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A S Bender

Publications and source records attributed to A S Bender.

At least 37 records · Page 2Linked to original sources
PubMed1994

Osmotic regulation of myo-inositol uptake in primary astrocyte cultures.

Uptake of myo-inositol by astrocytes in hypertonic medium (440 mosm/kg H2O) was increased near 3-fold after incubation for 24 hours, which continued for 72 hours, as compared with the uptake by cells cultured in isotonic medium (38 nmoles/mg protein). myo-Inositol uptake by astrocytes cultured in hypotonic medium (180 mosm/kg H2O) for periods up to 72 hours was reduced by 74% to 8 to 10 nmoles/mg protein. Astrocytes incubated in either hypotonic or hypertonic medium for 24 hours and then placed in isotonic medium reversed the initial down- or up-regulation of uptake. Activation of chronic RVD and RVI correlates with regulation of myo-inositol uptake. A 30 to 40 mosm/kg H2O deviation from physiological osmolality can influence myo-inositol homeostasis. The intracellular content of myo-inositol in astrocytes in isotonic medium was 25.6 +/- 1.3 micrograms/mg protein (28 mM). This level of myo-inositol is sufficient for this compound to function as an osmoregulator in primary astrocytes and it is likely to contribute to the maintenance of brain volume.

Animals↗
PubMed1994

Calcium dependence of hypoosmotically induced potassium release in cultured astrocytes.

A major mechanism in cell volume regulation after hypoosmotic stress is K+ release. Our studies show that in astrocytes, K+ release during hypoosmotic stress is a Ca(2+)-dependent process. Agents that increase intracellular Ca2+, such as ionomycin and ouabain, potentiated hypoosmotically stimulated K+ release, while compounds that block Ca2+ entry during hypoosmotic stress, such as nimodipine, bepridil, and MK-801, inhibited hypoosmotically stimulated K+ release. Similarly, chelation of intracellular Ca2+ blocked hypoosmotically induced K+ release. Caffeine and U-73122 also inhibited K+ efflux under hypoosmotic conditions, suggesting that intracellular Ca2+ release from Ca(2+)-induced Ca2+ release stores and inositol trisphosphate-sensitive intracellular Ca2+ stores play a role in the mechanism of K+ release. Blocking the activity of calmodulin, and of CaM kinase, attenuated hypoosmotically induced K+ release. Our findings indicate that entry of extracellular Ca2+ and Ca2+ release from intracellular stores play a key role in the activation of K+ release under hypoosmotic conditions and thus in cell volume regulation.

Animals↗
PubMed1993

Effect of external acidosis on basal and ATP-evoked calcium influx in cultured astrocytes.

The effect of lactic acidosis on calcium influx, accumulation and efflux was studied in primary cultures of neonatal cortical rat astrocytes. Treatment of cultures with 20 mM sodium lactate, pH 6.0, for 10-60 min resulted in a 35% reduction of 45Ca2+ influx. The decrease in calcium influx was pH dependent because a similar reduction was observed in cultures exposed to pH 6.0 without lactate, while no difference was observed in cultures treated with sodium lactate at pH 7.4. Calcium accumulation was also decreased by lactic acidosis (20% reduction), while calcium efflux was unaffected. Studies with lanthanum, an inhibitor of calcium transport, indicated that the effect of lactic acidosis was not due to non-specific leakage of calcium. The reduction in calcium influx was reversible, thereby indicating that the cells were not permanently damaged by lactic acidosis. In addition to basal calcium influx, stimulated influx (mediated by extracellular ATP, 100 microM) was also reduced by 20 mM sodium lactate, pH 6. These findings suggest that protonization of calcium channels or other calcium entry pathways leads to a reduction in calcium influx in astrocytes. This diminished calcium entry, by affecting calcium-dependent mechanisms necessary for such processes as volume regulation, glycogen metabolism, or regulation of ionic permeability, may alter the ability of astrocytes to elicit appropriate responses following CNS injury.

Adenosine Triphosphate↗
PubMed1993

Role of phosphoinositide hydrolysis in astrocyte volume regulation.

Astrocytes exposed to hypoosmotic stress swell and subsequently reduce their size to almost their original volume, a phenomenon called regulatory volume decrease (RVD). We found that during hypoosmotic swelling there was a twofold increase in phosphatidylinositol (PI) hydrolysis. This increase was inhibited by the phospholipase C inhibitor, U-73122 (10 microM). Inhibition of PI hydrolysis resulted in blockage of RVD. We also examined whether agents that stimulate PI hydrolysis would enhance RVD. These agents significantly accelerated RVD. The rank order of potency was endothelin (20 nM) > or = norepinephrine (100 microM) > endothelin-3 (7 nM) > thrombin (1 U/ml) > or = ATP (500 microM) > bradykinin (20 microM) > or = carbachol (500 microM), as indicated by RVD rate constants. The extent of PI hydrolysis induced by these agents at the beginning of RVD exhibited a logarithmic relationship with the magnitude of RVD enhancement. Also, there was a linear relationship between the rate of PI hydrolysis and RVD rate constants. Our results suggest that stimulated PI hydrolysis is involved in the regulation of cell volume in astrocytes.

Animals↗
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↗
PubMed1989

Potassium-stimulated calcium uptake in astrocytes and its potent inhibition by nimodipine.

Elevation of the extracellular potassium concentration above its "resting" level of 5.4 mM stimulated uptake of 45Ca2+ in primary cultures of astrocytes. This effect was only observed when cells were exposed to excess potassium shortly after their exposure to 45Ca2+ and was potently inhibited (IC50 congruent to 3 nM) by the calcium channel blocker nimodipine. In contrast, nimodipine exerted little effect on unstimulated basal uptake of 45Ca2+. These findings suggest that the therapeutic benefit of calcium channel blockers in epilepsy may result in part from the ability of these drugs to prevent calcium entry into astrocytes during seizures when the extracellular potassium is elevated four- to fivefold above normal.

Animals↗
PubMed1989

Beta-DL-methylene-aspartate, an inhibitor of aspartate aminotransferase, potently inhibits L-glutamate uptake into astrocytes.

[3H]Glutamate uptake into astrocytes in primary culture was potently inhibited by the aspartate analogues L- and D-aspartic acid, DL-threo-beta-hydroxy-aspartic acid-beta-hydroxymate (IC50's: 136, 259, 168, and 560 microM, respectively) and by beta-DL-methylene-aspartate, a suicide inhibitor of aspartate aminotransferase (IC50: 524 microM), and by the endogenous sulphur-containing amino acid L-cysteinesulfinic acid (IC50: 114 microM), [3H]Glutamate uptake was not significantly affected by either N-methyl-D-aspartate or DL-homocysteine thiolactone. These results demonstrate that other excitatory amino acids including aspartate and L-cysteinesulfinic acid (but excluding L-homocysteic acid) interact with the glutamate transport system of astrocytes. Inhibition of glutamate uptake may significantly increase the level of neuronal excitability.

Amino Acid Transport System X-AG↗
PubMed1988

Evidence for involvement of the astrocytic benzodiazepine receptor in the mechanism of action of convulsant and anticonvulsant drugs.

The anticonvulsant drugs carbamazepine, phenobarbital, trimethadione, valproic acid and ethosuximide at pharmacologically relevant concentrations inhibit [3H]diazepam binding to astrocytes in primary cultures but have much less effect on a corresponding preparation of neurons. Phenytoin as well as pentobarbital (which is not used chronically as an anticonvulsant) are equipotent in the two cell types. The convulsants picrotoxinin and pentylenetetrazol, the convulsant benzodiazepine RO 5-3663 and the two convulsant barbiturates DMBB and CHEB similarly inhibit diazepam binding to astrocytes but have little effect on neurons. On the basis of these findings it is suggested that these convulsants and anticonvulsants owe at least part of their effect to an interaction with the astrocytic benzodiazepine receptor, perhaps by interference with a calcium channel.

Animals↗
PubMed1987

Inhibition of [3H]diazepam binding in primary cultures of astrocytes by atrial natriuretic peptide and by a cyclic GMP analog.

Atrial natriuretic peptide (ANP) was found potently (IC50 = 88 nM) to inhibit [3H]diazepam binding to astrocytes in primary cultures but not to a corresponding preparation of neurons. Based upon this finding and literature data demonstrating a pronounced correlation between the distribution of 'peripheral-type' benzodiazepine receptors and ANP receptors, it is suggested that ANP may be an allosteric regulator of the astrocytic benzodiazepine binding site. Since ANP is known to increase the level of cGMP it was also investigated whether 8-bromoguanosine 3':5' cyclic monophosphate, a permeable cGMP analog, displaced diazepam binding. This was the case, suggesting that cGMP might be involved in the ANP effect on the astrocytic benzodiazepine receptor.

Animals↗
PubMed1987

Dissimilarities between benzodiazepine-binding sites and adenosine uptake sites in astrocytes and neurons in primary cultures.

The question whether the benzodiazepine receptor site in astrocytes or in neurons might be identical to the adenosine uptake site was studied by determining pharmacological profiles, inhibition types, and the effects of benzodiazepine antagonists in primary cultures of either astrocytes or neurons. Fourteen different benzodiazepines and five different adenosine uptake inhibitors displaced [3H] diazepam and inhibited adenosine uptake in both astrocytes and neurons. However, the rank orders (determined as IC50 values) with which these two parameters were affected were profoundly different, indicating dissimilarities between these two sites. For several of the compounds a difference in inhibition type (competitive vs. noncompetitive) was observed between the benzodiazepine-binding site and the adenosine uptake site in astrocytes and/or neurons, which further corroborated the conclusion of a difference between the benzodiazepine-binding site and the adenosine uptake site. Finally, the neuronal benzodiazepine antagonists RO 15-1788 and CGS-8216 and the astrocytic benzodiazepine antagonist PK 11195, which reverse the action of benzodiazepines, were not able to reverse inhibition of adenosine uptake by diazepam but exerted an inhibitory effect of their own.

Adenosine↗
PubMed1987

Pharmacological characteristics of diazepam receptors in neurons and astrocytes in primary cultures.

Benzodiazepine binding sites in mouse astrocytes and neurons in primary cultures were labeled with [3H]diazepam (1.8 nM), and their inhibition by 14 different benzodiazepines and 3 benzodiazepine antagonists was studied. RO 5-4864, RO 7-3351, and, especially, the antagonist PK 11195 were much more potent in astrocytes than in neurons, whereas the opposite was true for the agonists alprazolam, clonazepam, flurazepam, RO 11-3128, and chlordiazepoxide, and, especially, the antagonists CGS-8216 and RO 15-1788. Flunitrazepam, diazepam, midazolam, RO 11-6893, and RO 5-2181 were about equipotent in the two cell types. The neuronal, but not the astrocytic, binding site showed stereospecificity. In astrocytes most of the drugs had pseudo-Hill coefficients close to one, whereas the pseudo-Hill coefficients in neurons, except for RO 5-4864 and PK 11195, were distinctly lower than one. Thus, the benzodiazepine binding sites had profoundly different pharmacological characteristics in neurons and in astrocytes.

Animals↗
PubMed1986

Similarities of adenosine uptake systems in astrocytes and neurons in primary cultures.

Uptake of extracellular adenosine was studied in primary cultures of astrocytes or neurons. Both cell types showed a high affinity uptake. The Km values were not significantly different (6.5 +/- 3.75 microM in astrocytes and 6.1 +/- 1.86 microM in neurons), but the intensity of the uptake was higher in astrocytes than in neurons (Vmax values of 0.16 +/- 0.030 and 0.105 +/- 0.010 nmol x min-1 x mg-1 protein, respectively). The temperature sensitivity was similar in the two cell types. Adenosine uptake inhibitors and benzodiazepines inhibited the adenosine uptake systems in both astrocytes and neurons with IC50 values in the high nanomolar or the micromolar range and the rank order of potency was similar in the two cell types. In both cell types the (-) isomers of two sets of benzodiazepine stereoisomers were more potent than the (+) isomers. Dixon analysis showed that dipyridamole, papaverine, hexobendine and chlordiazepoxide inhibited the adenosine uptake competitively and clonazepam noncompetitively in both cell types.

Adenosine↗
PubMed1985

Binding of [3H]Ro 5-4864 in primary cultures of astrocytes.

Intact primary cultures of astrocytes display benzodiazepine receptors, which can be labeled with [3H]Ro 5-4864. Binding of [3H]Ro 5-4864 is specific, saturable and temperature dependent, being maximal at 0 degrees C. Scatchard analyses show a single population of high affinity binding sites with a Kd value of 6.7 nM and a Bmax value of congruent to 12,000 fmol/mg protein. The binding reaches equilibrium at congruent to 100 min, with k+1 of 0.0078 nM-1 X min-1 and is rapidly reversible with k-1 of 0.057 min-1. [3H]Ro 5-4864 binding is not modulated by GABA. Certain benzodiazepines (flunitrazepam, diazepam, Ro 7-3351) and dipyridamole displace this binding with IC50 values in the nanomolar range, whereas other benzodiazepines (alprazolam, clonazepam, chlordiazepoxide) as well as carbamazepine, phenytoin and phenobarbital have IC50 values in the micromolar range. These characteristics resemble those of Ro 5-4864 binding to brain membrane preparations reported by other authors and thus indicate that the cultured astrocytes are good models of their in vivo counterparts.

Animals↗