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Biomedical subjects

A S Bender

Publications and source records attributed to A S Bender.

At least 55 records · Page 3Linked to original sources
PubMed1985

Estradiol and progesterone potentiate adenosine's depressant action on rat cerebral cortical neurons.

The effects of 17 beta-estradiol and progesterone on the uptake of adenosine by rat cerebral cortical synaptosomes have been correlated with the ability of these gonadal steroids to potentiate the depressant actions of adenosine on the spontaneous firing of rat cerebral cortical neurons. 17 beta-estradiol hemisuccinate competitively inhibited adenosine uptake with a Ki of 0.5 microM (Lineweaver-Burk plot) or 0.78 microM (Dixon plot). The Ki for progesterone was 0.34 microM (Lineweaver-Burk plot) or 0.36 microM (Dixon plot). When applied by iontophoresis, both steroids potentiated the depressant effects of adenosine on the firing of rat cerebral cortical neurons. Potentiation of the effect of endogenously released adenosine would account for the central depressant actions of these steroids.

Action Potentials↗
PubMed1984

Flunitrazepam binding to intact and homogenized astrocytes and neurons in primary cultures.

[3H]Flunitrazepam binds to intact and homogenized mouse astrocytes and neurons in primary cultures. In intact cells, the binding is to a single, high-affinity, saturable population of benzodiazepine binding sites with a KD of 7 nM and Bmax of 6,033 fmol/mg protein in astrocytic cells and a KD of 5 nM and Bmax of 924 fmol/mg protein in neurons. After homogenization, the Bmax values decrease drastically in both cell types, but most in astrocytes. The temperature and time dependency are different for the two cell types, with a faster association and dissociation in astrocytes than in neurons and a greater temperature sensitivity in the astrocytes. Moreover, flunitrazepam binding sites on neuronal and astrocytic cells have different pharmacological profiles. In intact astrocytic cells, Ro 5-4864 (Ki = 4 nM) is the most potent displacing compound, followed by diazepam (Ki = 6 nM) and clonazepam (Ki = 600 nM). In intact neurons, the relative order of potency of these three compounds is different: diazepam (Ki = 7 nM) is the most potent, followed by clonazepam (Ki = 26 nM) and Ro 5-4864, which has little effect. After homogenization the potency of diazepam decreases. We conclude that both neuronal and astrocytic cells possess high-affinity [3H]flunitrazepam binding sites. The pharmacological profile and kinetic characteristics differ between the two cell types and are further altered by homogenization.

Animals↗
PubMed1983

Benzodiazepines and beta-adrenergic binding to primary cultures of astrocytes and neurons.

Characteristics of primary cultures of neurons or of astrocytes are discussed which suggest that such cultures are appropriate models for their in vivo counterparts. Advantages or disadvantages of these cultures for studies of receptor binding are: Neuronal and astrocytic binding can be studied separately. Binding can be studied to intact cells or after homogenization. Apparent binding to intact cells may include unspecific retention. This seems to be a problem for beta-adrenergic ligands but not for benzodiazepines.

Animals↗
PubMed1981

The rapid uptake and release of [3H]adenosine by rat cerebral cortical synaptosomes.

Adenosine, a putative inhibitory transmitter or modulator in the brain, is rapidly transported by rat cerebral cortical synaptosomes. The uptake may represent a facilitated diffusion process, which is saturable and temperature-dependent. In this study, the uptake process was very rapid, reaching completion within 60 s of incubation at 37 degrees C, and had an apparent Km value of 0.9 microM and a Vmax value of 5.26 pmol/mg protein/30 s. Over 70% of the adenosine taken up remained unchanged, whereas 14% was metabolized to inosine. Twelve percent of the adenosine was converted to nucleotides. Rapid uptake of adenosine into rat cerebral cortical synaptosomes was partially inhibited by replacing Na+ with choline chloride in the medium. Ca2+ ion is important for the uptake process, as inhibition of adenosine uptake occurs in the presence of either Co2+ or EGTA. Rapid uptake of adenosine is apparently mediated by a nucleoside carrier, a conclusion based on its inhibition by a variety of purine and pyrimidine nucleosides. Uptake was inhibited by dipyridamole, hexobendine, papaverine, flurazepam, and morphine. Over 60% of the adenosine taken up by the rapid uptake system (30 s) was released by depolarizing agents. In contrast, only 30% of the adenosine taken up during a 15-min incubation period was released under the same conditions. [3H]Adenosine was the predominant purine released in the presence or absence of depolarizing agents. The basal and KCl-evoked release mechanisms were found to be at least partially Ca2+-dependent, however, the release of adenosine by veratridine was increased in the presence of EGTA. This finding is in agreement with the reported Ca2+-independent release of ATP from brain synaptosomes. The present findings suggest that there are at least two functional pools of adenosine in synaptosomes. Adenosine taken up by different uptake systems may be destined for different uses (metabolism or release) in the neuron.

Adenosine↗
PubMed1981

Some biochemical properties of the rapid adenosine uptake system in rat brain synaptosomes.

The rapid uptake of adenosine into rat brain cortical synaptosomes is mediated by a facilitated diffusion process. The carrier mediated uptake is sensitive to pH and temperature. The average Q10 value for the system is approximately 1.77 and the necessary activation energy (Ea) is estimated to be 8870 cal/mol. These values are essentially in agreement with values reported for adenosine uptake carriers of other tissues. Substrate specificity of the uptake system in the CNS demonstrates that nucleotides do not interact with the carrier until they have been hydrolyzed to nucleosides. Structural modification of the purine moiety at the "2" position did not have a profound effect on the ability of the molecule to serve as a substrate for the uptake system. Competitive inhibition by sulfhydryl reagents, p-chloromercuribenzoate, and N-ethylmaleimide on adenosine uptake suggests a direct involvement of sulfhydryl group(s) in the uptake mechanism. Other purines and pyrimidines also inhibit adenosine uptake, suggesting that a variety of nucleosides can interact with a common carrier system.

Adenine Nucleotides↗
PubMed1980

The characterization of [3H] adenosine uptake into rat cerebral cortical synaptosomes.

Uptake of adenosine, a putative inhibitory transmitter or modulator, was investigated in rat cerebral cortical synaptosomes. The accumulation of [3H] adenosine into synaptosomes, using an adenosine concentration of 10 microM, was linear for 30 min at 30 degrees C. The uptake appeared to be mediated by kinetically saturable processes with apparent Km's of 1 microM ("high-affinity A") and 5 microM ("high-affinity B"), both of which were partially sensitive to the presence of external sodium and calcium ions. Both uptake processes were partially inhibited by 2,4-dinitrophenol, implying the presence of active uptake and diffusional components. A study of the metabolites of adenosine taken up by the two uptake systems indicates that the major metabolites were adenosine and nucleotides. However, adenosine incorporated by the high-affinity A uptake system is more likely to form deaminated metabolites, such as hypoxanthine and inosine, indicating a possible functional difference between the two uptake processes. A detailed comparison of the inhibitory properties of certain adenosine analogues and other pharmacological agents has revealed differences between the two adenosine uptake systems. Since the glial contamination in synaptosomal preparations is well established, one of the uptake systems we observed in the present study might be of glial origin. This notion is supported by the findings that the Km values and kinetic properties of papaverine action in he synaptosomal high-affinity A uptake system are similar to those of astrocytes reported in the literature. In conclusion, the uptake processes of synaptosomal preparations show that accumulation of adenosine into neuronal (and possibly glial) elements may play a major role in regulating the extracellular adenosine concentration. Uptake inhibitors, such as diazepam, may exert, at least in part, their pharmacological actions by interfering with the regulation of extracellular adenosine concentrations.

Adenosine↗
PubMedDate not supplied

Elevation of glutathione levels by ammonium ions in primary cultures of rat astrocytes.

It is well established that ammonia is detoxified in the brain to form glutamine and that astrocytes play a major role in this process. The synthesis of glutamine requires glutamate and ATP. Since glutamate and ATP are also required for the synthesis of glutathione (GSH), we examined the effect of pathophysiological concentrations of ammonia on levels of GSH in primary cultures of astrocytes. GSH content in the medium increased in a dose- and time-dependent manner in the presence of ammonia. After an initial decrease, cellular GSH content increased in a similar manner. The levels of glutathione disulfide (GSSG) were also increased. A linear relationship was observed between ammonia concentration and the increase in GSH levels. An increase in the efflux of GSH from cells into medium was also observed under these conditions. Buthionine sulfoximine and acivicin, but not methionine sulfoximine, blocked the ammonia induced increase in GSH levels. No, or minor, changes in the activities of enzymes (gamma-glutamyl transpeptidase, GSH reductase and GSH-peroxidase) that might influence GSH levels were identified and thus could not account for the ammonia induced increase in GSH levels in astrocytes. These findings indicate that pathophysiological concentrations of ammonium ions result in increased astroglial levels of GSH which may affect the metabolism and function of astrocytes.

Ammonia↗
PubMedDate not supplied

Characterization of cystine uptake in cultured astrocytes.

Glutathione is involved in the maintenance of the structural and functional integrity of membrane proteins, in protection against free radicals and oxidative stress, and in the detoxification of xenobiotics. The cellular uptake of cystine is the rate limiting step in the biosynthesis of glutathione. The precise mechanism for such uptake is not clear as some reports indicate that the uptake occurs through a glutamate-cystine antiporter (system X(c)(-)), whereas, others suggest that it is taken up by the glutamate transporter (system X(AG)). Our studies in cultured astrocytes derived from neonatal rats showed that glutamate, D- and L-aspartate inhibited cystine uptake; that factors that increased intracellular glutamate levels, which would have enhanced the activity of the antiporter, did not stimulate cystine uptake; that the uptake was sodium dependent and partially chloride dependent; that the b(o,+) and ASC systems, which have been shown to carry cystine in some cells, did not mediate cystine uptake in astrocytes; that glutamate uptake blockers such as L-aspartate-beta-hydroxamate (AbetaH) and L-trans-pyrrolidine-2,4-dicarboxylate (PDC), as well as cystine uptake inhibitor L-alpha-aminoadipate (AAA) potently reduced cystine uptake. Additionally, deferoxamine (100 microM) as well as ammonium chloride (5 mM), both of which inhibit glutamate uptake, also inhibited cystine uptake. Taken together, our findings indicate that astrocytes take up cystine through a similar, if not identical, system used to take up glutamate. Interference of cystine uptake by astrocytes through the glutamate transport system may have profound effects on the redox state and the structural and functional integrity of the CNS.

Amino Acids↗