Molecular mechanisms of glial swelling in vitro.

The pathophysiological chain of events occurring during cerebral ischemia is still poorly understood on a molecular level. Therefore, an in vitro model to study glial swelling mechanisms, using C6 glial cells under controlled extracellular conditions, has been established. Flow cytometry serves to determine even small cell volume changes. In this report, the effects of anoxia and acidosis on glial swelling are summarized. Anoxia alone, or in combination with iodoacetate to inhibit anaerobic glycolysis, did not cause an increase of glial volume for up to 2 h. Acidification of the incubation medium below pH 6.8, on the other hand, was immediately followed by cell swelling to 115% of normal. Amiloride or the absence of bicarbonate and Na+ in the medium significantly reduced glial swelling. The data support the contention that swelling results from an activation of the Na+/H+-antiporter to control intracellular pH. It is suggested that swelling in an ischemic penumbra is promoted by this mechanism. Therapeutic approaches to control cerebral pH might be useful to protect brain tissue in cerebral ischemia.

Reaction of periventricular tissue in the rat fourth ventricle to chronically placed shunt tubing implants.

The reaction of periventricular tissue to shunt tubing chronically implanted in the fourth ventricle of the rat was investigated by correlative scanning and transmission electron microscopy. Sterile silicone tubing with four 0.4 mm diameter holes was inserted into the fourth ventricle of adult Sprague-Dawley rats through an incision in the atlantooccipital membrane and the animals were killed at postoperative intervals of 5 and 8 weeks. Reactive changes that could be correlated with the extent of contact with the implant occurred in the periventricular tissue. The ependyma lining the ventricle underwent a progressive loss of cilia and microvilli, became attenuated and, in circumscribed areas, was lost entirely. A significant subependymal gliosis accompanied these changes. In regions denuded of ependyma, neurons and glia were exposed directly to the cerebrospinal fluid. Eruptions of periventricular tissue corresponding precisely to the location of holes in the implanted tubing were observed on both the vermal surface of the cerebellum and the floor of the ventricle. Evaginations from the surface of the inferior vermis and the floor of the ventricle were most prevalent at 5 and greatest at 8 weeks postimplantation, respectively. Gliosis combined with mechanical factors are believed to be responsible for development of these periventricular tissue evaginations, which may be a factor in the pathogenesis of cerebrospinal fluid shunt obstruction in treated human hydrocephalus.

Excitatory amino acids and epilepsy-induced changes in extracellular space size.

Convulsant and stimulus induced seizures are associated with Ca, Na, K and Cl concentration changes in the extracellular space (ES), which are a resultant of transmembrane ionic fluxes and of changes in the ES size. The ES decreases on average by 30% during a single seizure. An analysis of the causes of ES size changes reveal a large contribution from the spatial glia K buffer mechanism which may account for up to 60% of the ES decreases. NaCl and KCl uptake into cells as well as increases in intracellular osmolarity due to anaerobic glycolysis contribute less to the local cytotoxic edema but account for a net gain of osmotic active particle at the site of the focus. Excitatory amino acids such as glutamate, aspartate, N-methyl-D-aspartate (NMDA), kainate and quisqualate also lead to Na, Cl and eventually Ca uptake into cells and to release of K with dose dependent decreases in [Na]o, [Ca]o and [Cl]o, increases in [K]o and transient decreases in ES size by up to 80% which are possibly associated with a net reduction of osmotically active particles. The predominant cause for this cytotoxic edema is NaCl uptake into cells but spatial K buffering through glial cells also contributes to this type of edema. The possible consequences of the various ion movements and the changes in osmolarity as well as ES size for tissue vulnerability are discussed.

Effect of intrauterine growth retardation on developmental changes in DNA and [14C]thymidine metabolism in different regions of rat brain: histological and biochemical correlations.

Intrauterine growth retardation was induced in the rat by clamping the uterine artery on day 17 of gestation. The effect of hypotrophy on DNA synthesis was studied in two different cerebral structures: hippocampus and cerebellum. Accumulation of DNA in these structures was biochemically measured in parallel to the incorporation of methyl-[14C]thymidine into nucleic acid at different ages and correlated with autoradiography. The various metabolites of thymidine in acid-soluble fraction were determined by using chromatographic procedures. Phosphorylation defects or reduced utilization of thymidine were found in hypotrophic rats and may delay the DNA synthesis. An essay of catch-up occurred with a different timing according to the cerebral region studied. A morphological and DNA synthesis. An essay of catch-up occurred with a different timing according to the cerebral region studied. A morphological and autoradiographic study after incorporation of [3H]thymidine was carried out in parallel. The neuronal and glial components of cytogenesis were analyzed separately and a good correlation was observed between histological and biochemical data in both groups of animals.

Neoplastic transformation of newborn rat oligodendrocytes in culture.

We have developed a model to study the neoplastic transformation of rat oligodendrocytes in culture. This procedure utilizes a technique previously developed by McCarthy and de Vellis which allows the preparation of 99% pure astrocyte and oligodendrocyte populations from 1- to 2-day-old rat cerebral cortices. Pregnant rats on the 19th day of gestation were given injections with either ethyl nitrosourea (10 micrograms/g body weight) in phosphate-buffered saline or phosphate-buffered saline, and oligodendrocyte cultures were prepared. Oligodendrocytes appear to be unstable in culture since transformation was observed with cells derived from either pups from pregnant rats either treated with nitrosourea or phosphate-buffered saline. Transformation required 78 to 108 days and 3 to 9 passages, at which time a marked increase in cellular proliferation was observed. The possibility that the transformed cells were derived from a nonoligodendroglial cell was excluded by the following evidence. Light and scanning electron micrographs of the transformed cells revealed cytological features essentially similar to those of primary oligodendroglial cultures. Furthermore, 2 biochemical oligodendroglial markers, the induction of lactate dehydrogenase by N6,O6-dibutyryl cyclic adenosine 3':5'-monophosphate and the presence of 2',3'-cyclic nucleotide 3' phosphohydrolase, were also retained. Conversely, another oligodendroglial marker, the hydrocortisone induction of glycerol phosphate dehydrogenase, was not found in any of the cell lines. These transformed cells grew as tumors when injected intracranially into 21-day-old rats. Histologically, these tumors did not appear as classical oligodendrogliomas, but their oligodendroglial origin was confirmed since the tumor tissue contained 2':3'-cyclic nucleotide 3'-phosphohydrolase activity, and the cells which grew from tumor explant cultures morphologically appeared similar to the parent cell line. The transformed cells were also characterized for in vitro properties which correlate with the expression of tumorigenicity. The transformed cells exhibited anchorage-independent growth and were agglutinated by concanavalin A treatment. Changes in fibrinolytic activity were not an exclusive property of transformed glial cells. This model should now allow us to study various mechanisms involved in the neoplastic transformation of oligodendrocytes.

Thiamine deficiency limits glucose utilization and glial proliferation in brain lesions of symptomatic rats.

The effects of thiamine (B1) deficiency on local CMRglu (LMCRglu) in the vestibular nuclei were studied with the 14C-2-deoxyglucose autoradiographic method in awake asymptomatic and symptomatic rats. Animals on the B1-deficient diet for 98 days developed symptoms of ataxia and opisthotonos. The results show that B1 deficiency produces: (1) bilateral vestibular nuclei lesions in symptomatic animals; (2) very low LCMRglu rates in these lesions; and (3) limitation of glial proliferation in the lesions. Giving B1 to B1-deficient symptomatic animals produced a cellular proliferation response consisting mostly of microglia in the lesioned areas of the vestibular nuclei and a high LCMRglu rate in the regions of microglial proliferation.

Variations in the perineuronal glial changes after different types of nerve lesion: light and electron microscopic investigations on the facial nucleus of the rat.

The perineuronal glial changes were studied by light and electron microscopy after avulsion or a crush lesion of the facial nerve in rats. The changes consisted of proliferation of microglia, ensheathment of neurons by thin astrocytic processes, and separation of the synaptic boutons from the neuronal surface. Quantitative estimates of the glial proliferation were made with the light microscope. In spite of marked differences in the acute nerve cell reaction, 4 days after the two types of lesion the glial and synaptic changes did not differ significantly. Ultimately, all changes were reversible after crush lesions, while neuronophagia occurred after nerve avulsion. It is concluded that the acute synaptic and glial reactions were not influenced by the type of nerve lesion or the severity of the nerve cell reaction, but the latter stages differed depending upon whether the neurons recovered or disintegrated.

The early stages of Wallerian degeneration in the severed optic nerve of the newt (Triturus viridescens).

The initiation of Wallerian degeneration in the severed optic nerve of the newt (Triturus viridescens) was very rapid and intense. Significant degeneration of nonmyelinated axons was observed as early as six hours after lesion (h.a.l.) and was almost complete by 48 h.a.l. Initial degeneration of non-myelinated axons began in "extracellular digestion chambers" formed between burgeoning ependymoglial processes. The remaining fragments and debris were later phagocytized by surrounding ependymoglial processes. Many axons of myelinated fibers have degenerated as early as 6 h.a.l. However, the overall population of myelinated axons degenerates at a much slower rate than nonmyelinated ones, for many of them appear intact as late as 48 h.a.l. Some myelin sheaths show significant signs of degeneration by 6 h.a.l. Indeed, by this time a number of myelinated fibers have completely degenerated leaving only large vacuolated spaces in the nerve parenchyma. Swelling and vacuolization of the sheath are among the earliest signs of myelin degeneration. The ependymoglial cell response to optic nerve lesion is manyfold and dramatic. By 6 h.a.l. there are signs of burgeoning ependymoglial processes which begin to resemble scar formation (gliosis) by 48 h.a.l. The morphological evidence is consistent with the concept of an important phagocytic role of ependymoglial cells during the early stages of optic nerve degeneration.

[Posttraumatic epilepsy. Clinical aspects and electrophysiological foundations]. / Posttraumatische Epilepsie Klinik und elektrophysiologische Grundlagen

The review of the syndrome is centered on time course, prognosis and prophylaxis. Prophylactic anticonvulsive treatment depends on the prognosis. The latter is dependent on the risk to develop epilepsy after a certain type of head injury. Treatment should begin in the high risk group of patients before seizures become overt. In order to avoid preventive treatment in unselected posttraumatic cases, predicting the risk in each case is most important. Therefore the literature was reviewed selecting injuries and complications of high risk. Knowledge on pathophysiologic mechanisms in the epileptogenic focus to a large extent stems from animal experiments using a model of chronic recurrent seizures. Basic electrophysiologic mechanisms responsible for focal electroencephalographic changes were studied by recording from single cells. Profound abnormalities in patterns of neuronal discharge were found as well as morphological and functional changes of glia in the epileptogenic scar. It appears that glia is important in generating focal electroencephalographic spiking. By controlling intercellular potassium transport glia modulates neuronal activity.