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Biomedical subjects

M Fujimoto

Publications and source records attributed to M Fujimoto.

At least 505 records · Page 28Linked to original sources
PubMed1983

Influence of phospholipase treatments on ligand bindings to a benzodiazepine receptor-GABA receptor-chloride ionophore complex.

Treatment of rat cerebellar membranes with phospholipase A2 (PLA2) or phospholipase C (PLC) increased basal [3H]diazepam binding at 0 degrees C with concomitant disappearance of the stimulatory effect of Cl- ion on the binding. On the other hand, these treatments did not affect the stimulatory effect of GABA, nor the maximum enhancement obtained in the presence of both GABA and Cl- ion. These results suggest that PLA2 or PLC modified the phospholipids responsible for the interaction between the benzodiazepine receptor and the Cl- ionophore. This assumption was supported by the results of thermodynamic experiments which showed that the changes in thermodynamic parameters occurring after the addition of Cl- ion resembled those after PLA2 or PLC treatment. Since the effect of PLA2 was evident at very low concentrations, and a PLC concentration of at least one order of magnitude higher was required to induce a similar effect, the change of phospholipids especially to lysophospholipids seems to be of particular importance. Protein release from the membrane, which also occurs after PLA2 or PLC treatment, did not appear to be responsible for the present phenomenon.

Animals↗
PubMed1983

An exploration of the binding site of aldolase using alkanediol monoglycolate bisphosphoric esters.

Alkanediol monoglycolate bisphosphoric esters (P-O-CH2-CO-O-(CH2)n-O-P), which are analogues of the aldolase (D-fructose-1,6-bisphosphate D-glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13) substrate fructose 1,6-bisphosphate, were synthesized and used for probing its active site. The Ki value was lowest when the maximum distance between the phosphorus atoms of the bisphosphate was brought close to that of fructose 1,6-bisphosphate. The binding constants estimated from difference spectra correlate well with Ki values for the substrate analogues. Propanediol monoglycolate bisphosphoric ester protected aldolase from inactivation by 1,2-cyclohexanedione, which preferentially attacks arginine-55. However, propanol phosphate had little protective effect. The synthesized phosphate compounds protected the enzyme against inactivation by trypsin, and also against spontaneous denaturation. These results suggest that the synthesized phosphate compounds bind to aldolase at the active site, which tends to keep the distance constant between the two phosphate-binding sites for the open-chain form of fructose 1,6-bisphosphate, and stabilize the natural conformation of the enzyme. Both arginine-55 and lysine-146 are shown to participate in the phosphate-binding site for the C-1-phosphate of fructose 1,6-bisphosphate.

Animals↗
PubMed1983

Analyses of neural mechanisms mediating the effect of horizontal cell polarization.

Polarization of horizontal cells by current through an intracellular electrode elicited a response in certain other horizontal cells and in bipolar cells. Results were consistent with the hypothesis that the feedback from horizontal cells to photoreceptors play an important role in the center-surround antagonism of bipolar cells and also in the opponent color responses of horizontal cells. The feedback from L-type horizontal cells must be mediated by GABA. The red component of RG-type and RGB-type horizontal cells was suppressed by application of GABA antagonists. GABA receptors was not found in these cells, suggesting that the effect was mediated through photoreceptors.

Animals↗
PubMed1982

Ethyl beta-carboline-3-carboxylate reverses the diazepam effect on cerebellar cyclic GMP.

The administration of ethyl beta-carboline-3-carboxylate (beta-CCE), a ligand for benzodiazepine receptors, did not affect the cerebellar cyclic GMP level in mice, but giving beta-CCE together with diazepam significantly inhibited the diazepam-induced decrease in cyclic GMP. The fact that no antagonism was observed when beta-CCE was given 15 min before the diazepam treatment indicates that beta-CCE is short-acting. These biochemical observations support the conclusions from behavioral and electrophysiological studies which indicated that beta-CCE is a short-acting antagonist of benzodiazepines.

Animals↗
PubMed1982

Effects of geranylgeranylacetone on gastrointestinal secretion in rats.

The effects of geranylgeranylacetone (GGA), a new acyclic polyisoprenoid with a novel antiulcer action on gastrointestinal secretion were studied in rats. Intraduodenal administration of GGA (1-30 mg/kg) dose-relatedly reduced the gastric acid secretion caused by pentagastrin, histamine or insulin. On the other hand, GGA (3-30 mg/kg i.d.) dose-relatedly increased pancreatic secretion but did not affect biliary secretion. The above-mentioned findings seem consistent with the further findings that GGA depressed a plasma gastrin level enhanced by insulin while in increased the basal level of plasma secretin. These pharmacological features found in the present studies may partially, at least, account for the mechanism of GGA antiulcer action.

Animals↗
PubMed1982

Comparison of the effects of GABA and chloride ion on the affinities of ligands for the benzodiazepine receptor.

The effects of gamma-aminobutyric acid (GABA), Cl- ion, and their combination were examined on the affinity for the benzodiazepine (BDZ) receptor of various ligands including BDZ's, putative endogenous ligands, and CL218872. When the binding was performed at 0 degrees, the ligands could be classified into four types according to the action of the modulators. The affinity of the first type including some BDZ's increased with GABA, Cl- ion, or their combination while that of the second type consisting of 3-hydroxy-benzodiazepines, CL218872, and inosine increased only with GABA. In the third type, designated as the beta-carboline type, only Cl- ion increased the affinity of the ligands. The fourth inactive type included pentylenetetrazole and nicotinamide and the affinities of these ligands were not affected by GABA or Cl- ion. At 37 degrees, although GABA still retained the property to increase the affinity of prototype compounds of the first and second types, the effect of C1- ion became somewhat ambiguous. Comparison of the present results with published data on the pharmacological and electrophysiological activities of the ligands suggested that the compounds with affinity increased by GABA act as BDZ-mimetics, while those belonging to types 3 and 4 act as antagonists. Also, our results seem to throw some additional light upon the relationships among the benzodiazepine receptor, the GABA receptor, and the chloride ionophore.

Animals↗
PubMed1982

Double antibody enzyme immunoassay for the quantitation of adenosine 3', 5' -cyclic monophosphate (cyclic AMP) and guanosine 3', 5'-cyclic monophosphate (cyclic GMP) in tissue and plasma.

A sensitive double antibody enzyme immunoassay for the quantitation of cyclic AMP and cyclic GMP is presented. Specific antisera to each nucleotide were raised in rabbits by immunization with succinyl cyclic nucleotide-human serum albumin conjugates. For competitive reaction, antibodies were incubated with a mixture of succinyl cyclic nucleotide labelled with beta-D-galactosidase and unlabelled succinylated standard or sample cyclic nucleotides. The antibody-bound enzyme-hapten was separated from free hapten by anti-rabbit IgG immobilized to a polystyrene ball. Activity of the enzyme on the solid phase was fluorometrically determined. The assay system made it possible to ascertain values as low as 5 fmole of cyclic AMP or cyclic GMP. Cyclic nucleotides in plasma could be accurately determined by this method without requiring a deproteinizing reagent as the first step of assay.

Animals↗
PubMed1982

Intracellular measurement of Na activity using neutral carrier Na ion-selective microelectrode.

To measure intracellular Na+ activities, a double-barreled Na ion-selective microelectrode was constructed with a neutral carrier Na+ ligand (ETH 227). The slope constant and detection limit were 56 mV and 3-5 mM, respectively, the selectivity of Na+/K+ being 40-50. In the bullfrog, the cell Na+ was 14.4 mEq/liter for the proximal tubule and 12.6 mEq/liter for the sartorius muscle.

Animals↗
PubMed1982

Microinjection of macromolecules into normal murine lymphocytes by means of cell fusion. II. Enhancement and suppression of mitogenic responses by microinjection of monoclonal anti-cyclic AMP into B lymphocytes.

Reproducible methods are now available for introducing protein molecules such as antibodies into normal murine lymphocytes by fusion with protein molecule-containing erythrocyte ghosts. Monoclonal antibodies against cyclic AMP were raised by hybridoma technique and packed into erythrocyte ghosts. Then, monoclonal anti-cyclic AMP containing ghosts were fused with splenic B lymphocytes by polyethylene glycol-mediated fusion at various intervals after LPS stimulation. This method made it possible for us to quantitatively microinject antibodies into B lymphocytes. Microinjection of anti-cyclic AMP antibody molecules into lymphocytes at a very early stage of LPS stimulation resulted in a marked enhancement of DNA synthetic responses as well as increased numbers of plaque-forming cells. Intracellular cyclic AMP levels were found to be markedly decreased after microinjection of monoclonal anti-cyclic AMP, suggesting that lowering the intracellular cyclic-AMP level in the B lymphocytes at an early stage of stimulation might have induced the enhanced proliferative as well as differentiative responses to LPS. Similar enhancing effects on cell proliferation were obtained when antibodies were injected 18 hr after stimulation. Microinjection of anti-cyclic AMP at 12 hr after culture, however, inhibited the DNA synthetic responses, and induction of plaque-forming cells was suppressed when anti-cyclic AMP was injected 6 hr after LPS stimulation. The present data suggest the biphasic regulatory roles of cyclic AMP at the early stage of B lymphocyte activation. This approach may be useful in identifying regulatory molecules in B lymphocyte induced by mitogenic or antigenic stimulation.

Animals↗
PubMed1982

[Determination of dissociation exponent of CO2 used in Henderson-Hasselbalch equation by means of bicarbonate-selective microelectrode].

To measure intracellular ion activity of bicarbonate directly, a double-barreled bicarbonate-selective microelectrode was constructed with a liquid ion exchanger(LIX) containing tri-n-octylammonium, trifluoroacetyl-butyl benzene and octanol. A new dissociation exponent of CO2 (pK') was determined experimentally for a modified Henderson-Hasselbalch equation, in which HCO3 activity was used instead of HCO3 concentration. The temperature effects on pK' and solubility coefficient of CO2 were analyzed over the range of 10 to 40 degrees C. The average values of pK' at room temperature (22 degrees C) were 6.377 and 6.348 in water and frog serum, respectively, and the solubility coefficients of CO2 were 0.048 and 0.046 mM/L/mmHg. The values of pK' plotted on Arrhenius plot were shown linear with 1/T, indicating that the pK' thus obtained can be treated thermodynamically as a first-order kinetics. Under normal circumstances, the values for the intracellular pH predicted from the intracellular bicarbonate activities in the proximal tubule and sartorius muscle of bullfrogs were virtually identical with those obtained directly by the LIX-pH-microelectrodes.

Animals↗