocan, NG2, decorin, biglycan, and laminin. Results The extracellular space in cortical layer 1 was reduced significantly from 16.5 to 9.6% in adult rats with 2 weeks duration hydrocephalus. Western blot and immunohistochemistry showed that neurocan increased only in the periventricular white matter following neonatal induction and 3 weeks duration hydrocephalus. The same rats showed mild decorin increases in white matter and around cortical neurons. Juvenile and adult onset hydrocephalus was associated with no significant changes. Conclusion We conclude that compositional changes in the extracellular compartment are negligible in cerebral cortex of hydrocephalic rats at various ages. Therefore, the functional change related to extracellular fluid flow should be reversible.

Background The objective of this study was to develop a simple and inexpensive animal model of induced obstructive hydrocephalus with minimal tissue inflammation, as an alternative to kaolin injection. Materials Two-hundred and two male Sprague-Dawley rats aged 3 weeks received intracisternal injections of kaolin (25% suspension), Matrigel, type 1 collagen from rat tail, fibrin glue (Tisseel), n-butyl-cyanoacrylate (NBCA), or ethylene vinyl alcohol copolymer (Onyx-18 and Onyx-34). Magnetic resonance imaging was used to assess ventricle size. Animals were euthanized at 2, 5, 10 and 14 days post-injection for histological analysis. Results Kaolin was associated with 10% mortality and successful induction of hydrocephalus in 97% of survivors (ventricle area proportion 0.168 ± 0.018). Rapidly hardening agents (fibrin glue, NBCA, vinyl polymer) had high mortality rates and low success rates in survivors. Only Matrigel had relatively low mortality (17%) and moderate success rate (20%). An inflammatory response with macrophages and some lymphocytes was associated with kaolin. There was negligible inflammation associated with Matrigel. A severe inflammatory response with giant cell formation was associated with ethylene vinyl alcohol copolymer. Conclusion Kaolin predictably produces moderate to severe hydrocephalus with a mild chronic inflammatory reaction and fibrosis of the leptomeninges. Other synthetic polymers and biopolymers tested are unreliable and cause different types of inflammation.

Inflammation, demyelination, and blood-spinal cord barrier (BSB) breakdown occur in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. The purpose of this study was to evaluate the utility of MRI for detecting lesions and BSB disruption in vivo during EAE in the mouse lumbar spinal cord, to determine how MR features of BSB disruption change during the course of disease, and to relate such changes to clinical signs and histological features of disease. Following induction of EAE in C57BL/6 mice, contrast-enhanced (CE) T1-weighted MR images were acquired to detect BSB disruption in the lumbar spinal cord at the early stage of disease, at peak disease, and at remission, and T2weighted images were obtained to monitor spinal cord morphology. Following imaging the spinal cords were assessed in situ for general features of inflammation, BSB leakage, activated macrophages/microglia, and demyelination. No focal lesions were evident on T2-weighted MR images. BSB disruption was greatest at the onset of signs of disease, and decreased progressively thereafter. Inflammation and demyelination were pronounced at the initial stage of disease and at peak disease, and were decreased at remission. Nonuniform contrast enhancement indicated that breakdown of the BSB occurred predominantly within the white matter (WM) of the spinal cord. Magn Reson Med 58:298 –305, 2007. © 2007 Wiley-Liss, Inc.

Background Previous work with 3-week hydrocephalic rats showed that white matter damage could be reduced by the calcium channel antagonist magnesium sulfate (MgSO4). We hypothesized that MgSO4 therapy would improve outcomes in ferrets with hydrocephalus induced with kaolin at 15 days. Methods MRI was performed at 29 days to assess ventricle size and stratify ferrets to treatment conditions. Beginning at 31 days age, they were treated daily for 14 days with MgSO4 (9 mM/kg/day) or sham saline therapy, and then imaged again before sacrifice. Behavior was examined thrice weekly. Histological and biochemical ELISA and myelin enzyme activity assays were performed at 46 days age. Results Hydrocephalic ferrets exhibited some differences in weight and behavior between treatment groups. Those receiving MgSO4 weighed less, were more lethargic, and displayed reduced activity compared to those receiving saline injections. Hydrocephalic ferrets developed ventriculomegaly, which was not modified by MgSO4 treatment. Histological examination showed destruction of periventricular white matter. Glial fibrillary acidic protein content, myelin basic protein content, and myelin enzyme activity did not differ significantly between treatment groups. Conclusion The hydrocephalus-associated disturbances in juvenile ferret brains are not ameliorated by MgSO4 treatment, and lethargy is a significant side effect.

Medulloblastoma (MB) is the most common malignant primary pediatric brain tumor and is currently divided into 4 subtypes based on different genomic alterations, gene expression profiles and response to treatment: Wnt, Sonic Hedgehog (Shh), Group 3 and Group 4. This extensive heterogeneity has made it difficult to assess the functional relevance of genes to malignant progression. For example, expression of the transcription factor, Orthodenticle homeobox2 (Otx2) is frequently dysregulated in multiple MB variants; however, its role may be subtype-specific. We recently demonstrated that neural precursors derived from transformed human embryonic stem cells (trans-hENs), but not their normal counterparts (hENs), resemble Groups 3 and 4 MB in vitro and in vivo. These trans-hENs have >12-fold overexpression of Otx2. Here, we tested the utility of this model system as a means of dissecting the role of Otx2 in MB using gain and loss of function studies in hENs and trans-hENs respectively. Parallel experiments with MB cells revealed that Otx2 exerts tumor suppressive effects in hEN and Shh MB cells, likely by regulating self-renewal and decreasing expression of pluripotent genes such as Sox2, Lin28A, Oct4 and Nanog. Concomitant to the negative regulation of hESC genes, we observed a decrease in levels of the active histone mark, H3K4me3 at promoter loci of pluripotent genes as well as an increase in Otx2 recruitment at the Sox2 promoter by chromatin immunoprecipitation assays. This was supported by overexpression of Sox2 in Otx2+ Shh MB and hENs that resulted in significant rescue of self-renewal and cell migration. In contrast, Otx2 is oncogenic and promotes self-renewal of trans-hENs and Groups 3/4 MB independent of pluripotent genes. These results demonstrate a cell context-dependent function of Otx2. Our study underscores the value of hESC derivatives as developmental models of MB and for investigating key biological questions related to embryonal childhood cancers.

Germinal matrix hemorrhage refers to bleeding that arises from the subependymal (or periventricular) germinal region of the immature brain. Clinical studies have shown that infants who experience germinal matrix hemorrhage can develop hydrocephalus or suffer from long-term neurologic dysfunction, including cerebral palsy, seizures, and learning disabilities. Understanding the causative factors and the pathogenesis of subsequent brain damage is important if germinal matrix hemorrhage is to be prevented or treated. Appropriate animal models are necessary to achieve this understanding. A number of animal species, including mice, rats, rabbits, sheep, pigs, dogs, cats, and primates, have been used to model germinal matrix hemorrhage. This literature review critically evaluates the animal models of germinal matrix hemorrhage. Each model has its own advantages and disadvantages; no single model is suitable for the study of all aspects of brain damage.

The mechanism of brain cell injury associated with intracerebral hemorrhage may be in part related to proteolytic enzymes in blood, some of which are also functional in the developing brain. We hypothesized that there would be an age-dependent brain response following intracerebral injection of blood, thrombin, and plasminogen. Mice at 3 ages (neonatal, 10-day-old, and young adult) received autologous blood (15, 25, and 50 microl respectively), thrombin (3, 5, and 10 units respectively), plasminogen (0.03, 0.05, and 0.1 units respectively) (the doses expected in same volume blood), or saline injection into lateral striatum. Forty-eight hours later they were perfusion fixed. Hematoxylin and eosin, lectin histochemistry, Fluoro-Jade, and TUNEL staining were used to quantify changes related to the hemorrhagic lesion. Damage volume, dying neurons, neutrophils, and microglial reaction were significantly greater following injections of blood, plasminogen, and thrombin compared to saline in all three ages of mice. Plasminogen and thrombin associated brain damage was greatest in neonatal mice and, in that group unlike the other 2, greater than the damage caused by whole blood. These results suggest that the neonatal brain is relatively more sensitive to proteolytic plasma enzymes than the mature brain.

Neonatal periventricular hemorrhage (PVH) is a devastating complication of prematurity in the human infant. Based upon observations made primarily in adult rodents and the fact that the immature brain uses proteolytic systems for cell migration and growth, we hypothesized that thrombin and plasmin enzyme activities contribute to the brain damage after PVH. The viability of mixed brain cells derived from newborn rat periventricular region was suppressed by whole blood and thrombin, but not plasmin. Following injection of autologous blood into the periventricular region of newborn rat brain, proteolytic activity was detected in a halo around the hematoma using membrane overlays impregnated with thrombin and plasmin fluorogenic substrates. Two-day old rats received periventricular injection of blood, thrombin, and plasminogen. After 2 days, thrombin and blood were associated with significantly greater damage than saline or plasminogen. Two-day old mice received intracerebral injections of blood in combination with saline or the proteolytic inhibitors hirudin, alpha2macroglobulin, or plasminogen activator inhibitor-1. After 2 days, hirudin significantly reduced brain cell death and inflammation. Two-day-old mice then received low and high doses of hirudin mixed with blood after which behavioral testing was conducted repeatedly. At 10 weeks there was no statistically significant evidence for behavioral or structural brain protection. These results indicate that thrombin likely plays a role in neonatal periventricular brain damage following PVH. However, additional factors are likely important in the recovery from this result.

Premature infants with placental infection and adult stroke patients with fever have worse outcomes following intracerebral hemorrhage (ICH). We hypothesized that immune pre-activation would aggravate brain injury in mouse brain following ICH. The immune system of 2-day, 10-day and 7-week young adult CD1 mice was stimulated by intraperitoneal injection of concanavalin A (ConA), lipopolysaccharide (LPS) or polyinosinic-polycytidilic acid (PolyI:C) 12 h prior to intracerebral injection of blood. Two days later, brain damage and inflammation were worse in 2-day mice that had received LPS. The other agents had less consistent effects in 2-day mice. Brain damage in young adults was aggravated less after immune stimulation. These data suggest that immune pre-activation modifies hemorrhagic brain injury in immature mouse brain.

Glutaric acidemia type 1 (GA-1) is an autosomal recessive disorder characterized by a deficiency of glutaryl-CoA dehydrogenase (GCDH) activity. GA-1 is often associated with an acute encephalopathy between 6 and 18 months of age that causes striatal damage resulting in a severe dystonic movement disorder. Ten autopsy cases have been previously described. Our goal is to understand the disorder better so that treatments can be designed. Therefore, we present the neuropathological features of six additional cases (8 months-40 years), all North American aboriginals with the identical homozygous mutation. This cohort displays similar pathological characteristics to those previously described. Four had macroencephaly. All had striatal atrophy with severe loss of medium-sized neurons. We present several novel findings. This natural time course study allows us to conclude that neuron loss occurs shortly after the encephalopathical crisis and does not progress. In addition, we demonstrate mild loss of large striatal neurons, spongiform changes restricted to brainstem white matter and a mild lymphocytic infiltrate in the early stages. Reverse transcriptase-PCR to detect the GCDH mRNA revealed normal and truncated transcripts similar to those in fibroblasts. All brain regions demonstrated markedly elevated concentrations of GA (3770-21 200 nmol/g protein) and 3-OH-GA (280-740 nmol/g protein), with no evidence of striatal specificity or age dependency. The role of organic acids as toxic agents and as osmolytes is discussed. The pathogenesis of selective neuronal loss cannot be explained on the basis of regional genetic and/or metabolic differences. A suitable animal model for GA-1 is needed.

The literature concerning brain damage due to hydrocephalus, especially in children and animal models, is reviewed. The following conclusions are reached: 1.. Hydrocephalus has a deleterious effect on brain that is dependent on magnitude and duration of ventriculomegaly and modified by the age of onset. 2.. Animal models have many histopathological similarities to humans and can be used to understand the pathogenesis of brain damage. 3.. Periventricular axons and myelin are the primary targets of injury. The pathogenesis has similarities to traumatic and ischemic white matter injury. Secondary changes in neurons reflect compensation to the stress or ultimately the disconnection. 4.. Altered efflux of extracellular fluid could result in accumulation of waste products that might interfere with neuron function. Further research is needed in this as well as the blood‐brain barrier in hydrocephalus. 5.. Some, but not all, of the changes are preventable by shunting CSF. However, axon loss cannot be reversed, therefore shunting in a given case must be considered carefully. 6.. Experimental work has so far failed to show any benefit in reducing CSF production. Pharmacologic protection of the brain, at least as a temporary measure, holds some promise but more pre‐clinical research is required.

Cerebellar liponeurocytoma, a rare, newly identified CNS neoplasm of adults, is characterized by advanced neuronal/neurocytic and focal lipomatous differentiation, low proliferative potential and a favorable clinical prognosis. Despite the different age distribution and benign biological behavior, the cerebellar liponeurocytoma shares several features with the cerebellar medulloblastoma, which may include an origin from the periventricular matrix of the fourth ventricle or the external granular layer of the cerebellum. To establish the genetic profile of cerebellar liponeurocytomas, we have formed an international consortium and collected tumor samples from 20 patients. DNA sequencing revealed TP53 missense mutations in 4 (20%) of 20 cerebellar liponeurocytomas, a frequency higher than in medulloblastomas. There was no case with PTCH, APC, or beta-catenin mutations, each of which may be present in subsets of medulloblastomas. Isochromosome 17q, a genetic hallmark of classic medulloblastomas, was not observed in any of the cases investigated by FISH analysis. cDNA array analyses were carried out on 4 cerebellar liponeurocytomas, 4 central neurocytomas, and 4 classic medulloblastomas. Cluster analysis of the cDNA expression data of 1176 genes grouped cerebellar liponeurocytomas close to central neurocytomas, but distinct from medulloblastomas. These results suggest cerebellar liponeurocytoma as a distinct tumor entity that is genetically different from medulloblastoma. Furthermore, the cDNA expression array data suggest a relationship to central neurocytomas, but the presence of TP53 mutations, which are absent in central neurocytomas, suggests that their genetic pathways are different.

Hydrocephalus causes damage to periventricular axons. Tacrolimus, cyclosporine A (CsA) and calpain inhibitors have been shown to protect axons in rat models of acute traumatic brain injury. We hypothesized that these agents would ameliorate the axon damage and behavioral effects in experimental hydrocephalus. Hydrocephalus was induced in 3-week-old rats by injection of kaolin into the cisterna magna. Tests of cognitive and motor function were performed on a weekly basis. In a blinded and randomized manner, tacrolimus (FK506; 3.6 mg/kg body weight) or CsA (10 mg/kg) was administered once daily by subcutaneous injection for 2 weeks, beginning 2 weeks after induction of hydrocephalus. In a separate experiment, calpain inhibitor I (10 mg/kg/day) was administered by continuous subcutaneous infusion. The brains were subjected to histopathological and biochemical analyses after 2 weeks of treatment. There was no statistically significant protection in regard to behavior, brain structure or brain composition in any of the experiments. However, there was biochemical and histological evidence of renal injury following chronic tacrolimus and CsA administration. Calcineurin inhibition does not offer significant protection in this rat model of hydrocephalus.

Chronic hydrocephalus that begins in childhood and progresses only very gradually is sometimes called "arrested" hydrocephalus. Data suggest that this state eventually can become symptomatic and may be treatable by shunting. However, the pathological substrate of the disorder is not entirely understood. We studied chronic hydrocephalus in rats, 9 months after induction by kaolin injection into the cisterna magna, and in humans. In both circumstances, destruction of periventricular white matter structures was worst in those with the largest ventricles. Structures damaged include the corpus callosum, corticospinal tract, and fimbria/fornix projections from the hippocampus. Myelin turnover was increased. These changes were associated with deficits of motor and cognitive function. The cerebral cortex was largely spared. There appears to be a threshold of ventricle size beyond which functional changes manifest, but this undoubtedly is modified by factors such as age of onset and rate of enlargement. These data support the need for persistent follow-up of patients with chronic, apparently stable hydrocephalus. A slight increase in size of already enlarged ventricles might cause significant axonal damage.

Major research efforts have focused on the isolation and characterization of brain tumor stem cells, or propagating cells (BTPC) in a variety of brain cancers. Elucidating cell surface marker profiles that can be used to selectively isolate this cellular population is an imperative first step in the development of targeted therapies. Medulloblastoma (MB) is the most common form of pediatric brain cancer. MB is divided into 4 molecular subgroups; Wnt, Sonic Hedgehog (SHH), Group 3 and Group 4. Given the variable results obtained for currently utilized markers, as well as the cellular heterogeneity within and between MB sub-groups, it is likely there are additional surface marker profiles capable of selecting for sub-type specific MB BTPCs. We set out to identify novel surface marker combinations capable of selecting for TPCs in SHH MB. We employed the new BD Bioscience Lyoplate screening platform to compare 242 human cell surface marker levels across high and low self-renewing SHH MB sub-clones. The top 25 markers were refined by evaluating expression levels in Shh vs Group 3,4 and Wnt variants in transcriptome datasets representing 548 patient samples. Four markers, CD271, CD106/VCAM1, EGFR and CD171/NCAM-L1 showed consistent differential expression in the SHH subtype relative to the other variants. Flow cytometry validation in additional cell lines confirmed these findings. As a proof of principle, functional characterization of CD271 in SHH MB in vitro and in vivo was performed. Using fluorescence activated cell sorting and gain/loss of function studies, our results suggest that CD271 selects for MB progenitor cells. This work highlights a new approach to screening for differentially expressed surface markers across matched samples. We delineated a cell surface fingerprint for BTPC populations from MB molecular variants, however the utility can be seen in normal stem cell biology and across all forms of cancer.

The Acp2 gene encodes the beta subunit of lysosomal acid phosphatase, which is an isoenzyme that hydrolyzes orthophosphoric monoesters. In mice, a spontaneous mutation in Acp2 results in severe cerebellar defects. These include a reduced size, abnormal lobulation, and an apparent anterior cerebellar disorder with an absent or hypoplastic vermis. Based on differential gene expression in the cerebellum, the mouse cerebellar cortex can normally be compartmentalized anteroposteriorly into four transverse zones and mediolaterally into parasagittal stripes. In this study, immunohistochemistry was performed using various Purkinje cell compartmentation markers to examine their expression patterns in the Acp2 mutant. Despite the abnormal lobulation and anterior cerebellar defects, zebrin II and PLCβ4 showed similar expression patterns in the nax mutant and wild type cerebellum. However, fewer stripes were found in the anterior zone of the nax mutant, which could be due to a lack of Purkinje cells or altered expression of the stripe markers. HSP25 expression was uniform in the central zone of the nax mutant cerebellum at around postnatal day (P) 18-19, suggesting that HSP25 immunonegative Purkinje cells are absent or delayed in stripe pattern expression compared to the wild type. HSP25 expression became heterogeneous around P22-23, with twice the number of parasagittal stripes in the nax mutant compared to the wild type. Aside from reduced size and cortical disorganization, both the posterior zone and nodular zone in the nax mutant appeared less abnormal than the rest of the cerebellum. From these results, it is evident that the anterior zone of the nax mutant cerebellum is the most severely affected, and this extends beyond the primary fissure into the rostral central zone/vermis. This suggests that ACP2 has critical roles in the development of the anterior cerebellum and it may regulate anterior and central zone compartmentation. peerReviewed

Neuronal and glial cell death in the striatum of a rat model of collagenase-induced intracerebral hemorrhage begins at 1 day and continues for at least 3 weeks. We hypothesized that administration of a neurotrophic agent would reduce neuronal loss in this experimental model. Because it has been shown to protect striatal neurons against excitotoxic injury, a second-generation ciliary neurotrophic factor (CNTF) (AXOKINE) was administered by continuous intracerebral infusion (2 microg/day) beginning 28 h after hemorrhage and continuing for 2 weeks. Magnetic resonance imaging showed that the hematoma size was comparable in control and treated rats prior to treatment. Counts of medium-sized striatal neurons within 320 microm of the hematoma 8 weeks after the hemorrhage revealed a slight but statistically significant benefit with a 42.5% loss in treated rats compared to 51.7% loss in controls. The results suggest that AXOKINE might be protective of striatal neurons in the vicinity of a hemorrhagic lesion.

Background and Purpose — Extravasation of blood is associated with intracerebral hemorrhage and head trauma. The mechanism of brain cell injury associated with hemorrhage differs from that due to pure ischemia. The purpose of this study was to investigate the acute changes after intracerebral injections of proteins that are involved in blood clotting and clot lysis. Methods — Sixty-eight adult rats were subjected to stereotaxic intrastriatal injections of normal saline (5 μL), low- (2.5 U/5 μL) and high-dose (25 U/5 μL) thrombin, low- (0.1 μg/5 μL) and high-dose (1 μg/5 μL) tissue plasminogen activator, low- (0.05 U/5 μL) and high-dose (0.5 U/5 μL) plasminogen, and low- (0.335 U/5 μL) and high-dose (3.35 U/5 μL) plasmin. Forty-eight hours later rats were perfusion fixed. Brain damage area, eosinophilic neurons, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL)-positive cells, infiltrating neutrophils, CD8a immunoreactive leukocytes, and reactive microglia were quantified. Results — Damage area in striatum, dying cells, inflammatory cells, and microglial reaction were significantly greater after the high-dose plasminogen, plasmin, and thrombin injections. Tissue plasminogen activator injections were associated with mild inflammation. Conclusions — These results suggested that thrombin and plasmin are harmful to brain cells in vivo. Although the doses required to cause damage are relatively great in consideration of the plasma content of these proteins, their pathological effect might be enhanced through synergism with other mechanisms.

The ubiquitous neuromodulator adenosine inhibits the production of several proinflammatory cytokines through activation of specific cell-surface adenosine receptors. We demonstrated recently that antisense oligonucleotides to tumor necrosis factor-alpha (TNF-alpha) are neuroprotective in a rat model of intracerebral hemorrhage. Therefore, we hypothesized that activation of adenosine receptors would provide protection against intracerebral hemorrhage-induced TNF-alpha production and inflammatory events. In vitro experiments showed that adenosine A1, A2A, and A3 receptor subtypes were present on U937 cells, and activation of these subtypes inhibited TNF-alpha production with a rank order of A2A > > A1 > A3. Prolonged treatment of U937 cells with the A2A receptor agonist 2-p-(carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS 21680) desensitized adenosine A2A, A1, and A3 receptors. CGS 21680 administration directly into the striatum immediately prior to the induction of intracerebral hemorrhage inhibited TNF-alpha mRNA and, 24 hours following induction, reduced parenchymal neutrophil infiltration (p < 0.001) and TUNEL-positive cells (p < 0.002) within and bordering the hematoma. These results suggest that pharmacological strategies targeting A2A receptors may provide effective inhibition of acute neurotoxic proinflammatory events that occur following intracerebral hemorrhage.

Object. Hydrocephalus, a pathological dilation of the ventricles of the brain, causes damage to periventricular white matter, at least in part, through chronic ischemia. The authors tested the hypothesis that treatment with nimodipine, an L-type calcium channel-blocking agent with demonstrated efficacy in a range of cerebral ischemic disorders, would ameliorate the adverse effects of experimental hydrocephalus. Methods. Hydrocephalus was induced in 3-week-old rats by injection of kaolin into the cisterna magna. The rats were treated by continuous administration of nimodipine or control vehicle for 2 weeks, beginning 2 weeks after induction of hydrocephalus. During the treatment period, the animals underwent repeated tests of motor and cognitive behavior. At the end of the treatment period, the rat brains were analyzed by histopathological and biochemical means. Nimodipine treatment prevented the declines in motor and cognitive behavior that were observed in untreated control rats. During the treatment period, ventricular enlargement, determined by magnetic resonance imaging, was equal in the two groups, although the corpus callosum was thicker in the treated rats. Myelin content in white matter and synaptophysin content in gray matter, an indicator of synapses, did not differ. Conclusions. The protective effect of nimodipine is most likely based on improved blood flow, although prevention of calcium influx—mediated proteolytic processes in axons cannot be excluded. Adjunctive pharmacological therapy may be beneficial to patients with hydrocephalus.

Beginning 15 min after induction of intracerebral hemorrhage (ICH) by intrastriatal administration of collagenase, rats were treated intramuscularly with FK-506 (3 mg/kg) or with vehicle. Treatment was repeated daily for 7 days. MR imaging 1, 7, and 28 days post-ICH showed that treatment did not affect hematoma size or its subsequent resolution. Two days post-ICH, neutrophil infiltration around the hematoma was decreased in the FK-506-treated rats, as was the number of TUNEL-positive cells at the edge of the hematoma and in the peripheral region. The decreased inflammatory response was accompanied by functional improvement in the treated rats. The neurological deficit induced by the ICH (beam walking ability, postural reflex, spontaneous circling) was significantly decreased from 3 to 21 days post-ICH by treatment with FK-506. Skilled use of the forelimb ipsilateral to the ICH was improved and sensory neglect of the same limb was decreased 8-9 weeks post-ICH in rats treated with FK-506. However, neuronal loss assessed 9 weeks post-ICH was not different in the treated and untreated rats.

Hydrocephalus is a pathological dilatation of the cerebrospinal fluid (CSF)-containing ventricles of the brain. Damage to periventricular white matter is multifactorial with contributions by chronic ischemia and gradual physical distortion. Acute ischemic and traumatic brain injuries are associated with calcium-dependent activation of proteolytic enzymes. We hypothesized that hydrocephalus is associated with calcium ion accumulation and proteolytic enzyme activation in cerebral white matter. Hydrocephalus was induced in immature and adult rats by injection of kaolin into the cisterna magna and several different experimental approaches were used. Using the glyoxal bis (2-hydroxyanil) method, free calcium ion was detected in periventricular white matter at sites of histological injury. Western blot determinations showed accumulation of calpain I (mu-calpain) and immunoreactivity for calpain I was increased in periventricular axons of young hydrocephalic rats. Proteolytic cleavage of a fluorogenic calpain substrate was demonstrated in white matter. Immunoreactivity for spectrin breakdown products was detected in scattered callosal axons of young hydrocephalic rats. The findings support the hypothesis that periventricular white matter damage associated with experimental hydrocephalus is due, at least in part, to calcium-activated proteolytic processes. This may have implications for supplemental drug treatments of this disorder.

During attempts to develop the intraluminal suture model of transient middle cerebral artery occlusion (MCAO) in the Sprague Dawley strain of rats, we noticed a wide variability in lesion size seen with T2-weighed magnetic resonance imaging (MRI) or histopathology, as well as in scores for behavioral indices. It was our intent to examine the results of the study carefully and determine whether there were strong point-to-point correlations between the degree of lesion size determined from T2-weighted MRI or histopathology and intermediate or long-term neurologic/behavioral assessments. Baseline behavioral scores for forelimb dexterity (staircase test) were obtained on all animals in the period before receiving 60 minutes of transient MCAO. After MCAO, animals were tested at specified intervals from 1 to 21 days for composite neurologic deficits. T2-weighted MRI was taken at 2 and 7 days post-MCAO. At 30 and 60 days post-MCAO, animals were retested in the staircase test with subsequent histopathologic examination of the brains. Indeed, there were highly significant correlations between lesion size determined by MRI and histopathology. The damage observed in the T2-weighted MRI, as well as the size of the histopathologic lesions, were in turn highly correlated to deficiencies observed in the composite neurologic assessments, as well as to deficits at 30 and 60 days post-MCAO for skilled use of the contralateral forepaw (damaged side). In the latter test, the correlations were somewhat less significant for the ability of rats to reach for food with the ipsilateral forepaw (undamaged side).

Intracerebral hemorrhage is associated with stroke and head trauma. The purpose of this study was to study brain inflammation and cell death in adult rats 1 h to 4 weeks after injection of blood into the striatum. Terminal dUTP nick-end-labeling positive dying cells were evident 4 h to 4 weeks post-hemorrhage. Neutrophil infiltration was brief and peaked at 48 h. CD8a immunoreactive lymphocytes, possibly natural killer cells, became apparent at 48 h and persisted for 1 week. Microglial reaction was evident at 4 h and persisted for 4 weeks. We conclude that extravascular blood causes a mixed inflammatory cell reaction in brains that is maximal from 48-72 h following hemorrhage. This is associated with death of brain cells over a prolonged period of at least 4 weeks.

Background and Purpose —Hypertension is a major risk factor for ischemic and hemorrhagic stroke and may also cause more chronic and subtle brain injury. Progressive brain changes in a rat model of renal hypertension have been assessed to better understand the pathogenesis of hypertensive brain damage. Methods —Young adult rats were made hypertensive by partial occlusion of both renal arteries. MR images of brain were obtained weekly, and histopathological outcome was assessed. A separate group of rats was used to measure brain specific gravity and Evans blue dye content as an indicator of extravasation. Results —Rats developed maximal mean systolic blood pressures of 173 to >300 mm Hg, reaching a plateau in 6 to 8 weeks. Rats whose mean systolic pressure never exceeded 210 mm Hg never had brain lesions, while rats whose mean systolic pressure exceeded 276 mm Hg consistently developed brain lesions. Brain T2 values increased with increasing blood pressure. Lesions seen on MRI corresponded to those seen histologically. MRI also demonstrated transient brain expansion, probably due to diffusely increased water content, and rarely demonstrated focal cortical edema, which had no histological correlate. These transient phenomena, as well as hemorrhagic and ischemic infarcts, occurred mainly during the phase of climbing blood pressure and early stages of stable hypertension. Conclusions —Serial MRI reveals aspects of hypertensive brain disease that cannot be studied by histological examination alone. The observed phenomena are likely related to loss of autoregulation and/or blood-brain barrier integrity. Breach of blood vessel integrity is less likely once the vessels become accustomed to high pressures.

We investigated whether the changes detectable with magnetic resonance imaging techniques during and after an episode of cerebral hypoxia-ischemia differ in immature and older brain. Diffusion weighted (DW) and T2-weighted (T2W) images were repeatedly acquired before, during, and after an episode of cerebral hypoxia-ischemia (unilateral carotid artery occlusion plus hypoxia) in 2- and 4-wk-old rats lightly anesthetized with isoflurane. Areas of increased brightness were detected in DW images from both 2- and 4-wk-old rats by 10-20 min after the start of hypoxia. These hyperintense areas increased during hypoxia, comprising 60.8 ± 4.9% and 30.5 ± 2.7% of the brain image at the level of the thalamus in 2-wk-old and 4-wk-old animals, respectively (p < 0.003). Hyperintense areas (e.g. 27.0 ± 8.3%) also appeared in T2W images during hypoxia-ischemia in 2-wk-old animals, but these did not occur in 4-wk-old animals (p < 0.02). This observation was reflected in T2, which increased during hypoxia-ischemia in the 2-wk-old but not the 4-wk-old group. By 60 min after the termination of hypoxia-ischemia in either age group, areas of hyperintensity resolved and then reappeared 24 h later on both DW and T2W images. Thus, irrespective of age, magnetic resonance imaging changes during transient hypoxia-ischemia generally recover with a delayed or secondary increase in DW and T2W hyperintensity hours later. Immature brain differs from older brain primarily with respect to some combination of hypoxic/ischemic cellular or biochemical changes, that are detectable as increases in T2 within 2-wk-old but not 4-wk-old animals.

We hypothesized that hydrocephalus can cause death of brain cells and that generation of new brain cells might compensate for the cell loss. Hydrocephalus was induced in 3-week-old rats by injection of kaolin into the cisterna magna. The brains were studied 1 to 4 weeks later by histochemical, immunochemical, and ultrastructural methods. The ventricles enlarged progressively. Some axons in the corpus callosum were injured as early as 1 week, but axonal damage was not prevalent until 4 weeks when ventriculomegaly became severe. Dying cells detected by DNA end labeling and often identified as oligodendrocytes by electron microscopy were evident in white matter. Late-stage hydrocephalus was associated with a significant increase in the quantity of dying cells. Hydrocephalus was associated with increased Ki67 labeling and bromodeoxyuridine incorporation in the subependymal zone. Reactive changes were identified among astrocytes, oligodendroglia, and microglia. We conclude that hydrocephalus causes, in addition to axonal injury, gradual cell death in the cerebrum, particularly the white matter. The brain response includes production of new glial cells, but whether the new cells play any beneficial role remains unknown.

Background and Purpose —Deep intracerebral hemorrhage is associated with considerable mortality and morbidity, but the value of surgical therapy is debatable. The purpose of this study was to evaluate whether aspiration of the hematoma in a rodent model of intracerebral hemorrhage could improve final neurological outcome. Methods —Intracerebral hemorrhage was induced in 2 groups of rats by injection of bacterial collagenase into the caudate nucleus. In 1 group of rats, streptokinase was used to lyse the hematoma 4 hours after hemorrhage induction, and the clot was then aspirated. Behavioral function was evaluated repeatedly until the rats were killed 7 weeks after collagenase injection. Histology was used to assess neuronal loss, astroglial proliferation, and overall brain morphology. In a second experiment, brain water was measured at 24 hours. Results —The treated rats performed significantly better than controls on a motor-behavior evaluation on days 1, 2, and 28 after aspiration. Skilled forelimb testing performed for 3 weeks after the global behavior evaluations showed a significant deficit of contralateral forelimb function in both groups, but there was no significant difference between the 2 groups. Neuronal loss in the perihematoma striatum was significantly greater in untreated compared with treated rats. In most rats, structural damage extended into the internal capsule and thalamus. Conclusions —Aspiration of the hematoma after collagenase-induced hemorrhage slightly improved acute functional outcome and reduced neuronal loss from the striatum. Further studies are required to delineate the mechanism of the effect.

Hydrocephalus is characterized by enlargement of the cerebral ventricles. The behavioral disturbances are, in some cases, rapidly reversible by surgical treatment suggesting that there may be a functional impairment of neurons. Hydrocephalus was induced in 3-week old rats by kaolin injection into the cisterna magna. Parietal cerebrum and striatum content of monoamine neurotransmitters and amino acids were assayed by high performance liquid chromatography (HPLC), 1, 2, or 4 weeks after induction of hydrocephalus. The ventricles exhibited progressive enlargement which was partially reversed by surgical treatment. Cerebral water content was increased at all stages. Increased levels of cerebral aspartate and glutamate suggest that there is the potential for excitatory neurotoxicity. The increase in cerebral taurine correlated negatively with the increase in water content. Cerebral concentrations of norepinephrine and serotonin, and its metabolite 5-HIAA, were increased at 1 and 2 weeks suggesting an increase in their turnover during the early stages of ventricular dilatation. Dopamine and its metabolite DOPAC were transiently diminished in the striatum at 1 and 2 weeks, respectively, suggesting that axonal projections from the brainstem may be impaired. We conclude that the effect of hydrocephalus on amino acids and monoamines varies regionally. Due to increased water content, there may be dilution effects in whole tissue, therefore, it is important to make determinations on the basis of protein content.

To determine the role of the Tat protein of the human immunodeficiency virus type 1 (HIV-1) in the pathogenesis of HIV-1 associated dementia, recombinant Tat was injected intraventricularly as a single or repeated dose into male Sprague-Dawley rats. Histopathological evaluation showed an initial infiltration of neutrophils one day after Tat injection, followed by macrophages and lymphocytes by 7 days. Tat-injected brains also exhibited astrocytosis, apoptotic cells, and ventricular enlargement 7 days following the last injection. Nuclear magnetic resonance spectroscopic analysis of tissue extracts of hippocampi from Tat-injected rats showed a decrease in the glutamate/g aminobutyric acid ratio. We conclude that the transient extracellular exposure of the central nervous system to Tat protein of HIV can cause a cascade of events leading to the influx of inflammatory cells, glial cell activation, and neurotoxicity.

Objective: The objective of this study was to investigate flow distribution during retrograde and antegrade cerebral perfusion with India ink as a marker. Methods: Ten pigs received cerebral perfusion with a solution containing 50% filtered India ink for 5 minutes either antegradely through both internal carotid arteries at a flow of 180 to 200 ml/min ( n = 5) or retrogradely via the superior vena cava at a flow of 300 to 500 ml/min ( n = 5). The brains were then fixed for quantitative measurement of the density of ink-filled capillaries (reported as a percentage of the total selected area). The assessment was done with the use of an in-house software program. Results: In the antegrade cerebral perfusion group, the intracranial arterial and venous systems were completely filled with ink. The gray matter was colored uniformly black, and light coloring was observed in the white matter. During retrograde cerebral perfusion, the majority of ink was returned to the inferior vena cava, and only a small amount of ink was found in the innominate artery draining from the brain. Massive ink filling was observed in the sagittal sinus and other venous sinuses in all the pigs. Vessels on the surface of the brain and large vessels in the brain were also well filled with ink. However, only 10% of capillaries were filled with ink during retrograde cerebral perfusion relative to the number observed with antegrade cerebral perfusion. Conclusions: Retrograde cerebral perfusion supplies a limited amount of blood to brain tissue, which flows mainly through superficial and large deep cerebral vessels. (J Thorac Cardiovasc Surg 1997;114:660-5)

Apoptotic cell death is associated with condensation and fragmentation of the nuclear chromatin into apoptotic bodies. Such nuclei occur frequently following cerebral hypoxia-ischemia in 1 week old rats but seem sparse in older animals. We investigated the age dependence by counting pyknotic or punctate chromatin containing cells in the cerebral cortex of 1,2 and 4 week old animals subjected to cerebral hypoxia-ischemia. In 1 week old animals the majority of injured neurons in the cortex had nuclei with punctate condensed chromatin but few were pyknotic. In the 2 and 4 week old animals the majority of injured cells were pyknotic (P < 0.002). Electron microscopic studies demonstrated the presence of apoptotic bodies. The results suggest that immature brain retains a part of the developmental cell death "program' that is activated following hypoxia-ischemia.

Microtubule-associated protein 2 (MAP2) immunohistochemical labeling in the hippocampus was studied to assess the protective effect of brain perfusion during surgery requiring hypothermic circulatory arrest in 24 pigs exposed to anesthesia alone (control), 120 min of complete circulatory arrest at 15 degrees C, min of retrograde cerebral perfusion at 15 degrees C, or 120 min of anterograde cerebral perfusion at 15 degrees C. Pigs were reperfused for 60 min and sacrificed. In the control anterograde perfusion groups, the intensity of MAP2 labeling was similar in all regions of the hippocampus. Circulatory arrest and retrograde perfusion resulted in significant reduction of MAP2 labeling (28% and 38% respectively of control, P0.001) of neurons in the CA1 region. MAP2 labeling may be useful for assessing early damage in the hippocampus in this model.

Background. Antegrade and retrograde cerebral perfusion during hypothermic circulatory arrest (HCA) has been reported to provide better brain protection during operation than hypothermic circulatory arrest alone. However, the efficacy of these techniques remains to be fully determined, especially when used for prolonged periods. We used a pig model to evaluate the histophathologic consequences of HCA and the potential benefit of cerebral perfusion during HCA. Methods. Twenty-two pigs were divided into four groups and exposed to either anesthesia alone, 120 minutes of HCA (15°C), 120 minutes of retrograde cerebral perfusion at 15°C during HCA, or 120 minutes of antegrade cerebral perfusion at 15°C during HCA, and then reperfused for 60 minutes under cardiopulmonary bypass at 37°C. The brains were perfusion fixed at the end of the experiments and examined by light microscopy. Results. There were no morphologic changes in any areas of the brains in the anesthesia group, and very minor changes in some areas of the brains in the antegrade cerebral perfusion group. Varying severity of neuronal damage was found in the brains of all the pigs in the HCA and retrograde cerebral perfusion groups. The severity of ischemic damage in the brain showed the following descending order: hippocampus (CA4), caudate nucleus, cerebral cortex, putamen, thalamus, Purkinje cells of the cerebellum, pons, and mesencephalic gray matter. In the hippocampus the order of damage was CA4, CA3, polymorphous layer of the dentate gyrus, prosubiculum, CA2, CA1, and granule cell layer of the dentate gyrus. The damage in the retrograde cerebral perfusion group was less severe relative to the HCA group in many areas (no significance except mesencephalic gray matter). Conclusions. These results demonstrate that the pattern of neuronal damage in pigs subjected to HCA and retrograde cerebral perfusion differs from the traditional pattern in that the caudate nucleus and hippocampal CA4 region are the most vulnerable to ischemia-hypoxia. Our results also suggest that antegrade cerebral perfusion prevented ischemic damage to the brain and retrograde cerebral perfusion provided some protection but moderately severe damage occurred.

In developing rabbit brain we studied expression of metalloproteinases (MMP) 1 and 3 by in situ hybridization and MMP2 and tissue and urokinase-type plasminogen activators (tPA and uPA) by immunohistochemistry. All are detected in developing cell populations. Mature olfactory bulb neurons express MMP1 and MMP3. uPA is expressed by glial cells during myelination and by mature cortical neurons. MMP2 is expressed by mature subpial and perivascular astrocytes.

Background The chemokine CCL2 has an important role in the recruitment of inflammatory cells into the central nervous system (CNS). A transgenic mouse model that overexpresses CCL2 in the CNS shows an accumulation of leukocytes within the perivascular space surrounding vessels, and which infiltrate into the brain parenchyma following the administration of pertussis toxin (PTx). Methods This study used contrast-enhanced magnetic resonance imaging (MRI) to quantify the extent of blood–brain barrier (BBB) disruption in this model pre- and post-PTx administration compared to wild-type mice. Contrast-enhanced MR images were obtained before and 1, 3, and 5 days after PTx injection in each animal. After the final imaging session fluorescent dextran tracers were administered intravenously to each mouse and brains were examined histologically for cellular infiltrates, BBB leakage and tight junction protein. Results BBB breakdown, defined as a disruption of both the endothelium and glia limitans, was found only in CCL2 transgenic mice following PTx administration and seen on MR images as focal areas of contrast enhancement and histologically as dextrans leaking from blood vessels. No evidence of disruption in endothelial tight junctions was observed. Conclusion Genetic and environmental stimuli were needed to disrupt the integrity of the BBB in this model of neuroinflammation.

U primigravide de 13 ans a ete soumise a une evaluation echographique a 24+4 semaines de gestation. Dans la tete fœtale, des zones echogenes ont ete constatees dans le lobe frontal droit, aux cotes du ventricule lateral (Figure 1). L’IRM a confirme qu’il s’agissait d’une hemorragie intraparenchymateuse (Figure 2). Le travail a ete provoque a 26+4 semaines; la patiente a accouche d’un fœtus mort-ne de sexe masculin. L’autopsie a confirme la presence d’une importante hemorragie subaigue de l’eminence ganglionnaire du cote droit mettant en cause le parenchyme des regions frontales parietale et posterieure.

The brain is enclosed in an unyielding container. Any process that causes expansion of one intracranial component necessarily causes compression of the other components. When there is rapid expansion of an intracranial space-occupying lesion, for example edema in the area of injured brain parenchyma, cerebrospinal fluid and venous blood are displaced from the intracranial compartment and the brain is distorted. Ultimately intracranial pressure rises and, when it exceeds the arterial perfusion pressure, blood flow to the brain is impaired. Thereafter neurons die and the opportunity for functional recovery is lost. Because we are currently incapable of resurrecting neurons which die during the initial ischemic or traumatic insult, we must concentrate our therapeutic efforts on the prevention of secondary neuronal injury. This is most often the consequence of brain swelling due to edema and in order to control brain swelling it is imperative that we understand the pathophysiology of brain edema.