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Biomedical subjects

M Negishi

Publications and source records attributed to M Negishi.

At least 271 records · Page 15Linked to original sources
PubMed1991

Heterogeneous response in calcium signaling by adrenergic and cholinergic stimulation in cultured bovine trabecular cells.

Adrenergic and cholinergic drug-dependent Ca(2+)-signaling in cultured bovine trabecular cells was studied using a fluorescent calcium dye, fura-2, and digital video-imaging system. The resting calcium concentration in the cultured cell was 178.7 +/- 1.8 nM. Upon stimulation with norepinephrine, isoproterenol and carbachol trabecular cells responded heterogeneously. Many types of reaction to the stimuli were found; about 25% of the cells responded to the stimuli and about 30% showed no response to all of the stimuli. Carbachol induced response was blocked with pretreatment of atropine, and timolol inhibited the isoproterenol-induced response. Ca2+ signaling in trabecular cells may mediate the action of adrenergic and cholinergic drugs and possibly play significant roles in the regulation of intraocular pressure.

Adrenergic beta-Agonists↗
PubMed1991

TEI-9063, a stable and highly specific prostacyclin analogue for the prostacyclin receptor in mastocytoma P-815 cells.

The prostacyclin (PGI2) analogues, TEI-9063 and its methyl ester, TEI-1324, have been compared with another stable analogue, iloprost, with respect to binding to the PGI2 receptor, stimulation of adenylate cyclase activity and inhibition of thrombin-induced Ca2+ mobilization in mastocytoma P-815 cells. TEI-9063 displaced the [3H]iloprost binding to the membrane fraction, the IC50 value being 3 nM, but showed very low affinity for the PGE receptor. TEI-9063 dose dependently stimulated cAMP formation in the cells and GTP-dependent adenylate cyclase activity in the membrane fraction, the EC50 value being 50 and 10 nM, respectively. Furthermore, TEI-9063 prevented the thrombin-induced increase in the intracellular Ca2+ concentration, the IC50 value being 50 nM. These IC50 and EC50 values are lower than those obtained for iloprost. On the other hand, those of TEI-1324 were about two-orders higher. Although PGI2 lost its ability to stimulate cAMP formation by preincubation for 20 min at 37 degrees C, TEI-9063 completely retained its ability after 60-min preincubation. These results demonstrate that TEI-9063 is a stable and stronger agonist for the PGI2 receptor than iloprost, and that it prevents thrombin-induced Ca2+ mobilization through stimulation of the adenylate cyclase system in mastocytoma cells.

Adenylyl Cyclases↗
PubMed1991

Central administration of Lys-D-Pro-Thr, an interleukin-1 beta 193-195 analogue, stimulates feeding in rats.

We determined the effect on feeding of Lys-D-Pro-Thr (LDPT), an interleukin-1 beta 193-195 analogue which antagonizes the analgesic effects of interleukin-1 beta. Intracerebroventricular administration of 2 micrograms/rat LDPT increased food consumption at the 0-1 h time period, although food intake was reduced by LDPT at the 1-2 h time period. There was no effect on colonic temperature 1 h later. Subcutaneously injected LDPT (2 micrograms/rat) failed to increase food intake for 1 h. These data suggest that brain interleukin-1 beta may have a physiological role in feeding suppression.

Amino Acid Sequence↗
PubMed1991

Sucrose feeding at weaning alters the preference for sucrose in adolescence.

The present studies were undertaken to examine the hypothesis that sucrose feeding at weaning may alter the preference for sucrose in the adolescence. Chronological changes of hypothalamic dopamine (DA), its metabolite, 3,4-dihydroxy-phenylacetic acid (DOPAC) and norepinephrine (NE) contents were also measured by HPLC. In 21 day-aged rats, 10% sucrose or water was given as drinking water for 3 weeks. From 9 weeks, all animals were maintained under a free choice between 30% sucrose solution and water. Sucrose-ingested rats more preferred to 30% sucrose solution than control rats and body weight gain of sucrose group was significantly greater than that of controls. Hypothalamic DA content was significantly decreased at 6 and 13 weeks and the DOPAC/DA ratio increased at 3, 6, and 13 weeks later. In contrast, hypothalamic NE concentration was not changed at all. The data obtained herein suggest that sucrose feeding at weaning alters the preference for sucrose, resulting in an overweight gain, and that the observed increase of hypothalamic DA metabolism may be involved in the altered preference for sucrose.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1991

Sodium fluoride mimics the effect of prostaglandin E2 on catecholamine release from bovine adrenal chromaffin cells.

We have reported recently that prostaglandin E2 (PGE2) stimulated phosphoinositide metabolism in bovine adrenal chromaffin cells and that PGE2 and ouabain, an inhibitor of Na+, K(+)-ATPase, synergistically induced a gradual secretion of catecholamines from the cells. Here we examined the involvement of a GTP-binding protein(s) in PGE receptor-induced responses by using NaF. In the presence of Ca2+ in the medium, NaF stimulated the formation of all three inositol phosphates, i.e., inositol monophosphate, bisphosphate, and trisphosphate, linearly over 30 min in a dose-dependent manner (15-30 mM). This effect on phosphoinositide metabolism was accompanied by an increase in cytosolic free Ca2+. NaF also induced catecholamine release from chromaffin cells, and the dependency of stimulation of the release on NaF concentration was well correlated with those of NaF-enhanced inositol phosphate formation and increase in cytosolic free Ca2+. Although the effect of NaF on PGE2-induced catecholamine release in the presence of ouabain was additive at concentrations below 20 mM, there was no additive effect at 25 mM NaF. Furthermore, the time course of catecholamine release stimulated by 20 mM NaF in the presence of ouabain was quite similar to that by 1 microM PGE2, and both stimulations were markedly inhibited by amiloride, with half-maximal inhibition at 10 microM. Pretreatment of the cells with pertussis toxin did not prevent, but rather enhanced, PGE2-induced catecholamine release over the range of concentrations examined. These results demonstrate that NaF mimics the effect of PGE2 on catecholamine release from chromaffin cells and suggest that PGE2-evoked catecholamine release may be mediated by the stimulation of phosphoinositide metabolism through a putative GTP-binding protein insensitive to pertussis toxin.

Adrenal Medulla↗
PubMed1991

Characterization of prostaglandin E2-induced Ca2+ mobilization in single bovine adrenal chromaffin cells by digital image microscopy.

We recently reported that prostaglandin E2 (PGE2) stimulates phosphoinositide metabolism accompanied by an increase in intracellular free Ca2+ concentration ([Ca2+]i) in cultured bovine adrenal chromaffin cells. In the present study, temporal and spatial changes in [Ca2+]i induced by PGE2 in fura-2-loaded individual cells were investigated by digital image microscopy and were compared with those induced by nicotine and histamine. Image analysis of single cells revealed that responses to PGE2 showed asynchrony with the onset of [Ca2+]i changes. After a lag time of 10-30 s, PGE2-induced [Ca2+]i changes took a similar prolonged time course in almost all cells: a rapid rise followed by a slower decline to the basal level over 5 min. Few cells exhibited oscillations in [Ca2+]i. In contrast, nicotine and histamine induced rapid and transient [Ca2+]i changes, and these [Ca2+]i changes were characteristic of each stimulant. Whereas pretreatment of the cells with pertussis toxin (100 ng/ml, 6 h) did not block the response to any of these stimulants, treatment with 12-O-tetradecanoylphorbol 13-acetate (100 nM, 10 min) completely abolished [Ca2+]i changes elicited by PGE2 and histamine. In a Ca2(+)-free medium containing 3 mM EGTA, or in medium to which La3+ was added, the [Ca2+]i response to nicotine disappeared, but that to histamine was not affected significantly. Under the same conditions, the percentage of the cells that responded to PGE2 was reduced to 37% and the prolonged [Ca2+]i changes induced by PGE2 became transient in responding cells, suggesting that the maintained [Ca2+]i increase seen in normal medium is the result of a PGE2-stimulated entry of extracellular Ca2+. Whereas the organic Ca2(+)-channel blocker nicardipine inhibited [Ca2+]i changes by all stimulants at 10 microM, these [Ca2+]i changes were not affected by any of the organic Ca2(+)-channel blockers, i.e., verapamil, diltiazem, nifedipine, and nicardipine, at 1 microM, a concentration high enough to inhibit voltage-sensitive Ca2+ channels. These results demonstrate that PGE2 may promote Ca2+ entry with concomitant release of Ca2+ from intracellular stores and that the mechanism(s) triggered by PGE2 is apparently different from that by histamine or nicotine.

Adrenal Medulla↗
PubMed1991

Prostaglandin E2 activates Ca2+ channels in bovine adrenal chromaffin cells.

We have demonstrated that prostaglandin E2 (PGE2) treatment of bovine adrenal chromaffin cells results in a sustained elevation of intracellular Ca2+ concentration ([Ca2+]i) in these cells. Because the continued elevation of [Ca2+]i was dependent on extracellular Ca2+ concentration, it can be assumed that the PGE2-induced [Ca2+]i increase is due, at least in part, to an opening of membrane Ca2+ channels. In this study, we used electrophysiological methods to examine the mechanism of the PGE2-induced [Ca2+]i increase directly. Puff application of PGE2 to the external medium resulted in a prolonged depolarization in about half of the chromaffin cells examined. In whole-cell voltage-clamp recordings, an increase in inward current was observed over a 6-7 min period following bath application of PGE2 (greater than or equal to 10 microM), even in the absence of external Na+. This inward current was abolished when the recordings were made with the cells in a Ca2(+)-free medium, but it was not inhibited by Mn2+, a blocker of voltage-dependent Ca2+ channels. In cell-attached patch-clamp configuration, PGE2 produced an increase in the opening frequency of inward currents. The reversal potential of the PGE2-induced currents was about +40 mV, which is close to the reversal potential of the Ca2+ channel. The opening frequency was not affected by membrane potential changes. In inside-out patch-clamp configuration, inositol 1,4,5-trisphosphate (2 microM) added to the cytoplasmic side activated the Ca2(+)-channel currents, but PGE2 was ineffective when applied to the cytoplasmic side. These results suggest that PGE2 activates voltage-independent Ca2+ channels in chromaffin cells through a diffusible second messenger, possibly inositol 1,4,5-trisphosphate.

Adrenal Medulla↗
PubMed1991

[Cryptosporidium diarrhea developing in two Japanese adults--one in AIDS and the other in a normal host. Research Group for Infectious Enteric Diseases, Japan].

Case I: A middle-age homosexual male developed AIDS with Pneumocystis carinii pneumonia (PCP) and esophageal candidiasis in 1986 during his stay in an European country about five months prior to transfer to Tokyo Metropolitan Komagome Hospital, Tokyo, in 1987. He was also diagnosed as having cryptosporidiosis presenting with mild diarrhea a month following the diagnosis of PCP. Diarrhea was successfully treated with spiramycin. On transfer to Tokyo Metropolitan Komagome Hospital, he was febrile but had no diarrhea. Serum HIV and TPHA were positive and his blood lymphocyte subset T4a was markedly decreased. On the 13th day after transfer to the hospital, watery diarrhea appeared. Cryptosporidium oocysts were detected from the feces taken on the 17th hospital day. The patient died of Escherichia coli septicemia on the 38th hospital day. Autopsy finding yielded Cryptosporidium infection widely spread over the stomach, ileum, bile and pancreatic ducts. Case II: A 31-year-old previously healthy female presented with abrupt onset of mucous stool five times daily. Mucous passage continued on the subsequent days despite administration of loperamide, and the passage increased to 20 times daily with mucous to watery diarrhea associated with mild abdominal cramps and nausea on the 4th day after onset of illness. On the 6th day of illness, she visited Tokyo Metropolitan Komagome Hospital. She denied close contact with pet animals or contact with any person presenting diarrhea. She had no recent history of travelling anywhere outside Tokyo. On examination she was an apparently healthy woman except for a slightly distended abdomen with localized tenderness in the right upper quadrant.(ABSTRACT TRUNCATED AT 250 WORDS)

Acquired Immunodeficiency Syndrome↗
PubMed1991

[Clinical research on patients with typhoid and paratyphoid fever (1984-1987). Research Group for Infectious Enteric Diseases, Japan].

Patients with typhoid or paratyphoid fever, admitted to 14 hospitals for infectious diseases during 1984-1987, were epidemically and clinically studied. Of the total number of 183 typhoid, 49 paratyphoid fever patients, those infected overseas was 44.3% and 71.4% respectively, giving an overall annual decrease, yet marking an increased ratio of overseas infection. Patients aged 20s-30s and males were dominant. One hundred and seventy six cases (96.2%) of typhoid and all the paratyphoid fever cases were bacteriologically diagnosed. The period from the onset to the diagnosis was around 14 days in most cases, but beyond 29 days in over 10% of the cases. We would like to emphasize that enteric fever, focusing on high fever, bradycardia, roseola, hepatosplenomegaly, leukopenia, elevated serum-GOT GPT and LDH, can be easily diagnosed by blood/stool culture before beginning chemotherapy. Intestinal bleeding was recognized in 24 cases (13.1%) of typhoid and 4 (8.2%) of paratyphoid fever, intestinal perforation in 2 (1.1%) and death in 1 (0.5%) of typhoid fever. CP was most commonly used in chemotherapy. Bacteriological relapse was recognized in 7/127 cases (5.5%) of typhoid, 6/48 (13.0%) of paratyphoid fever those followed beyond 3 weeks, though eradication was attained by retreatment. One strain of S. typhi resistant to CP.ABPC.KM.SM was isolated in 1986 from a patient infected overseas. New quinolones seem reliable in our preliminary studies.

Adolescent↗
PubMed1991

Stimulatory effect of guanine nucleotides on prostaglandin E1 binding to murine renal outer medulla.

Prostaglandin E1 (PGE1) specifically bound to the membrane prepared from murine renal outer medulla. The extent of binding of [3H]PGE1 to the membrane was increased about 4-fold by guanosine triphosphate (GTP) and its analogs, but the dissociation of bound [3H]PGE1 from the membrane was in turn enhanced by GTP gamma S. Scatchard plot analyses revealed that GTP gamma S increased the binding affinity more than 2-fold without a major change in the number of binding sites. When [3H]PGE1-bound proteins were cross-linked in the membrane by dithiobis (succinimidyl propionate), bound [3H]PGE1 was no longer dissociated by GTP gamma S treatment, suggesting that cross-linking produced a stable complex of PGE receptor with a GTP-binding protein. The cross-linked [3H]PGE1-specifically bound proteins solubilized from the membranes labeled with [3H] PGE1 in the presence or absence of GTP gamma S were eluted as an apparently single radioactive peak at the same position of Mr = 15000 by gel filtration, indicating that the PGE receptor forms a complex with a GTP-binding protein regardless of the treatment with GTP gamma S by which [3H]PGE1 binding is promoted. The ability of GTP gamma S to stimulate [3H]PGE1 binding was eliminated by pretreatment of the membrane with pertussis toxin, but not cholera toxin, indicating that the PGE receptor is coupled to a pertussis toxin-sensitive GTP-binding protein.

Alprostadil↗
PubMed1990

Inhibited insulin secretion by recombinant human interleukin-1 beta in adrenalectomized rats: involvement of prostaglandin.

An involvement of prostaglandin synthesis in reduced insulin secretion by interleukin-1 was investigated in adrenalectomized (ADX) rats. The recombinant human interleukin-1 beta (IL-1) significantly reduced insulin secretion in ADX rats 2 and 4 hr after the injection, although IL-1 stimulated insulin secretion in intact rats. In ADX rat, IL-1 showed dose-dependent inhibition of pancreatic insulin secretion. In addition, insulin response to intravenous glucose loading was also attenuated in ADX rats with pretreatment by IL-1. At 4 hours after injection, ibuprofen (IBP; 0.5-50.0 mg/kg, ip), selective cyclooxygenase blocker, attenuated insulin inhibition by IL-1 in a dose-dependent manner. These data suggest that IL-1 may suppress in vivo insulin release at least in part through the mediation of prostaglandin synthesis in the absence of adrenal glands.

Adrenal Glands↗
PubMed1990

Involvement of phosphoinositide metabolism in GABA-induced catecholamine release from cultured bovine adrenal chromaffin cells.

The effects of GABA on catecholamine release and phosphoinositide metabolism were studied in cultured bovine adrenal chromaffin cells. GABA and muscimol, a specific agonist for the GABAA receptor, each evoked a gradual secretion of catecholamines from the cells in the presence of ouabain, an inhibitor of Na+, K(+)-ATPase. This release was inhibited by bicuculline, a specific antagonist for the GABAA receptor, or by picrotoxin, a blocker of GABA-gated Cl- channels, and was potentiated by diazepam or pentobarbital. GABA or muscimol induced a concentration-dependent formation of inositol phosphates. This accumulation of inositol phosphates was also inhibited by bicuculline, picrotoxin or removal of extracellular Ca2+, and also potentiated by diazepam and pentobarbital. Nicardipine suppressed GABA-induced catecholamine release in the presence of ouabain and accumulation of inositol phosphates, while verapamil, diltiazem, and omega-conotoxin failed to inhibit these responses to GABA. The phosphoinositide-specific phospholipase C inhibitor neomycin also inhibited both GABA-induced accumulation of inositol phosphates and stimulation of catecholamine release in the presence of ouabain. These results taken together indicate that GABA evoked catecholamine release from the chromaffin cells in the presence of ouabain by stimulation of phosphoinositide metabolism in a Ca2(+)-sensitive manner via activation of GABAA receptor-coupled Cl- channels.

Adrenal Medulla↗
PubMed1990

A specific cis-acting element regulates in vitro transcription of sex-dependent mouse steroid 16 alpha-hydroxylase (C-P450(16 alpha)) gene.

The mouse steroid 16 alpha-hydroxylase (C-P450(16 alpha)) family consists of structurally similar genes with very divergent expressions (Wong, G., Itakura, T., Kawajiri, K., Skow, L., and Negishi, M. (1989) J. Biol. Chem. 264, 2920-2927). Of five family members, the C-P450(16 alpha) gene exhibits a male-specific expression whereas the P450cb gene is equally expressed in both males and females although the level of expression is much lower than that of the C-P450(16 alpha) gene. By DNase I footprinting and in vitro transcription of the C-P450(16 alpha) gene, we determined two cis-acting transcription elements. SDI (sex difference information) in the -84/-102 region from the transcription start site is one of the two cis-acting transcription elements, which is specific to the sex-specific C-P450(16 alpha) gene and confers a high transcription level of this gene. We substituted the nucleotide(s) in the SDI sequence to the corresponding base(s) in the P450cb gene promoter and tested each substituted gene promoter by in vitro transcription. The results indicate that the function of SDI as the transcription element depends most critically on the nucleotides at positions -100 to -98. Another cis-acting transcription element, CTE (common transcription element), is located between -44 and -68 in both C-P450(16 alpha) and P450cb genes. A column chromatography of nuclear extract indicates that the proteins bound to SDI and CTE are different. We speculate therefore that SDI is a specific and necessary element involved in the male-specific C-P450(16 alpha) gene transcription. The sequence comparisons of the corresponding regions of SDI and CTE among the mouse and the closely related rat P450 genes suggest that SDI is a newly evolved cis-acting element in the sex-specific C-P450(16 alpha) gene which arose through gene duplication and selective nucleotide substitution.

Animals↗
PubMed1990

Arachidonic acid stimulates phosphoinositide metabolism and catecholamine release from bovine adrenal chromaffin cells.

Arachidonic acid (AA) evoked a dose-dependent increase in the accumulation of inositol phosphates in cultured bovine adrenal chromaffin cells, and this effect was specific for AA. AA also induced a rise in [Ca2+]i, but this rise was markedly reduced by removal of extracellular Ca2+. AA-induced accumulation of inositol phosphates was absolutely dependent on extracellular Ca2+, and nicardipine and nifedine partially reduced it but verapamil had no effect. Moreover, AA dose-dependently stimulated catecholamine release from chromaffin cells in the presence of ouabain, and this effect was specific for AA. AA-induced catecholamine release in the presence of ouabain was also inhibited by nicardipine and nifedipine but not by verapamil. Furthermore, the phospholipase C inhibitor neomycin inhibited the release. These results taken together suggest that AA stimulates catecholamine release in the presence of ouabain by stimulation of phosphoinositide metabolism in a Ca2(+)-dependent manner.

Adrenal Medulla↗