Biomedical subjects

N Mizuno

Publications and source records attributed to N Mizuno.

At least 73 records · Page 4Linked to original sources

PubMed1998

Metabotropic glutamate receptor subtypes in axon terminals of projection fibers from the main and accessory olfactory bulbs: a light and electron microscopic immunohistochemical study in the rat.

Localization of metabotropic glutamate receptor subtypes, mGluR1, mGluR1alpha, mGluR2/3, mGluR4a, mGluR5, mGluR7a, mGluR7b, and mGluR8, was examined in some of the target areas of projection fibers from the main and accessory olfactory bulbs (MOB and AOB) by using subtype-specific antibodies. The superficial layer of the olfactory tubercle and layer Ia of the piriform cortex, the target areas of MOB, showed marked mGluR1-, mGluR5-, mGluR7a-, and mGluR8-like immunoreactivities (-LI), and rather weak mGluR2/3-LI. The periamygdaloid cortical region including the target areas of both MOB and AOB showed intense mGluR2/3-LI as well as marked mGluR1-, mGluR5-, mGluR7a-, and mGluR8-LI. No significant mGluR1alpha-, mGluR4a-, or mGluR7b-LI was seen in these regions. After transection of the lateral olfactory tract, mGluR2/3-, mGluR7a-, and mGluR8-LI were reduced markedly in the target regions on the side ipsilateral to the transection; no significant changes were detected in mGluR1- or mGluR5-LI. Double labeling experiments indicated light and electron microscopically colocalization of mGluR7a- and mGluR8-LI in axon terminals on dendritic shafts of presumed interneurons in the superficial layer of the olfactory tubercle and layer Ia of the piriform cortex. Electron microscopically mGluR2/3-LI was seen in preterminal and terminal portions of axons, whereas mGluR7a- and mGluR8-LI were associated with presynaptic membrane specialization. Immunolabeled axon terminals were filled with round synaptic vesicles and constituted asymmetric synapses with dendritic profiles. The results suggest that glutamate release from axon terminals of projection fibers from MOB and AOB is regulated presynaptically and differentially through mGluR2/3, mGluR7a, and/or mGluR8.

Animals↗

PubMed1998

Diurnal change of the cold-inducible RNA-binding protein (Cirp) expression in mouse brain.

RNA-binding proteins are believed to play important roles in regulation of neural functions. Recently, a mouse cDNA encoding the cold-inducible RNA-binding protein, Cirp, has been isolated, the amino acid sequence of which showed similarity to plant circadian rhythm proteins. In the present study, diurnal expression of Cirp in the mouse nervous system was examined. Northern blot analysis showed that the level of Cirp mRNA was diurnally regulated in the brain but not in the testis and liver. The level increased during the daytime and decreased during the nighttime. Immunohistochemistry using an anti-Cirp antibody showed that Cirp was expressed in the nucleus of neurons and that the level of Cirp was diurnally regulated in the suprachiasmatic nucleus and the cerebral cortex. The diurnal regulation was not observed in the brain of adult mice kept in constant darkness nor that of 3-day-old mice. These findings suggest that Cirp plays a role in biological rhythms as known for plant proteins and that expression of Cirp is regulated differentially in discrete brain regions.

Animals↗

PubMed1998

Immunohistochemical localization of metabotropic glutamate receptors, mGluR7a and mGluR7b, in the central nervous system of the adult rat and mouse: a light and electron microscopic study.

The distributions of two alternative splicing variants of metabotropic glutamate receptor mGluR7, mGluR7a and mGluR7b, were examined immunohistochemically in the rat and mouse by using variant-specific antibodies raised against C-terminal portions of rat mGluR7a and human mGluR7b. Many regions throughout the central nervous system (CNS) showed mGluR7-like immunoreactivities (LI). The distribution patterns of mGluR7-LI in the rat were substantially the same as those in the mouse, although some species differences were observed in a few regions. Intense mGluR7a-LI was seen in the main and accessory olfactory bulbs, anterior olfactory nucleus, islands of Calleja, superficial layers of the olfactory tubercle, piriform cortex and entorhinal cortex, periamygdaloid cortex, amygdalohippocampal area, hippocampus, layer I of the neocortical regions, globus pallidus, superficial layers of the superior colliculus, locus coeruleus, and superficial layers of the medullary and spinal dorsal horns. The distribution of mGluR7b was more restricted. It was intense in the islands of Calleja, substantia innominata, hippocampus, ventral pallidum, and globus pallidus. The medial habenular nucleus also showed intense mGluR7a-LI in the rat but not in the mouse. For both mGluR7a- and mGluR7b-LI, localization in the active zones of presynaptic axon terminals was confirmed electron microscopically at synapses of both the asymmetrical and symmetrical types. It is noteworthy that mGluR7a-LI is seen preferentially in relay nuclei of the sensory pathways and that both mGluR7a- and mGluR7b-LI are observed not only in presumed glutamatergic axon terminals, but also in non-glutamatergic axon terminals including presumed inhibitory ones. Thus, mGluR7 may play roles not only as an autoreceptor in glutamatergic axon terminals, but also as a presynaptic heteroreceptor in non-glutamatergic axon terminals in various CNS regions.

Amygdala↗

PubMed1998

Cytosolic nuclease activated by caspase-3 and inhibited by DFF-45.

To study the intracellular apoptotic signaling pathways, we have established a cell-free system, in which DNA fragmentation of the isolated mouse liver nuclei was induced with lysates from the Fas-activated cells. We have found that the inactive nuclease present in the intact cell cytosol is activated by a caspase-3-like protease and the activated nuclease induces the nucleosomal DNA fragmentation. We attempted the purification of the inactive nuclease from bovine liver cytosol. The partially purified nuclease was activated by recombinant caspase-3, and to a lesser extent by caspase-6. The activated nuclease was able to digest plasmid DNA in addition to induce the DNA fragmentation of nuclei. DFF-45, which is a subunit of heterodimeric protein leading to DNA fragmentation upon its digestion by caspase-3, is found to inhibit the activity of the activated nuclease. These results suggest that the inactive nuclease in cytoplasm is converted to the active form by caspases, and the activated nuclease enters into nucleus to induce the DNA fragmentation. It is suggested that DFF-45 may function as an inhibitory factor of the caspase-sensitive nuclease in vivo.

Animals↗

PubMed1998

Necessity of endogenous GTP derived from glucose-6-phosphate for insulin secretion augmented by glucose under protein kinase A activation.

To investigate the possible involvement of some intracellular metabolic signaling other than the ATP derived from glucose metabolism under protein kinase A (PKA) activation, we measured the insulin secretory capacity stimulated by glucose and other fuel secretagogues using diazoxide-treated pancreatic islets. Under these conditions, we found a signal from a site proximal to glyceraldehyde-3-phosphate (GA-3-P) in the glycolysis to be necessary for glucose-induced insulin secretion. By using several different glycolytic enzyme inhibitors, we found that this proximal signal is derived from glucose-6-phosphate (G-6-P), and that metabolic signaling distal to GA-3-P also is necessary. Mycophenolic acid completely inhibited the augmented glucose-induced insulin secretion, which guanosine could reverse, indicating that the proximal signaling is coupling with endogenous GTP production. In this novel system of metabolic signaling, endogenous GTP derived from G-6-P in the glycolysis elicits the augmentation of glucose-induced insulin secretion under PKA activation in diazoxide-treated pancreatic islets.

Animals↗

PubMed1998

Role of Mitf in differentiation and transdifferentiation of chicken pigmented epithelial cell.

Mitf encodes a basic helix-loop-helix-leucine-zipper (bHLHzip) protein that is known to function in the development of melanocytes, pigmented epithelial cells (PECs), osteoclasts, and mast cells. In this paper, we report on the isolation, expression, and overexpression of the chicken Mitf and discuss the role of its protein product in the differentiation and transdifferentiation of PECs. Northern blotting showed that chicken Mitf is predominantly expressed in embryonic retinal pigmented epithelium (PE), but is expressed at low levels in other tissues. A 5' RACE analysis revealed differences in the 5' region Mitf nRNA in PE and other tissues. Immunological analysis revealed that Mitf, the protein encoded by Mitf, is first detected in the nuclei of the optic vesicle cells at embryonic stage 13 in a restricted region covered with mesenchymal cells. From stage 14 to 24, the specific staining is observable in the PE and precursor of the PE, the outer layer of the optic cup. In embryos at stages later than stage 29 the signals for Mitf in the future iris, ciliary body, and posterior retinal regions become faint. These results show that expression of Mitf starts at the optic vesicle stage at which no other marker genes for PECs such as mmp115 and tyrosinase are expressed. Dedifferentiation of cultured retinal PECs (rPECs) was induced by phenylthiourea and testicular hyaluronidase, bFGF, or TGF-beta. Mitf expression was inhibited by these factors and reactivated during redifferentiation of the dedifferentiated cells into rPECs, showing the correlation between Mitf expression and rPEC differentiation. Retrovirus-mediated overexpression of Mtif inhibited bFGF-induced dedifferentiation and transdifferentiation of rPECs to both lens and neural cells. These findings showed that downregulation of Mitf expression is essential for the transdifferentiation of rPEC. Mitf overexpression caused hyperpigmentation in cultured rPECs and suppressed the changes in gene expression induced by bFGF. Mitf overexpression promoted expression of mmp115 and tyrosinase in bFGF-treated rPECs suggesting a critical role for Mitf in rPEC differentiation. Mitf overexpression, however, did not promote expression of another rPEC-specific gene, pP344, in bFGF-treated rPECs. This result suggests the presence of other regulatory genes promoting rPEC differentiation. The expression patterns of pax6 and Mitf are complementary both in vivo in vitro. Overexpression of Mitf inhibited expression of pax6 in cultured rPECs. These observations suggest that Mitf regulates pax6 expression negatively.

Amino Acid Sequence↗

PubMed1998

Pharmacokinetics of CPT-11 in rhesus monkeys.

PURPOSE: To examine the pharmacokinetic relationships between humans and monkeys, we studied the disposition of 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11) and its active metabolite, 7-ethyl-10-hydroxycamptothecin (SN-38), in rhesus monkeys. METHODS: CPT-11 was administered to a total of six monkeys at doses of 3, 7, 15 and 25 mg/kg by intravenous infusion for 10 min and plasma concentrations and pharmacokinetic parameters of CPT-11 determined. RESULTS: Maximum plasma concentrations at 25 mg/kg reached around 10000 ng/ml, and dropped to 500 ng/ml in 8 h. Plasma concentrations of SN-38 remained between 2 and 10 ng/ml. Mean values of systemic clearance, mean residence time and distribution volume at steady state, the major pharmacokinetic parameters for CPT-11, were 13.3 (ml/min per kg), 192 (min) and 2553 (ml/kg), respectively. The initial plasma concentration ratio of lactone to total CPT-11, 76%, declined to about 20% within 75 min, and the final ratio was about 40% at 8 h; the initial ratio of SN-38 was 72%, dropped to 34% within 70 min and finally recovered to 55% at 8 h. CONCLUSION: Comparison with human data revealed that systemic clearances of CPT-11 and the maximum AUC of SN-38 were not as different between humans and monkeys as between humans and mice, but the metabolic conversion of CPT-11 into SN-38 in monkeys was significantly lower than in humans.

Animals↗

PubMed1998

Distribution of a metabotropic glutamate receptor, mGluR2, in the central nervous system of the rat and mouse: an immunohistochemical study with a monoclonal antibody.

The distribution of a metabotropic glutamate receptor mGluR2 in the central nervous system was immunohistochemically examined in the rat and mouse with a monoclonal antibody raised against an N-terminal sequence of rat mGluR2 (amino acid residues 87-134). Neuronal cell bodies with mGluR2-like immunoreactivity (mGluR2-LI) were clearly shown in the horizontal cells of Cajal in the cerebral cortex, neurons in the triangular septal nucleus and medial mammillary nucleus, Golgi cells and the unipolar brush cells in the cerebellar cortex, and Golgi-like and unipolar brush-like cells in the cochlear nucleus. Neuropil was intensely immunostained in the accessory olfactory bulb, bed nucleus of the accessory olfactory tract, neocortex, cingulate cortex, retrosplenial cortex, subicular and entorhinal cortices, stratum lacunosum-moleculare of CA1 and CA3, molecular layer of the dentate gyrus, periamygdaloid cortex, basolateral amygdaloid nucleus, bed nucleus of the anterior commissure, caudate-putamen, accumbens nucleus, thalamic reticular nucleus, anteroventral and paraventricular thalamic nuclei, granular layer of the cerebellar cortex, anterior and ventral tegmental nuclei, granular layer of the cochlear nucleus, and parvicellular part of the lateral reticular nucleus. Many axons in the white matter and fiber bundles were also immunostained. No glial cells with mGluR2-LI were found. No particular species differences were found in the distribution pattern of mGluR2-LI between the rat and mouse. The results indicate that mGluR2 is expressed not only in somato-dendritic domain, but also in axonal domain of excitatory and inhibitory neurons.

Animals↗

PubMed1998

Substance P receptor (NK1)-immunoreactive neurons projecting to the periaqueductal gray: distribution in the spinal trigeminal nucleus and the spinal cord of the rat.

Substance P receptor (SPR)-immunoreactive neurons projecting to the periaqueductal gray (PAG) were examined in the rat spinal trigeminal nucleus and spinal cord by a retrograde tracing method combined with immunofluorescence histochemistry. After injection of Fluoro-gold (FG) into the PAG, SPR-immunoreactive neurons labeled with FG were observed mainly in the lateral spinal nucleus and lamina I of the medullary and spinal dorsal horns and additionally in laminae V and X of the spinal cord.

Animals↗

PubMed1998

Direct projections from the entorhinal cortical layers to the dentate gyrus, hippocampus, and subicular complex in the cat.

Projections from each layer of the entorhinal cortex (EC) of the cat were traced to the dentate gyrus (DG), Ammon's horn (CA), prosubiculum (ProSb), subiculum (Sb), presubiculum (PreSb) and parasubiculum (ParaSb); the anterograde or retrograde labeling method was used after stereotaxic injection of wheat germ agglutinin-horseradish peroxidase, cholera toxin B subunit, or Phaseolus vulgaris leucoagglutinin. On the side ipsilateral to the tracer-injection, layer II of the EC projected most abundantly to the outer half of the molecular layer (ML) of the DG, less abundantly to the almost entire thickness of the stratum lacunosum-moleculare (SLM) of CA2-3, moderately to the almost entire thickness of the SLM of CA1, and less to the outer part of the ML of the ProSb and Sb. Layer III projected abundantly to the almost entire thickness of the SLM of CA1 and outer part of the ML of the ProSb and Sb, and sparsely to the SLM of CA2-3. Layer IV projected sparsely to the pyramidal cell layer of the ProSb and Sb; Layer IV of the medial part (toward the ParaSb) of the EC projected further to the ML of the DG. Layer VI projected sparsely to the outer part of the ML of the DG, almost entire thickness of the SLM of CA1-3, and outer part of the ML of the ProSb and Sb. More temporal parts of the hippocampal region received the projections from progressively more medial and more rostral parts of layers II and III, and from progressively more rostral parts of layers IV and VI. The ML of the PreSb and ParaSb received projections from all layers of the medial part of the ipsilateral EC. The SLM of CA1 and ML of the ProSb, Sb and ParaSb received projections from layer II and/or III of the contralateral medial entorhinal area.

Animals↗

PubMed1998

Importance of hepatic first-pass removal in metastasis of colon carcinoma cells.

BACKGROUND/AIMS: It might be thought that colon carcinoma tends to metastasize to the liver because tumor cells leaving the primary colon tumor pass initially through the liver. Therefore we elucidated the kinetics of tumor cells in the body in order to understand the effect of the location of the liver on hepatic metastasis, that is to examine the hepatic first-pass effect of tumor cells. METHODS: Based on a physiological kinetic model, we examined quantitatively the hepatic metastasis and hepatic distribution of KM12-H1X cells administered by different routes. RESULTS: Both the number and incidence of colonies of hepatic metastasis were much greater after intrasplenic injection than after intravenous injection. The distribution of radioactivity to the liver after intrasplenic injection of [3H] thymidine-labeled cells was also much higher than that after intravenous injection. The number of colonies of hepatic metastasis correlated well with the area under the curve of the distributed amount of the tumor cells in the liver, regardless of the injection route; the correlation line was identical for each injection route. CONCLUSIONS: These results suggest that the hepatic first-pass effect is an important factor for the hepatic metastasis and that the cumulative number of tumor cells distributed in the liver is a determining factor for the degree of metastasis. Mathematical analysis based on a physiological model also suggests that hepatic metastasis depends on hepatic first-pass trapping of tumor cells.

Animals↗

PubMed1998

Mechanism of initial distribution of blood-borne colon carcinoma cells in the liver.

BACKGROUND/AIMS/METHODS: The distribution characteristics of a human colon carcinoma cell line, KM12-HX cells, were examined. After intraportal vein (i.p.v.) or intravenous (i.v.) injection into rats, almost all the injected tumor cells are distributed to liver or lung, respectively, both after 30 s and 30 min. Our previous kinetic analysis of the fate of tumor cells revealed that the cumulative amount of tumor cells distributed in the liver is a factor determining the degree of metastasis. Thus, we examined the mechanism of initial efficient trapping of tumor cells by the liver in more detail. RESULTS: Thirty minutes after tumor cells were injected into the left ventricle of the heart, the distribution of tumor cells was more restricted in several tissues (kidney, small intestine, large intestine and spleen), as compared with the distribution of microspheres undergoing 100% extraction, indicating that the first-pass extraction of KM12-HX cells is incomplete in these organs. The hepatic first-pass distribution of these tumor cells was unaffected by pretreatment of liposomes, such that the preinjected amount was sufficient to saturate the phagocytotic function of macrophages. Thus, the mechanism of initial distribution of the tumor cells to the liver is different from the mechanism of liposome uptake by macrophages. Considering that the diameter of microvessels in sinusoid and KM12-HX cells is approximately 7 and 12 microm, respectively, it is possible that these tumor cells are trapped physically in hepatic microvessels. In fact, after i.p.v. injection of microspheres 5 microm in diameter, only 20% of the dose was distributed to liver and the rest to other tissues. In contrast, almost 100% of microspheres 10 microm in diameter were distributed to the liver. CONCLUSIONS: These results support the hypothesis that the initial organ distribution of blood-borne tumor cells is determined by mechanical and physical properties of the cells.

Animals↗

PubMed1998

Characterization of neocortical non-pyramidal neurons expressing preprotachykinins A and B: a double immunofluorescence study in the rat.

Neurons expressing preprotachykinin A and preprotachykinin B, which are the precursor prepropeptides of substance P and neurokinin B (neuromedin K), respectively, were characterized immunocytochemically in the rat neocortex. Antibodies raised against C-terminal portions of preprotachykinins were used for labeling cell bodies of preprotachykinin-producing neurons. Neurons immunoreactive for preprotachykinin B were encountered four times more frequently in the neocortex than those immunoreactive for preprotachykinin A. Preprotachykinin A-immunoreactive neurons were scattered more frequently in the deep cortical layers (layers IV-VI) than in the superficial layers (layers I-III), whereas preprotachykinin B-immunoreactive neurons were distributed more frequently in the superficial layers than in the deep layers. Almost all preprotachykinin-expressing neurons were immunoreactive for GABA, suggesting that they were non-pyramidal cells. However, co-expression of the two preprotachykinin immunoreactivities in single neurons was not found. Preprotachykinin-expressing neocortical neurons were further examined with markers for subpopulations of GABAergic cortical neurons. Immunoreactivities for parvalbumin, calbindin and somatostatin were found in 69%, 27% and 11%, respectively, of preprotachykinin A-immunoreactive neurons. Conversely, preprotachykinin A-immunoreactive neurons constituted only 6% of parvalbumin-immunoreactive neurons, 4% of calbindin-immunoreactive neurons and 1% of somatostatin-immunoreactive neurons. Immunoreactivities for calretinin, choline acetyltransferase, vasoactive intestinal polypeptide, corticotropin-releasing factor and cholecystokinin were detected in 13-39% of preprotachykinin B-immunoreactive neurons. Preprotachykinin B immunoreactivity was seen in 33% of calretinin-positive neurons, 45% of cholinergic neurons, 47% of vasoactive intestinal polypeptide-positive neurons, 59% of corticotropin-releasing factor-positive neurons and 83% of cholecystokinin-positive neurons. These results indicate that preprotachykinin A- and preprotachykinin B-expressing neurons constitute separate populations of GABAergic non-pyramidal neurons in the neocortex. Since receptors for substance P and neurokinin B are expressed in GABAergic neurons [Kaneko T. et al. (1994) Neuroscience 60, 199-211] and pyramidal neurons [Ding Y. Q. et al. (1996) J. comp. Neurol. 364, 290-310], respectively, cortical neurons may use two separate lines of tachykinin signals; substance P serves as a signal between GABAergic non-pyramidal neurons, whereas neurokinin B acts as a signal of GABAergic neurons to pyramidal neurons.

Amino Acid Sequence↗

PubMed1998

A critical role for the optic vesicle in lens development; a reinvestigation of free lens formation in Cynops pyrrhogaster.

The role of the optic vesicle in lens development was reinvestigated in Cynops pyrrhogaster. To study the necessity for the optic vesicle in early lens development, the optic anlages of stage 17-27 embryos were ablated and the frequency of free lens formation was examined with lens specific markers. Free lens formation was not observed when operations were performed prior to contact between the head surface epidermis and the optic vesicle (stages 17-18). On the contrary, free lens formation occurred in all cases where the optic vesicles were removed after the initiation of lens placode formation in the head surface epidermis (stage 27). However, no lens fiber formation was observed in these free lenses as judged by the absence of lens fiber specific gene expression, namely gamma-crystallin, at stages when secondary lens fiber formation could be found in the control lenses of the unoperated sides. The pattern of expression of alpha A-crystallin in the developing free lens also differed from that of the normally developing lens. This paper is the first report to indicate that the coordinated and sequential expression of crystallin genes are influenced by the optic vesicle; the optic vesicle is required for proper regulation of the alpha A- and gamma-crystallin but not beta B1-crystallin genes.

Animals↗

PubMed1998

Beneficial effect of long-term combined treatment with voglibose and pioglitazone on pancreatic islet function of genetically diabetic GK rats.

Effects of voglibose (an alpha-glucosidase inhibitor) and pioglitazone (an insulin sensitizer) on glycemic control and on the function of pancreatic islets were evaluated using Goto-Kakizaki (GK) rats with non-insulin-dependent diabetes mellitus (NIDDM). Five week administration (8-13 weeks of age in GK rats) of voglibose alone (added to the chow at a concentration of 10 ppm), pioglitazone alone (10 mg/kg daily p.o.), or both of the agents together significantly improved fasting plasma glucose levels and those at 120 min in oral glucose tolerance tests. Insulin secretory capacity in response to glucose of the isolated islets, assessed by batch incubation, was significantly improved in the voglibose and in the voglibose plus pioglitazone groups. Eight-week administration (5-13 weeks of age) of voglibose and voglibose plus pioglitazone successfully lowered the fasting levels of plasma glucose and triglyceride. The glucose-responsiveness in insulin release from the islets was also significantly recovered by the therapy. The treatment increased the insulin content of the islets to almost twice that in untreated controls. Thus, treatment by these drugs can not only effectively ameliorate the metabolic derangement of NIDDM in GK rats, but it can also restore the deteriorated islet function, possibly through protection from glucose toxicity.

Animals↗

PubMed1998

Effect of cyclosporine A on the gastrointestinal absorption of acyclovir in neonatal and adult rats.

The effect of cyclosporine A (CyA) on the pharmacokinetics of acyclovir in neonatal and adult rats was studied. CyA at 25 and 50 mg/kg for 2-week-old and adult rats, respectively, given as a subcutaneous injection, reduced growth of the 2-week-old rats and inhibited growth of adult rats. The plasma concentration of acyclovir after intravenous administration (20 mg/kg) to neonatal CyA-treated rats increased and the total body clearance decreased compared with the neonatal controls. The bioavailabilities of acyclovir for neonatal control and CyA-treated rats after oral administration were significantly different at 15.6 and 22.0%, respectively, but those for the adult control and CyA-treated rats were the same at 15.6 and 14.0%, respectively. Experiments using the everted sac method showed that the amount of acyclovir transferred was higher in neonatal CyA-treated rats than the controls, but there was no difference in adult rats. A good relationship was observed between the bioavailabilities (in vivo) and cumulative transferred amounts (in vitro) in CyA-treated and control rats. Lactase activity in the brush border membrane of the intestine in the neonatal CyA-treated rats was significantly higher than in the controls. These results suggest that the gastrointestinal maturation of CyA-treated neonates is suppressed, resulting in increased bioavailability of acyclovir, while the gastrointestinal absorption of acyclovir does not differ between adult CyA-treated and control rats.

Acyclovir↗

PubMed1998

The novel insulinotropic mechanism of pimobendan: direct enhancement of the exocytotic process of insulin secretory granules by increased Ca2+ sensitivity in beta-cells.

Pimobendan is a new class of inotropic drug that augments Ca2+ sensitivity and inhibits phosphodiesterase (PDE) activity in cardiomyocytes. To examine the insulinotropic effect of pimobendan in pancreatic beta-cells, which have an intracellular signaling mechanism similar to that of cardiomyocytes, we measured insulin release from rat isolated islets of Langerhans. Pimobendan augmented glucose-induced insulin release in a dose-dependent manner, but did not increase cAMP content in pancreatic islets, indicating that the PDE inhibitory effects may not be important in beta-cells. This agent increased the intracellular Ca2+ concentration ([Ca2+]i) in the presence of 30 mM K+, 16.7 mM glucose, and 200 microM diazoxide, but failed to enhance the 30 mM K+-evoked [Ca2+]i rise in the presence of 3.3 mM glucose. Insulin release evoked by 30 mM K+ in 3.3 mM glucose was augmented. Then, the direct effects of pimobendan on the Ca2+-sensitive exocytotic apparatus were examined using electrically permeabilized islets in which [Ca2+]i can be manipulated. Pimobendan (50 microM) significantly augmented insulin release at 0.32 microM Ca2+, and a lower threshold for Ca2+-induced insulin release was apparent in pimobendan-treated islets. Moreover, 1 microM KN93 (Ca2+/calmodulin-dependent protein kinase II inhibitor) significantly suppressed this augmentation. Pimobendan, therefore, enhances insulin release by directly sensitizing the intracellular Ca2+-sensitive exocytotic mechanism distal to the [Ca2+]i rise. In addition, Ca2+/calmodulin-dependent protein kinase II activation may at least in part be involved in this Ca2+ sensitization for exocytosis of insulin secretory granules.

Androstadienes↗

PubMed1998

Altered bcl-2 and bax expression and intracellular Ca2+ signaling in apoptosis of pancreatic cells and the impairment of glucose-induced insulin secretion.

Apoptosis is the process of cellular self-destruction, and genes such as bcl-2 and bax are known to inhibit and promote apoptosis, respectively. In this study, we show that apoptosis can be induced in pancreatic beta-cell lines, and we investigate the apoptotic pathways through the bcl-2 and bax genes and intracellular Ca2+. Serum deprivation induces apoptosis in the MIN6 and RINm5F pancreatic beta-cell lines, and alters the bcl-2 messenger RNA (mRNA) and protein. KCl, BayK, A23187, and ionomycin elicit an elevation of cytosolic/nuclear Ca2+, which, however, is insufficient to evoke apoptosis or to alter bcl-2 or bax mRNA expression in MIN6 cells. The extracellular Ca2+ chelators, EGTA and 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, tetrapotassium salt, hydrate, evoke apoptosis and also alter the ratio of bcl-2 to bax mRNA and protein concomitantly with the depletion of cytosolic/nuclear Ca2+. This indicates that there are at least two apoptotic pathways in pancreatic beta-cells: through serum deprivation and through a decrease in cytosolic/nuclear Ca2+. MIN6 cells exhibit reduced insulin secretion induced by glucose regardless of the molecular pathway of apoptosis. Apoptosis in pancreatic beta-cells, therefore, may be closely related to the impairment of insulin secretion in certain pathological conditions such as diabetes mellitus.

Animals↗