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Biomedical subjects

H Kushida

Publications and source records attributed to H Kushida.

At least 19 recordsLinked to original sources
PubMed1992

Induction of preneoplastic lesions by a low dose of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in the livers of rats treated with carbon tetrachloride.

The multifactorial nature of carcinogenesis in man has impelled us to study the effects of various chemicals and conditions in combination. In the present investigation, we examined the effects of low doses of 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline (MeIQx) in combination with carbon tetrachloride (CCl4) on the formation of glutathione S-transferase placental form (GST-P)-positive foci in rat liver. Administration of diet containing MeIQx at 0.4, 4 or 40 p.p.m., representing one-thousandth, one-hundredth and one-tenth of the dose proved to induce hepatocellular carcinomas (400 p.p.m.), for 8 or 12 weeks did not induce GST-P-positive foci. However, 40 p.p.m. of MeIQx when co-administered with CCl4 (0.7 ml/kg, s.c. twice a week) induced preneoplastic lesions: 7- and 3-fold increases in the numbers and areas of GST-P positive foci in week 8, and 8- and 6-fold increases respectively in week 12, over those with CCl4 alone. Furthermore, a marked increase in the number of hyperplastic nodules was observed in this group of rats in week 12. No significant increases of GST-P-positive foci were observed at doses of 0.4 or 4 p.p.m. MeIQx in combination with CCl4. Thus, it is predicted that chronic exposure to 40 p.p.m. of MeIQx eventually results in induction of hepatocellular carcinomas in injured rat liver.

Animals↗
PubMed1992

Suppression of spontaneous hepatocellular carcinoma development in C3H/HeNCrj mice by the lipophilic ascorbic acid, 2-O-octadecylascorbic acid (CV-3611).

The study was performed to examine the effects of the lipophilic ascorbic acid, 2-O-octadecylascorbic acid (CV-3611), with a strong scavenging capacity for active oxygen species, on the spontaneous development of liver tumors in male C3H/HeNCrj mice. Animals were given a diet containing 0.1% CV-3611 for a total experimental period of 16 months. Hepatocellular carcinomas developed in 2/39 (5%) of these experimental mice and in 11/43 (26%) of control mice fed a basal diet. The numbers of carcinoma per mouse were 0.05 +/- 0.22 and 0.33 +/- 0.61 respectively. Thus, CV-3611 clearly suppressed the development of hepatocellular carcinomas. However, the scavenger did not affect either the incidence or the number of hepatocellular adenomas, suggesting that active oxygen species might be involved in the conversion of adenomas to carcinomas in spontaneous liver carcinogenesis in C3H/HeNCrj mice.

Animals↗
PubMed1991

Pharmacological characteristics of adenosine-induced inhibition of dog ventricular contractility: dependence on the pre-existing level of beta-adrenoceptor activation.

Experiments were carried out to characterize the adenosine-induced negative inotropic effect in relation to the extent of beta-adrenoceptor activation in the isolated dog left ventricular myocardium. Adenosine and R-N6-phenylisopropyladenosine inhibited the positive inotropic effect of isoprenaline (10(-7) mol/l and lower) about 20% of its maximal response, which was antagonized by an A1 adenosine receptor antagonist 1,3-dipropyl-8-cyclopentylxanthine in a concentration-dependent manner. The negative inotropic effect of adenosine disappeared and that of R-N6-phenylisopropyl-adenosine decreased when the isoprenaline concentration was elevated to the level higher than 10(-7) mol/l. Adenosine deaminase (1.5 U/ml) that abolished the negative inotropic effect of adenosine enhanced the effect of R-N6-phenylisopropyladenosine, indicating that endogenous adenosine released by high isoprenaline concentration (10(-6) mol/l) modulates the interaction. The maximal response to adenosine and R-N6-phenylisopropyladenosine determined in the presence of 10(-7) mol/l isoprenaline was 50% of that of carbachol which elicited the maximal inhibition even in the presence of 10(-6) mol/l isoprenaline. The negative inotropic effects of R-N6-phenylisopropyladenosine and carbachol were additive to the maximal response equivalent to that of carbachol. The difference in the efficiency between the adenosine and muscarinic receptor agonists may be partly ascribed to the difference in densities of the respective receptors in the dog ventricular myocardium. The negative inotropic effect of R-N6-phenylisopropyladenosine in the presence of isoprenaline was associated with decrease in cyclic AMP levels elevated previously by isoprenaline.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine↗
PubMed1991

Inhibitory effect of 2-O-octadecylascorbic acid in agglutination assay with concanavalin A; short-term examination of rat urinary bladder carcinogenesis.

A derivative of ascorbic acid, 2-O-octadecylascorbic acid (CV-3611), is a strong scavenger of active oxygen species. We examined the effect of CV-3611 on a short-term test of bladder carcinogenesis, using concanavalin A (Con A)-dependent agglutination of isolated bladder epithelial cells. Rats were given 0.01% N-butyl-N-(4-hydroxybutyl)nitrosamine (BHBN) for 1 week, and then 5% sodium saccharin or 2% DL-tryptophan or 0.01% BHBN alone or with 0.002, 0.006 or 0.02% CV-3611 for 3 weeks. Treatment with CV-3611 reduced the effects of the bladder tumor promoters sodium saccharin and DL-tryptophan by 48-86 and 65-87%, respectively. CV-3611 also reduced the number of aggregates of bladder epithelial cells from rats treated with BHBN for 4 weeks. These results suggest that CV-3611 has a suppressive effect on rat bladder carcinogenesis.

Agglutination Tests↗
PubMed1991

Histochemical demonstration of heavy metals in the hippocampal formation embedded in Quetol 523M.

To facilitate improvement of investigations on the distribution of mossy fibers in the hippocampal formation, a method is described using Timm's stained preparations after methacrylate embedding with the hydrophilic resin, Quetol 523M. Fixation with a mixture of formaldehyde and glutaraldehyde yielded satisfactory staining results and good structural preservation. During the course of histochemical experiments employing Timm's staining, examinations revealed that sulfide silver reaction products were consistently present in both the mossy fibers themselves and their terminals associated with the dendrites of pyramidal cells in tissue sections of 1-2 microns in thickness. The results obtained also revealed that variations of the mossy fiber system occurred in the neurological mutant mouse dreher (dr). The bundles of mossy fibers forming the intrapyramidal synaptic field may be considered to reflect genotype-dependent differences in the mutation. The present method is adequate for allowing the histochemical demonstration of mossy fibers and their giant boutons by light microscopy.

Animals↗
PubMed1991

En bloc staining available for stereoscopic observation of epoxy resin Quetol 651-embedded thick sections under a high voltage transmission electron microscope.

This method has been devised for easy en block staining for stereoscopic observation of thick sections under a high voltage transmission electron microscope (HVTEM). It uses carbohydrazide as an osmium bridging agent and both osmium tetroxide and uranyl acetate as electron staining agents. Osmium tetroxide-fixed and en bloc-stained tissue blocks are embedded in a Quetol 651 resin mixture. Thick sections (2-3 microns thick) without double staining are observed at an accelerating potential of 300 kV and a tilt angle of +/- 10 degrees by an H-9000 TEM with a side-entry goniometer. Stereoscopic electron micrographs can be obtained.

Animals↗
PubMed1991

Correlative light and electron microscopic observations on ectopic neurons in the cerebellum of dreher mutant mouse.

The structure of ectopic neurons in the cerebellum of dreher mutant mouse was investigated by correlative light and electron microscopic observations. Tissue blocks were fixed in buffered aldehyde and embedded in a mixture of 2-hydroxypropyl methacrylate, Quetol 523, and methyl methacrylate. Sections at 0.4-0.5 microns in thickness were examined by electron microscopy after observation under a light microscope. By comparing the electron images with those of light microscopy in the same sites, the structures of ectopic cells were confirmed. Ectopic Purkinje cells were arranged with cell bodies that contained an oval, spherical or wrinkled nucleus without deep invagination and the thin layers of endoplasmic reticulum at the perinuclear regions. Granule cells were ectopically matured in the external granular layer and within the cluster at the cortical region. This method provides a useful procedure for understanding structures of the cerebellar neurons of the mutant.

Animals↗
PubMed1990

The preferential inhibition of alpha 1- over beta-adrenoceptor-mediated positive inotropic effect by organic calcium antagonists in the rabbit papillary muscle.

Experiments were carried out to elucidate the mechanism that the positive inotropic effect mediated by alpha 1-adrenoceptors is more susceptible to organic calcium antagonists than the beta-adrenoceptor-mediated effect. Verapamil and diltiazem displaced the specific binding of [3H]prazosin to the membrane fraction derived from the rabbit ventricular myocardium, verapamil being about 70 times more potent than diltiazem. Nifedipine did not displace the binding. While these compounds suppressed the positive inotropic effect mediated via alpha 1-adrenoceptors in a concentration-dependent manner, there was no correlation between the potency of the compounds to displace the [3H]prazosin binding and to inhibit the alpha-mediated positive inotropic effect. The relative potency of three calcium antagonists to decrease the basal force of contraction and the alpha 1-mediated effect (of the same extent as compared to basal force of contraction) was consistent to each other. The positive inotropic effect mediated by beta-adrenoceptors was inhibited much less, and was enhanced by low concentrations of organic calcium antagonists. The differential action of calcium antagonists on the alpha- and beta-mediated positive inotropic effect was mimicked by lowering the extracellular calcium concentration to 1/2, 1/4 and 1/8 of that in normal Krebs-Henseleit solution (2.5 mmol/l). These results indicate that the alpha 1-adrenoceptor blocking activity does not play an essential role for the preferential inhibition of alpha-mediated positive inotropic effect by organic calcium antagonists. Difference in the subcellular mechanism involved in mobilization of intracellular Ca2+ subsequent to alpha 1- and beta-adrenoceptor activation may be responsible for the differential inhibitory action of calcium antagonists in the rabbit heart.

Animals↗
PubMed1990

Staining of intestinal goblet cells with ruthenium red in semithin sections.

For a correlative light and electron microscopy of intestinal goblet cells, postembedding staining with ruthenium red (RR) was performed in epoxy-embedded sections. Tissue blocks were fixed in buffered aldehyde and embedded in a mixture of Quetol 651, nonenyl succinic anhydride (NSA), methyl nadic anhydride (MNA), and DMP-30. Sections at 0.4-0.5 micron in thickness were mounted on grids and were treated with an aqueous solution of RR followed by osmium tetroxide, uranyl acetate and lead citrate. Postembedding staining of epoxy sections revealed the interaction between RR and anionic groups by both light and electron microscopy. Light and electron microscopic observation of identical sites in semithin sections was successful for the correlations of colored reaction with electron density.

Animals↗
PubMed1989

Preponderance of beta- over alpha-adrenoceptors in mediating the positive inotropic effect of phenylephrine in the ferret ventricular myocardium.

[3H]prazosin bound to the membrane fraction derived from the ferret ventricular muscle with high affinity in a saturable manner (Kd = 0.25 nmol/l and Bmax = 27 fmol/mg protein in the right ventricle). [3H]CGP-12177, a beta-adrenoceptor ligand, bound to the membrane fraction with a Kd value of 0.29 nmol/l and a Bmax of 42 fmol/mg protein. In the isolated ferret papillary muscle driven at 1 Hz at 37 degrees C, phenylephrine elicited a concentration-dependent positive inotropic effect. The maximal effect of phenylephrine was comparable to that of isoprenaline. Prazosin (0.3 mumol/l) shifted the concentration-response curve for phenylephrine slightly but significantly to the right, the maximal response being unaffected. In contrast, bupranolol (0.3 mumol/l) shifted the curve for phenylephrine markedly downwards: the maximal response was depressed significantly to 40% and the curve became less steep. In the presence of prazosin and bupranolol the curve was shifted to the right, being essentially parallel to the control curve. These results indicate that in the ferret ventricular myocardium both alpha- and beta-adrenoceptors mediate the positive inotropic effect of phenylephrine. The extent of contribution of the two classes of adrenoceptor is quite different from that in other mammalian species. In the ferret heart, beta-adrenoceptors predominate over alpha-adrenoceptors in mediating the positive inotropic effect of phenylephrine, although the number of beta-adrenoceptors is not especially high when compared with other species.

Adrenergic beta-Antagonists↗
PubMed1989

Use of semithin sections embedded in a water-miscible methacrylate for light microscopy of central nerve tissues.

Semithin sections embedded in water-miscible methacrylates were used for the study of fine structures of cells and tissues in the central nervous system by light microscopy instead of the conventional paraffin sections. This method used a water-miscible methacrylate mixture consisting of 2-hydroxypropyl methacrylate (HPMA), Quetol 523 and methyl methacrylate (MMA) as an embedding medium. The mixture had a low viscosity, was easy to handle and penetrated readily and completely into the specimen, producing a homogenous block from which it was easy to make sections 1.5 microns thick. Staining could be localized far more precisely in these sections than in paraffin sections owing to the thickness of the semithin sections and to the excellent structural preservation of cellular components.

Acrylates↗
PubMed1988

Further characterization of the myocardial alpha-adrenoceptors mediating positive inotropic effects in the rabbit myocardium.

[3H]Prazosin bound with high affinity to the membrane fraction derived from the rabbit ventricular myocardium. Oxymetazoline displaced [3H]prazosin from its binding site, did not elicit a positive inotropic effect but antagonized the positive inotropic effect of phenylephrine mediated by alpha-adrenoceptors in the presence of a beta-antagonist. Naphazoline was more potent in displacing [3H]prazosin and behaved as a weak partial agonist. YM-12617 (5-[2-[[2-(2-ethoxyphenoxy)ethyl]amino]propyl]-2- methoxybenzenesulfonamide HCl), a potent selective alpha 1-antagonist, displaced [3H]prazosin and antagonized the alpha-mediated positive inotropic effect with equal potency. Thus, a good correlation was found between the potency of alpha-antagonists to displace [3H]prazosin and their ability to antagonize the alpha-mediated positive inotropic effect. On the other hand, there was no significant correlation between the Ki and the pD2 value of the alpha-agonists (norepinephrine, epinephrine, phenylephrine and naphazoline), indicating that there is a non-linear relationship between agonist binding to myocardial alpha 1-adrenoceptors and subsequent functional changes. Myocardial alpha 1-adrenoceptors showed some pharmacological characteristics which appear to be different from those in smooth muscle tissues.

Adrenergic alpha-Agonists↗
PubMed1988

Phorbol ester does not mimic, but antagonizes the alpha-adrenoceptor-mediated positive inotropic effect in the rabbit papillary muscle.

The phorbol ester 12-O-tetradecanoyl phorbol-13-acetate (TPA) was used to examine the hypothesis that phosphoinositide turnover is involved in the regulation of myocardial contractility mediated by stimulation of alpha-adrenoceptors in the mammalian cardiac muscle. Exposure of the isolated rabbit papillary muscle electrically driven at a rate of 1 Hz at a temperature of 37 degrees C to TPA in concentrations of 10-1000 nmol/l for 30 min did not affect the basal force of contraction. The concentration-response curve for the positive inotropic effect of (-)-phenylephrine mediated by stimulation of alpha-adrenoceptors in the presence of (+/-)-bupranolol (100 nmol/l) was shifted to the right and downward by TPA in concentrations of 30-1000 nmol/l, while the effect of (-)-phenylephrine mediated by stimulation of beta-adrenoceptors in the presence of prazosin (100 nmol/l) was not decreased, but slightly enhanced by exposure of the muscle to relatively low concentrations of TPA (10-100 nmol/l). Incubation of the membrane fraction isolated from the rabbit ventricular muscle with TPA in vitro under the same condition as employed in the physiological experiments decreased the specific binding of [3H]prazosin but not that of [3H]CGP-12177, while the non-tumor promoting phorbol ester, alpha PDD, was ineffective. These results indicate that activation of protein kinase C by TPA does not mimic the positive inotropic effect of catecholamines mediated by activation of myocardial alpha-adrenoceptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Antagonists↗
PubMed1988

Enantiomers of dobutamine increase the force of contraction via beta adrenoceptors, but antagonize competitively the positive inotropic effect mediated by alpha-1 adrenoceptors in the rabbit ventricular myocardium.

Experiments were carried out to characterize the pharmacological properties of enantiomers and racemic mixture of dobutamine to modulate the myocardial contractility through alpha and beta adrenoceptors in the rabbit papillary muscle. Dobutamine caused the concentration-dependent positive inotropic effect: the rank order of potency was R-(+)- greater than (+/-) - greater than S-(-)-dobutamine. The positive inotropic effect of (+)-, (-)- and (+/-)-dobutamine was antagonized by a beta adrenoceptor antagonist, (+/-)-bupranolol in a competitive manner, but was not affected by an alpha-1 adrenoceptor antagonist, prazosin. The concentration-response curve for (-)-phenylephrine mediated by alpha adrenoceptors in the presence of 10(-6) M (+/-)-bupranolol was shifted by enantiomers of dobutamine to the right in a concentration-dependent manner. Thus, enantiomers of dobutamine antagonized the positive inotropic effect of (-)-phenylephrine in a competitive manner, and pA2 values [negative logarithm of the dissociation constant (KB)] for (+)- and (-)-dobutamine were 6.67 and 5.99, respectively. The specific binding of [3H]prazosin to membrane fractions of rabbit ventricular myocardium was displaced by dobutamine with a high potency: the -log Ki values for (+)- and (-)-dobutamine were 6.43 and 5.97, respectively, which correspond well with pA2 values of these compounds for functional modification. These findings indicate that enantiomers of dobutamine elicit the positive inotropic effect through activation of beta adrenoceptors, whereas both enantiomers behave as the competitive antagonist of myocardial alpha adrenoceptors mediating the positive inotropic effect in the isolated rabbit papillary muscle.

Animals↗
PubMed1986

PAS staining of eosinophils in semi-thin sections of bone marrow embedded in glycol methacrylate.

The usual periodic acid-Schiff (PAS) reaction of glycosaminoglycans is applicable to paraffin embedded material. A modification for water-miscible methacrylate embedded tissue suitable for correlative light and electron microscopic studies, which makes it possible to find the same stained cell in a semi-thin tissue section, is described. Eosinophil leukocytes in semi-thin sections from bone marrow were verified by electron microscopy after staining with the PAS reaction. Immature eosinophil leukocyte granules reacted with and without previous salivary treatment. This method facilitates the search for localization of glycosaminoglycans in scattered blood cells.

Animals↗