Janet M Miller

Reduction of astrogliosis and microgliosis by cerebrospinal fluid shunting in experimental hydrocephalus.

BackgroundReactive gliosis has the potential to alter biomechanical properties of the brain, impede neuronal regeneration and affect plasticity. Determining the onset and progression of reactive astrogliosis and microgliosis due to hydrocephalus is important for designing better clinical treatments.MethodsReactive astrogliosis and microgliosis were evaluated as the severity of hydrocephalus increased with age in hydrocephalic H-Tx rats and control littermates. Previous studies have suggested that gliosis may persist after short-term drainage (shunt treatment) of the cerebrospinal fluid. Therefore shunts were placed in 15d hydrocephalic rats that were sacrificed after 6d (21d of age) or after 21d (36d of age). Tissue was processed for Western blot procedures and immunohistochemistry, and probed for the astrocytic protein, Glial Fibrillary Acidic Protein (GFAP) and for microglial protein, Isolectin B4 (ILB4).ResultsIn the parietal cortex of untreated hydrocephalic animals, GFAP levels increased significantly at 5d and at 12d compared to age-matched control rats. There was a continued increase in GFAP levels over control at 21d and at 36d. Shunting prevented some of the increase in GFAP levels in the parietal cortex. In the occipital cortex of untreated hydrocephalic animals, there was a significant increase over control in levels of GFAP at 5d. This trend continued in the 12d animals, although not significantly. Significant increases in GFAP levels were present in 21d and in 36d animals. Shunting significantly reduced GFAP levels in the 36d shunted group. Quantitative grading of immuno-stained sections showed similar changes in GFAP stained astrocytes.Immuno-stained microglia were altered in shape in hydrocephalic animals. At 5d and 12d, they appeared to be developmentally delayed with a lack of processes. Older 21d and 36d hydrocephalic animals exhibited the characteristics of activated microglia, with thicker processes and enlarged cell bodies. Following shunting, fewer activated microglia were present.Histologic examination of the periventricular area and the periaqueductal area showed similar findings with the 21d and 36d animals having increased populations of both astrocytes and microglia which were reduced following shunting with a more dramatic reduction in the long term shunted animals.ConclusionOverall, these results suggest that reactive astrocytosis and microgliosis are associated with progressive untreated ventriculomegaly, but that shunt treatment can reduce the gliosis occurring with hydrocephalus.

Effects of hydrocephalus and ventriculoperitoneal shunt therapy on afferent and efferent connections in the feline sensorimotor cortex.

OBJECT The authors of previous studies have suggested that connectivity within the cerebral cortex may be irreversibly altered by hydrocephalus. To examine connectivity-related changes directly, the authors conducted a study in which they used an axonal tracer in an animal model of infantile hydrocephalus. METHODS In five hydrocephalic kittens low-pressure ventriculoperitoneal (VP) shunts were placed 10 to 14 days after induction of hydrocephalus by intracisternal kaolin injections. Wheat germ agglutinin-conjugated horseradish peroxidase was injected laterally into the motor cortex in hydrocephalic animals 9 to 15 days after kaolin injection, and 1, 2, and 4 weeks after VP shunt insertion in shunt-treated animals, and in age-matched controls. Reduction of antero- and retrograde labeling was most profound within the contralateral cortex and portions of the midbrain. Thalamic nuclei exhibited reductions in anterograde and retrograde labeling. Labeling within cell bodies of the ventral tegmental area decreased greatly in animals with untreated hydrocephalus, in which retrograde labeling was reduced in the locus coeruleus but did not affect the raphe nucleus. Shunt treatment increased both antero- and retrograde labeling of contralateral motor cortex to near-normal levels. Thalamic relay nuclei recovered antero- and retrograde labeling, although not to levels exhibited in controls. Shunt therapy restored cellular labeling within the ventral tegmental area and locus coeruleus. Recovery of labeling occurred as early as 7 days after shunt insertion. CONCLUSIONS Collectively, analysis of these data indicates the following. 1) Cortical connectivity involving both afferent and efferent pathways was impaired in untreated hydrocephalic animals. 2) Shunt therapy improved both cortical afferent and efferent connectivity. 3) Complete reestablishment of the cortical efferent pathways, however, did not occur. Cortical pathway dysfunction, if permanent, could cause many of the motor and cognitive deficits seen clinically in children with hydrocephalus.

Gene expression analysis of the development of congenital hydrocephalus in the H-Tx rat

To discover candidate genes in the pathogenesis of congenital hydrocephalus, gene arrays were utilized to analyze transcripts from the midbrain region of 5-day-old H-Tx rats; these animals develop hydrocephalus due to closure of their cerebral aqueduct between embryonic day 18 and post-natal day 5. Of the 15,924 transcripts assayed, we detected 47 differentially expressed transcripts representing 23 genes and 24 expressed sequence tags (ESTs); 17 transcripts (7 genes and 10 ESTs) were upregulated and 30 (16 genes and 14 ESTs) were downregulated in the hydrocephalic animals relative to control non-hydrocephalic animals. Seven of these genes, Cck, Nfix, Lgals3, Gsta1, Xdh, Tnf, and Tfpi-2, can be linked to hydrocephalus. In addition, 17 genes that displayed altered expression in our study are not currently known to be associated with the presence or development of hydrocephalus. These results indicate that a relatively few number of transcripts were found to be altered in the development of hydrocephalus in this model. This is the first experiment of its kind to identify changes in gene expression in a congenital model of rodent hydrocephalus that are occurring locally in the area surrounding the cerebral aqueduct. Studies are now needed to examine these candidate genes and their cognate proteins to delineate their role in hydrocephalus.

Inhibitory effects of minocycline on gliosis in the hydrocephalic H-Tx rat

Background Persistent gliosis, if present in the hydrocephalic brain, has the potential to alter the biomechanical properties of the brain, impair cerebral perfusion, impede neuronal regeneration and affect plasticity. The purpose of this study was to determine the ability of minocycline, a specific inhibitor of glial reactivity, to reduce glial scar formation in the H-Tx rat model of congenital hydrocephalus.

Lymphatic cerebrospinal fluid absorption is impaired in a kaolin-induced hydrocephalus model in the rat

Materials and methods Kaolin (n = 10) or saline as control (n = 9) was introduced into the basal cisterns of Sprague Dawley rats and the development of hydrocephalus was assessed using MRI. The degree of ventriculomegaly was calculated for each animal as the Evans ratio. Human serum albumin (125IHSA) was injected into the lateral ventricles. The enrichment of 125I-HSA in the olfactory turbinates at 30 minutes post injection provided an estimate of CSF transport through the cribriform plate into the nasal lymphatic vessels.

Pathophysiology of communicating hydrocephalus in two novel animal models

Background Communicating hydrocephalus (CH) occurs frequently but clinically-relevant animal models amenable to diagnostic imaging and cerebrospinal fluid shunting do not exist. This deficiency has inhibited our understanding of the pathophysiology of CH, which may be quite different from obstructive hydrocephalus. The purpose of this study was to perfect two novel models of CH and to characterize the histopathology that occurs in each.

Progression and reversibility of gliosis due to hydrocephalus in the H-Tx rat

The 49 Annual Meeting of the Society for Research into Hydrocephalus and Spina Bifida was held in Barcelona, Spain, from 29 June to 2 July 2005. We are especially grateful to Amparo Cuxart and Esther Pages who, together with their team, made us all very welcome. The city of Barcelona has much to offer, from the medieval elegance of the Gothic Quarter, to the rather more idiosyncratic architecture of Antonio Gaudi. Nor should it be forgotten that Picasso who spent much of his working life in Paris, was a Spaniard, who lived for many years in Barcelona. Barcelona is the main Catellan city, and we were treated to a feast of Catellan history, culture and language, as well as the more modern icons who play for Barcelona FC. The scientific sessions, held in the comfortable venue of the Hospital Universitari Vall d’Hebron, produced a great variety of topics, from cutting edge scientific research on hydrocephalus to very valuable work on the functional problems of children with spina bifida. Undoubtedly the highlight of the scientific programme was the Casey Holter Lecture given by John Mazur. John treated us to a fascinating insight into the orthopaedic management of patients with spina bifida, and brought to the task a working lifetime of experience. Finally a warm welcome was extended by Hugh Richards, to the 50 Annual Meeting which will be held in Cambridge, England from 30 August to 2nd September 2006.

Effects of hydrocephalus and ventriculoperitoneal shunting on afferent and efferent connections of the feline sensorimotor cortex

Materials and methods Obstructive hydrocephalus was induced in 10-day old kittens by intracisternal injections of kaolin; saline-injected animals served as controls. Hydrocephalic kittens received low-pressure ventriculoperitoneal (VP) shunts 10–14 days post-kaolin. Unilateral injections of wheat-germ agglutinin-conjugated horseradish peroxidase (HRP) were made into the sensorimotor cortex (Brodmanns areas 4, 6 and 3) in hydrocephalic animals (n = 5) at 9–15 days post-kaolin to represent the pre-shunted condition. Shunted animals (n = 5) received HRP injections at 1, 2 and 4 weeks post-shunt. Control animals (n = 5) received HRP injections at set time-points between 12–47 days of age to broadly match the sacrifice times of the shunted animals.

Gene alterations associated with closure of the cerebral aqueduct in hydrocephalic H-Tx rats

Materials and methods Midbrain regions which contain the cerebral aqueduct were micro-dissected from hydrocephalic and control animals at 5 days of age (n = 5). After RNA extraction and purification, total RNA was subjected to PCR techniques to generate cDNA that was subsequently labeled and hybridized (one brain per array) to the Rat 230 A oligonucleotide array from Affymetrix. Hybridization intensity for each array was measured using a confocal scanner, and results were normalized and reported as fold change differences. Raw expression data were subjected to a Students t-test as well as the Bayesian t-test, which is a method that helps control for variations resulting from small sample size. Only those transcripts passing both the fold change of 1.5 fold and t-test cut-offs (p < 0.05) were examined further.