PubMed Health⌕ Search

Biomedical subjects

M D Norenberg

Publications and source records attributed to M D Norenberg.

At least 19 recordsLinked to original sources
PubMed1992

Role of calmodulin and protein kinase C in astrocytic cell volume regulation.

We investigated the role of Ca(2+)-dependent protein kinases in the regulation of astrocytic cell volume. Calmodulin (CaM) antagonists were used to inhibit CaM and thus Ca2+/CaM-dependent protein kinase. The effect of these inhibitors as well as activators and inhibitors of protein kinase C (PKC) on astrocytic volume was measured in response to hypoosmotic stress and under isoosmotic conditions. In conditions of hypoosmolarity, CaM antagonists had no effect on swelling, but inhibited the regulatory volume decrease. PKC activation facilitated the swelling induced by hypoosmotic stress. PKC inhibitors induced cell shrinkage and inhibited the initial phase of regulatory volume decrease, whereas PKC down-regulation caused pronounced swelling and partial inhibition of regulatory volume decrease. In isoosmotic conditions, CaM antagonists and PKC activation did not affect astrocytic volume, but PKC inhibitors caused shrinking and PKC down-regulation led to swelling of these cells. These studies indicate the importance of Ca(2+)-dependent protein kinases in the regulation of astrocytic cell volume.

Animals↗
PubMed1992

Involvement of second messengers and protein phosphorylation in astrocyte swelling.

In a hypoosmotic model of astrocyte swelling, we found that Ca2+ and intracellular signals such as diacylglycerol and inositol phosphate, as well as protein phosphorylation systems, are implicated in the generation and (or) modulation of volume regulatory processes. Cyclic AMP, which also has a significant effect on astrocyte volume regulation, in addition influences some of these second messengers.

Animals↗
PubMed1991

ATP-evoked calcium signal stimulates protein phosphorylation/dephosphorylation in astrocytes.

Extracellular adenosine 5'-triphosphate (ATP)-evoked increases in intracellular calcium and the consequent stimulation of calcium-mediated protein phosphorylation systems were investigated in primary cultures of rat cerebral cortical astrocytes. Measurement of calcium responses in fura-2-loaded astrocytes indicated that extracellular ATP stimulated a transient calcium peak followed by a sustained increase in intracellular calcium which declined to baseline when external calcium was removed, thereby indicating that ATP evokes mobilization of internal calcium as well as influx of external calcium. Protein phosphorylation studies revealed that application of extracellular ATP resulted in increased phosphorylation of 55 and 52 kDa proteins (4-fold and 2-fold, respectively) and decreased phosphorylation of 24 and 21 kDa proteins (approximately 50% for each protein). These effects were time- and dose-dependent. The changes in phosphate incorporation were (a) inhibited by lanthanum, (b) reduced when calcium was omitted from the bath and (c) mimicked by ionomycin, thus suggesting that the ATP-induced changes in protein phosphorylation were dependent on increased levels of intracellular calcium. Adenosine diphosphate (ADP) gave similar, but reduced, effects while adenosine and guanosine triphosphate (GTP) were ineffective, findings consistent with activation of P2 purinergic receptors. The 52 kDa protein co-migrated with glial fibrillary acidic protein. These results support the premise that calcium-dependent protein kinases and phosphatases are transducing elements for the calcium signal brought about by activation of P2 purinergic receptors in astrocytes. Since ATP is released from neurons and endothelial cells, this signal transduction mechanism may be an important component of neuronal- and endothelial-astrocytic communication.

Adenosine Triphosphate↗
PubMed1991

Peripheral-type benzodiazepines inhibit proliferation of astrocytes in culture.

Peripheral-type benzodiazepine (BZD) receptors have been identified in brain and are predominantly localized to astrocytes. To determine their potential role in controlling astroglial proliferation, DNA synthesis, growth curves and mitotic index were investigated in primary astrocyte cultures which had been exposed to Ro5-4864 (a peripheral-type BZD ligand) and PK11195 (a peripheral-type BZD receptor antagonist). There was a dose-dependent inhibition of mitosis when two-week-old cells in culture were exposed to 50 nM, 500 nM, 1 microM and 10 microM Ro5-4864 for 24 h. Exposure of 5-, 8-, 12- and 15-day-old cultures to Ro5-4864 and PK11195 for 24 h did not affect growth rate and DNA synthesis; however, continuous exposure to 10 microM Ro5-4864 caused a persistent inhibition of cell growth and [3H]thymidine incorporation (P less than 0.05) while nanomolar concentrations did not cause any significant change. Concurrent administration of Ro5-4864 with PK11195 resulted in a partial reversal of Ro5-4864-induced inhibition in DNA synthesis and mitosis. These results indicate that peripheral-type BZDs are capable of inhibiting proliferation of astrocytes in culture.

Animals↗
PubMed1991

Aluminum chloride stimulates the release of endogenous glutamate, taurine and adenosine from cultured rat cortical astrocytes.

Primary astrocyte cultures derived from neonatal rat cerebral cortex were treated for 5 min with 0.5 mM or 5.0 mM AlCl3, and the incubation medium was analyzed by HPLC for the content of released glutamate (Glu), taurine (Tau), serine (Ser) and the nucleoside adenosine (Ade). At 0.5 mM, AlCl3 stimulated Tau release to about 170% of basal levels, but did not affect the release of the other compounds. Treatment with 5.0 mM AlCl3 enhanced the release of Tau, Glu and Ade, to 800%, 1000% and 250%, respectively, but decreased the release of Ser to 70% compared to basal levels. The enhanced release of these neuroactive compounds from astrocytes may contribute to changes in neural transmission known to accompany exposure to aluminum.

Adenosine↗
PubMed1991

Ammonia-induced astrocyte swelling in primary culture.

The effect of ammonia on water space of astrocytes in culture was determined as a means of studying the neurotoxicity of ammonia in fulminant hepatic failure (FHF). Treatment of primary astrocyte cultures obtained from neonatal rat cortices with 10 mM NH4Cl for 4 days resulted in a 29% increase in astrocytic water space, as measured by an isotopic method utilizing 3-O-methyl-[3H]-glucose. This effect was time- and dose-dependent. The ammonia-induced swelling was reversible as the water space in cultures treated with 10 mH NH4Cl for 3 days, and then returned to normal culture media for 1 day, was similar to control cultures. These findings suggest that elevated levels of ammonia lead to astrocyte swelling and may contribute to the brain edema in FHF.

3-O-Methylglucose↗
PubMed1991

Effect of methotrexate on glial fibrillary acidic protein content of astrocytes in primary culture.

We have recently reported that methotrexate (MTX) causes degenerative as well as reactive-like astroglial changes and alters the cell cycle kinetics of astrocytes in vitro. To further characterize the nature of the reactive-like changes that were noted by light and electron microscopy following MTX exposure, the glial fibrillary acidic protein (GFAP) content of astrocytes in culture was investigated by enzyme-linked immunosorbent assay, flow cytometry and double-immunofluorescent staining. An increase in GFAP content which did not correlate with drug dosage or DNA synthesis was noted in the MTX-treated cultures. It is postulated that this increase in GFAP content of astrocytes reflects an adaptive response to MTX-induced injury and partly explains the gliosis that is seen in methotrexate encephalopathy.

Animals↗
PubMed1990

The benzodiazepine receptor in cultured astrocytes from genetically epilepsy-prone rats.

Peripheral-type benzodiazepine (BZD) receptors were studied in cultured astrocytes derived from genetically epilepsy-prone and control rats. Scatchard analysis of the binding of [3H]Ro 5-4864 to astrocyte homogenates from epilepsy-prone rats showed 38% fewer BZD receptors (Bmax) as compared to controls. No significant change in affinity (Kd) was observed. These findings suggest that the astrocyte peripheral-type BZD receptor may be involved in some forms of epilepsy.

Animals↗
PubMed1990

Effect of ammonia on calcium homeostasis in primary astrocyte cultures.

Calcium influx, accumulation and efflux were studied in primary cultures of rat astrocytes treated with ammonium chloride. Treatment of the cells for 3 days with 10 mMN4Cl resulted in a 35% reduction in 45Ca influx. The decrease in calcium influx was dose-dependent between 2 and 10 mM NH4Cl. Short-term (30 min) exposure to ammonia had no effect on calcium influx. Calcium accumulation, as measured by 20-min exposure to 45Ca, decreased after treating cultures with 10 mM NH4Cl for one or 3 days; a greater effect was observed after the 3-day treatment. Studies with lanthanum, an inhibitor of calcium transport, indicated that the effect of ammonia was not due to non-specific leakage of calcium. Calcium efflux was not affected by exposure of the cultures to ammonium chloride. Purinergic-evoked calcium influx and mobilization was not altered by ammonia. While the mechanism(s) of calcium homeostasis affected by long-term hyperammonemia remain to be defined, these results suggest that reduced astrocytic calcium may be related to the pathogenesis of ammonia-related disorders such as hepatic encephalopathy.

Adenosine Triphosphate↗
PubMed1990

L-methionine-DL-sulfoximine induces massive efflux of glutamine from cortical astrocytes in primary culture.

L-Methionine-DL-sulfoximine (MSO) is a potent convulsant which metabolically and morphologically primarily affects astroglia, and the links between the gliotoxic and neurophysiological effects of MSO are not clear. A 5 min treatment with 3 mM MSO increased the rate of efflux of newly loaded radiolabelled glutamine (Gln) from rat cortical astrocytes in primary culture to more than 400% of the basal efflux. MSO did not affect the efflux of the neurotransmitter amino acids GABA or D-aspartate (a non-metabolizable analogue of L-glutamate), under the same experimental conditions. MSO-induced overflow with Gln, which according to a recent account strongly interacts with the N-methyl-D-aspartate (NMDA) receptor complex, may contribute to the convulsive action of the drug.

Animals↗
PubMed1990

Effect of phenol and sodium octanoate on the astrocyte benzodiazepine receptor.

Alterations in the benzodiazepine (BZD) receptor system have been proposed as key factors in the pathogenesis of hepatic encephalopathy (HE). To date, the focus of research has been exclusively on the central-type neuronal receptor. However, astrocytes also possess BZD receptors which are of the peripheral-type. In recent studies we found an increased affinity of the astrocyte BZD receptor, using [3H]Ro5-4864 as the ligand, after treatment of cell cultures with ammonia, an agent strongly implicated in HE. The present study was undertaken to determine whether other suspected toxins in HE (phenol and octanoic acid) produce comparable effects. Scatchard analysis of the binding of [3H]Ro5-4864 to astrocyte homogenates showed a significant decrease in Bmax in cells that had been treated with 0.5, 1.0 and 3.0 mM phenol (46%, 58% and 68%, respectively). The same homogenates also showed a significant decrease in Kd after treatment with 0.5 mM phenol. No change in either affinity or receptor number was seen with 0.5, 1.0 and 3.0 mM sodium octanoate. Our results indicate that phenol, but not sodium octanoate, has an effect on the astrocyte BZD receptor. Thus, different agents that have been implicated in HE produce varying effects on the astrocytic BZD receptor. These findings suggest that the astrocyte benzodiazepine receptor may be involved in the pathogenesis of HE.

Animals↗
PubMed1990

Herpes simplex virus type 1 infection of rat astrocytes in primary culture: effects of dibutyryl cyclic AMP.

Monolayer cultures of primary rat astrocytes grown with or without dibutyryl cyclic AMP (dBcAMP) for two weeks or longer were infected with round plaque-forming (Rd) or syncytia-forming (Syn) variants of herpes simplex virus type 1 (HSV-1). Infection with HSV-1 did not stimulate synthesis of glial fibrillary acidic protein (GFAP) or alter the general organization of the intermediate (glial) filaments in astrocyte cultures. However, the dBcAMP-treated astrocytes produced 10- to 100-fold lower titers of cell-free progeny HSV-1 than the untreated astrocyte cultures. Radiolabeled amino acid or glucosamine incorporated into acid precipitable cellular or viral glycoproteins was decreased by 10-25% in dBcAMP-treated astrocytes. Distinctive cell-rounding or syncytial cytopathology was produced by HSV-1 strains infecting untreated astrocytes, but the infected dBcAMP-treated astrocytes displayed only cell-rounding cytopathology. The dBcAMP-related effects on HSV-1 infection were specific to primary astrocyte cultures; they were not observed in HSV-1-infected human fibroblast cultures treated with dBcAMP. Comparison of HSV-1 infection of untreated versus dBcAMP-treated astrocytes suggests that the dBcAMP-induced "reactive" or differentiated state of the astrocyte can affect expression of virus-induced cytopathology and virus-specific polypeptide synthesis. The dBcAMP-treated primary astrocyte culture may afford a non-neoplastic, differentiated in vitro system for studying HSV-neural cell interactions.

Animals↗
PubMed1990

Ammonia induced decrease in glial fibrillary acidic protein in cultured astrocytes.

Previous studies of human hepatic encephalopathy (HE) have shown decreased levels of glial fibrillary acidic protein (GFAP) in Alzheimer type II astrocytes. In view of the important role of ammonia in the pathogenesis of HE, we carried out immunocytochemical and enzyme-linked immunosorbent assay (ELISA) studies on the effect of ammonium chloride (10 mM) on GFAP content in primary astrocyte cultures. There was a 39% loss of GFAP after a four day treatment. There was no fall in total cell protein. Potential mechanisms for this apparent selective loss of GFAP are discussed.

Ammonia↗