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S Ohkuma

Publications and source records attributed to S Ohkuma.

At least 91 records · Page 5Linked to original sources
PubMed1995

Role of nitric oxide in central synaptic transmission: effects on neurotransmitter release.

Nitric oxide (NO), an unstable radical formed via oxidative deamination of L-arginine by NO synthase, has the activity to evoke release of several neurotransmitters including acetylcholine, catecholamines and neuroactive amino acids. N-methyl-D-aspartate (NMDA) receptor stimulation also causes neurotransmitter release through NO formation, which is supported by the data extracellular hemoglobin completely abolishes the stimulatory effect of NMDA on neurotransmitter release. In addition, NO formed by NMDA receptor activation exhibits its stimulatory action on neurotransmitter release extracellularly. A product formed by the reaction of NO and superoxide, peroxynitrite, is also considered to be partly involved in NO-evoked neurotransmitter release. The removal of Ca2+ or Na+ significantly reduces the release of GABA evoked by an NO generator, S-nitroso-N-acetylpenicillamine (SNAP), and simultaneous withdrawal of these ions completely abolishes the SNAP-induced release of GABA. Either in the presence or absence of Ca2+, GABA transport inhibitors such as nipecotic acid dose-dependently suppress the SNAP-induced GABA release in the presence of Na+. These results indicate that NO-evoked neurotransmitter release is mediated by two distinct release systems, a Ca2(+)-dependent system and the reverse process of a Na(+)-dependent carrier-mediated transport system.

Acetylcholine↗
PubMed1995

Increase of diazepam binding inhibitor mRNA levels in the brains of chronically ethanol-treated and -withdrawn mice.

Although the molecular mechanisms underlying behavioral responses observed in ethanol (EtOH)-dependent and -withdrawn animals are poorly understood, several lines of evidence have suggested that some of the pharmacological actions of EtOH are mediated via the potentiation of gamma-aminobutyric acid (GABA)ergic transmission. In the present study, the effects of acute and chronic EtOH treatment on the expression of diazepam binding inhibitor (DBI) mRNA in the mouse brain and the functional involvement of the benzodiazepine (BDZ) receptor in EtOH-induced alteration of the DBI mRNA expression were examined. The expression of cerebral DBI mRNA significantly increased in EtOH-treated (130% of control) and EtOH-withdrawn (220% of control) mice in comparison with untreated mice, whereas the DBI mRNA level was not altered after a single administration of EtOH (3 g/kg i.p.). The increase in the DBI mRNA expression in the mouse cerebral cortex after EtOH withdrawal diminished over 14 days despite the disappearance of withdrawal signs within 2 days after the withdrawal of EtOH. Simultaneous administration of flunitrazepam (10 mg/kg i.p.) with EtOH completely abolished the EtOH-induced increase in DBI mRNA expression. On the other hand, the level of beta-actin mRNA was not affected by similar EtOH treatment. These results indicate that changes in the expression of cerebral DBI mRNA induced by long-term EtOH treatment may be involved in the establishment of alcohol dependence, and such changes may be also regulated by BDZ receptors.

Animals↗
PubMed1995

[Metabolic pathway of the degradation of macromolecules by lysosomal enzymes].

The lysosomes of most cells function principally in intracellular digestion. They contain several dozens of enzymes, mostly acid hydrolyases. Vacuolar-ATPase on lysosomal membrane establishes low-pH environment required for efficient expression of hydrolyase activity. Of several dozen disorders of human and various animals known to be due to lysosomal dysfunctions, most of the lysosomal storage diseases are of genetic origin. Metabolic pathways of intracellular macromolecules within lysosomes have been established from studies on the molecular and biochemical defects on lysosomal enzymes. In the lysosomal storage diseases most of all, the metabolic pathways of breakdown of lipids, glycosaminoglycans or oligosaccharides are severely affected. The degradation pathways of glycoproteins, proteoglycans, and glycolipids by lysosomal enzymes are described in this brief review.

Animals↗
PubMed1994

Mechanisms for pancreatic hypertrophy induced by long-term administration of bethanechol.

Mechanisms for the hypertrophy of rat pancreas induced by long-term administration of bethanechol were investigated. The administration of bethanechol, an acetylcholine receptor agonist, to male Wistar rats for 14 days induced significant increases in the pancreatic weight and contents of protein, amylase and RNA in the pancreas without altering the content of DNA and the incorporation of [3H]thymidine into DNA. Simultaneous administration of atropine with bethanechol suppressed the bethanechol-induced pancreatic hypertrophy. Long-term administration of other acetylcholine receptor agonists also showed similar effects as produced by bethanechol. CR1505 (loxiglumide; D,L-4-(3,4-dichlorobenzoyl-amino)-5-(N-3-methoxypropyl-pentylam ino)-5- oxopentanoic acid), an antagonist of cholecystokinin receptors, inhibited pancreatic growth induced by long-term administation of pentagastrin, whereas pancreatic hypertrophy induced by bethanechol was not inhibited by CR1505. These results suggest that long-term administration of bethanechol induces pancreatic hypertrophy through direct activation of muscarinic receptors in the pancreas.

Amylases↗
PubMed1994

Nicotine increases diazepam binding inhibitor (DBI) mRNA in primary cultured neurons.

The effect of nicotine on the expression of diazepam binding inhibitor (DBI) mRNA in primary cultured cerebral cortical neurons was examined using Northern blot analysis. Nicotine exposure (0.001-10 microM) for 24 h increased the DBI mRNA level in a dose-dependent manner, whereas the beta-actin mRNA level showed no change. This effect of nicotine was faded out over 48 h of its exposure. Hexamethonium (100 microM) completely abolished the nicotine-induced increase in DBI mRNA expression. These results indicate that nicotine increases the expression of DBI mRNA in cerebral cortical neurons via the activation of nicotinic acetylcholine receptor.

Animals↗
PubMed1994

Cytosol treated with GTP gamma S disintegrates lysosomes in vitro.

Cytosol that was treated with guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) disintegrated lysosomes in a dose dependent manner, as detected by the release of preloaded fluorescein isothiocyanate-dextran from lysosomes. This phenomenon is a consequence of the two successive steps of incubation, (i) the treatment of cytosol with GTP gamma S, and (ii) the disintegration of lysosomes by the treated cytosol. The first step was specific for GTP gamma S and the effect of GTP gamma S was suppressed by GTP or GDP. The second step required ATP but it was not affected by bafilomycin A1. GTP gamma S treatment of lysosomes themselves did not induce their lysis. The lytic "factor(s)" produced in GTP gamma S-treated cytosol consisted of high molecular weight protein(s).

Adenosine Triphosphate↗
PubMed1994

Induction of phagocytic activity of M1 cells by an inhibitor of vacuolar H+-ATPase, bafilomycin A1.

Bafilomycin A1, a selective inhibitor of vacuolar H+-ATPase, time- and dose-dependently induced the differentiation of M1 cells, a murine myeloid leukemic cell line, into macrophage-like cells as revealed by the phagocytosis of polystyrene latex particles. This differentiation was inhibited not only by actinomycin D and cycloheximide but also by ST-638 (an inhibitor of tyrosine kinase). However, it was affected neither by K-252a (an inhibitor of C-kinase) nor by H-89 (an inhibitor of A-kinase), in contrast to the M1 cell differentiation induced by leukemia inhibitory factor (LIF). Okadaic acid inhibited both the bafilomycin A1-induced and LIF-induced differentiation of M1 cells.

Animals↗
PubMed1994

Inhibition of NGF-induced neurite outgrowth of PC12 cells by Bacillus cereus sphingomyelinase, a bacterial hemolysin.

Sphingomyelinase of Bacillus cereus, a bacterial hemolysin, reduced nerve growth factor (NGF)-induced neurite outgrowth of PC12 cells in a dose-dependent manner. At 200 mU/ml, sphingomyelinase repressed half the neurite outgrowth of the cells at 250 ng/ml NGF. The c-fos superinduction, one of the early responses induced by NGF, was not influenced by this treatment, suggesting that the repression by sphingomyelinase occurred via a protein kinase C-independent pathway.

Animals↗
PubMed1994

Presence of N-methyl-D-aspartate (NMDA) receptors in neuroblastoma x glioma hybrid NG108-15 cells-analysis using [45Ca2+]influx and [3H]MK-801 binding as functional measures.

In the present study, we have attempted to clarify whether neuroblastoma glioma hybrid NG 108-15 cells (NG cells) possess the NMDA receptor complex using [45Ca2+]influx and [3H]MK-801 binding as functional measures. Glutamate and NMDA dose-dependently increased [45Ca2+]influx and these increases were further enhanced by glycine. Scatchard analysis revealed the presence of a high-affinity binding site for [3H]MK-801 with a KD of 18.8 nM and a Bmax of 0.328 pmol/mg protein. This [3H]MK-801 binding was also increased by NMDA in a dose-dependent manner and this increase was further enhanced by glycine. Both ketamine and MK-801 inhibited glutamate- and NMDA-induced [45Ca2+]influx as well as the increase of [3H]MK-801 binding in a dose-dependent manner. Similarly, Mg2+ and Zn2+ dose-dependently reduced both glutamate-induced [45Ca2+]influx and [3H]MK-801 binding. Spermine, one of the polyamines, showed a biphasic stimulatory effects on glutamate-induced [45Ca2+]influx and [3H]MK-801 binding. These results indicate that NG cells possess a pharmacologically distinct NMDA receptor complex and suggest that these cells may be useful for the analyses on pharmacological and biochemical characteristics of the NMDA receptor complex.

Calcium Radioisotopes↗
PubMed1994

GABAA receptor stimulation enhances NMDA-induced Ca2+ influx in mouse cerebral cortical neurons in primary culture.

The effect of GABAA receptor stimulation on N-methyl-D-aspartate(NMDA)-induced [45Ca2+]influx has been examined using primary cultured cerebral cortical neurons. NMDA induced a dose-dependent increase in [45Ca2+]influx, which was blocked by MK-801 in a dose-dependent manner. GABAA receptor agonists significantly enhanced the NMDA-induced [45Ca2+]influx, and this enhancement was dose-dependently inhibited by bicuculline, although picrotoxin and tert-butyl-bicyclo[2.2.2]phosphoro-thionate (TBPS) exhibited no alterations in this stimulatory action of GABAA receptor agonists. Blockers of L-type voltage-dependent calcium channels significantly reduced the NMDA-induced [45Ca2+]influx. The increased [45Ca2+]influx by both NMDA and GABAA receptor agonists was also reduced by verapamil and nifedipine. These results suggest that the enhancement of NMDA-induced [45Ca2+]influx by GABAA receptor stimulation in immature cerebral cortical neurons may be due to the increased opening of voltage-dependent calcium channel by synergestic actions between NMDA and GABAA receptors.

Animals↗
PubMed1994

Ethanol-induced alteration in activities of cerebral phosphatidylinositol 4,5-biphosphate-specific and cytosolic phospholipase C in the brain: analysis using NG 108-15 cells and brains from ethanol-inhaled mice.

Effect of long-term exposure to ethanol (EtOH) on the phosphatidylinositol 4,5-biphosphate (PIP2)-specific and cytosolic phospholipase C (PLC) activities in neuroblastoma x glioma hybrid (NG 108-15) cells and the brains from EtOH-inhaled mice were investigated. Long-term (2 days) exposure of NG 108-15 cells to EtOH induced significant decrease in PIP2-specific PLC activity dependent on concentration and duration of exposure, although the presence of EtOH in the enzyme assay system induced no alteration in PIP2-specific PLC activity. On the other hand, cytosolic PLC activity in NG 108-15 cells significantly increased by both the long-term exposure of the cells to EtOH and the addition of EtOH into the assay system. These changes in activities of both types of PLC in NG 108-15 cells observed after EtOH exposure recovered rapidly by the removal of EtOH. Moreover, the changes in activities of PIP2-specific and cytosolic PLC in the brain of EtOH-inhaled mice were similar to those found in NG 108-15 cells. These results indicate that EtOH inhibits the activity of PIP2-specific PLC and activates cytosolic PLC in the brain. These changes in cerebral PLC activities are suggested to involve in central action of EtOH and establishment of alcohol dependence.

Administration, Inhalation↗
PubMed1994

Muscimol prevents neuronal injury induced by NMDA.

The effect of muscimol on N-methyl-D-aspartate (NMDA)-induced injury of primary cultured cerebral cortical neurons was examined. NMDA induced a dose-dependent leakage of LDH activity, which was significantly inhibited by (+-)-5-methyl-10,11-dihydro-5H-dibenzo-[a,d]cyclopentan-5,10-imine (MK-801). Muscimol significantly reduced the NMDA-induced increase of lactic dehydrogenase (LDH) leakage, and bicuculline abolished this protective effect of muscimol. Similarly, muscimol reduced the NMDA-induced increase in trypan blue staining of the cells, and bicuculline suppressed this inhibitory action of muscimol. These results suggest that GABAA-receptor stimulation exerts a protective action against the neuronal injury induced by NMDA-receptor activation.

Animals↗
PubMed1993

Immunological detection of F1-ATPase gamma-subunit in rat liver peroxisomes.

The gamma-subunit of F1-ATPase was detected in highly purified rat liver peroxisomes by immunoblot analysis using an antiserum against purified mitochondrial F1-ATPase. Other subunits of F1-ATPase such as alpha, beta and delta were not detected in the peroxisomes. The polypeptide corresponding to the gamma-subunit in peroxisomes was also recognized by affinity purified antibody against F1-ATPase gamma-subunit. The amount of gamma-subunit in liver homogenate increased in parallel with proliferation of peroxisomes after administration of clofibrate in rats, whereas the amounts of alpha, beta and delta-subunits were not increased. These results suggest that the gamma-subunit of F1-ATPase is a constituent of peroxisomes in rat liver.

Animals↗
PubMed1993

Isolation and influenza virus receptor activity of glycophorins B, C and D from human erythrocyte membranes.

(1) Glycophorins (GPs) AM, AN, B, C and D were each isolated into a high state of purity from human erythrocyte membranes by a combination of lithium diiodosalicylate (LIS)-phenol extraction, gel-filtration with Bio-Gel A1.5m and HPLC with LiChrospher 1000 TMAE. (2) GPs-B, -C and -D reacted with influenza A and B viruses as well as GPs-AM and -AN and the order of reactivities against two viruses of the glycophorins was as follows: GP-B > GP-C > GP-AM = GP-AN >> GP-D for the former virus and GP-C > GP-B > GP-AM = GP-AN >> GP-D for the latter virus.

Amino Acids↗
PubMed1993

Purification and characterization of lysosomal H(+)-ATPase. An anion-sensitive v-type H(+)-ATPase from rat liver lysosomes.

Lysosomal H(+)-ATPase was purified to homogeneity from rat liver lysosomes. It is a bafilomycin A1-sensitive Mg(2+)-ATPase, which reacts with antibodies against the 16- and 70-kDa subunits of vacuolar H(+)-ATPase (Nezu, J., Motojima, K., Tamura, H., and Ohkuma, S. (1992) J. Biochem. (Tokyo) 112, 212-219), and has been separated from both the N-ethylmaleimide (NEM)-sensitive/bafilomycin A1-insensitive Mg(2+)-ATPase (ATPase I) and the NEM-insensitive Mg2+/Ca(2+)-ATPase (ATPase II) (Hayashi, H., Arai, K., Sato, O., Shimaya, A., Sai, Y., and Ohkuma, S. (1992) Chem. Pharm. Bull. 40, 2783-2786). The purified enzyme had the subunit structure of vacuolar H(+)-ATPase, consisting of 110-, 70-, 56-, 42-, 39-, 34-, (32-,) and 16-kDa proteins. It had optimal activity at a pH of 7.0-8.0, with an apparently single Km value for ATP of 95 microM. It hydrolyzed ATP > or = dATP >> GTP, ITP >> UTP, but not CTP, and was inhibited by ADP. It demonstrated divalent cation specificity in the order of Mg2+, Mn2+ > Fe2+, Co2+ > Ca2+. Among various anions, Cl-, Br-, and F- activated ATPase activity, whereas NO3- inhibited activity. It was inhibited not only by bafilomycin A1 but also by NEM, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole, quercetin, and dicyclohexylcarbodi-imide. The purified enzyme was incorporated into proteoliposomes where its proton pump activity was reconstituted. This suggested that the isolated enzyme maintains its H+ translocation activity. These findings suggest that the isolated enzyme is an anion-sensitive vacuolar type H(+)-ATPase that functions as a lysosomal proton pump.

Animals↗
PubMed1993

Inhibition of cell growth by bafilomycin A1, a selective inhibitor of vacuolar H(+)-ATPase.

Bafilomycin A1, a potent selective inhibitor of vacuolar H(+)-ATPase, inhibited the growth of a variety of cultured cells dose-dependently, including golden hamster embryo and NIH-3T3 fibroblasts, whether or not they were transformed, and PC12 and HeLa cells. The concentration of bafilomycin A1 for 50% inhibition of cell growth ranged from 10 to 50 nM. The dose response was nearly parallel with that of the bafilomycin A1-induced lysosomal pH increase. The degree of pH increase for growth inhibition produced by bafilomycin A1 was similar to that produced by NH4Cl in which little difference was recognized in effect among cell types.

3T3 Cells↗
PubMed1993

Inorganic pyrophosphatase of clofibrate-induced rat liver peroxisomes.

Clofibrate increased inorganic pyrophosphatase (PPase) activity in peroxisomes more than 12-fold (740 milliunits/head, 15.9 +/- 5.0 milliunits/mg protein) in rat liver. The distribution of cytochrome c oxidase and that of the PPase in a Nycodenz gradient suggested that the PPase is an original peroxisomal enzyme and not a mitochondrial contaminant: This was confirmed by second Nycodenz gradient centrifugation. The optimum pH of the peroxisomal PPase was about 8.5. The activity was specific to inorganic pyrophosphate (PPi), the Km value for PPi being 34.1 +/- 3.3 microM. It was strictly dependent on Mg2+ and showed a sigmoidal dose-response for Mg2+ with an S0.5 value of 100 microM. The activity was inhibited by Ca2+, p-chloromercuriphenylsulfonic acid, Hg2+, N-ethylmaleimide, and NaF. The functions of peroxisomal PPase are discussed.

Animals↗