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Biomedical subjects

S Seino

Publications and source records attributed to S Seino.

At least 91 records · Page 5Linked to original sources
PubMed1996

Cellular distribution of the P2X4 ATP receptor mRNA in the brain and non-neuronal organs of rats.

RNA blot analysis has shown that an ionotropic ATP receptor P2X4 is widely distributed, as compared with other P2X subtypes. Cellular distribution of the P2X4 ATP receptor mRNA was investigated by in situ hybridization technique in the brain and non-neuronal organs. In the brain, transcripts of the P2X4 were widely distributed in various regions, with the highest signals in the cerebellar Purkinje cells and granule cells. In other organs, intense and selective expression was detected in seminiferous tubules of the testis and intestinal crypts of the colon, suggesting an involvement of ATP in spermatogenesis and intestinal secretion. A low but significant level of the P2X4 mRNA was detected in the adenohypophysis and the cortex of the adrenal gland. The tissue- or cell-specific expression of the P2X4 mRNA is of importance to understand the functional roles of the purinergic transmission system.

Adrenal Glands↗
PubMed1996

Recent progress in the molecular genetic aspects of non-insulin-dependent diabetes mellitus.

Non-insulin-dependent diabetes mellitus (NIDDM) is one of the most common metabolic and endocrine disorders in developed countries. NIDDM is a multifactorial disorder in which genetic factors play a complex role in its development. Until recently, little has been known of the genetic determinants of NIDDM. Advances in molecular biological and molecular genetic approaches have now made it possible to begin to identify the diabetes susceptibility genes in NIDDM. This review describes the progress of recent research in the molecular genetic aspects of NIDDM.

Age of Onset↗
PubMed1996

Identification of microsatellite markers near the human genes encoding the beta-cell ATP-sensitive K+ channel and linkage studies with NIDDM in Japanese.

ATP-sensitive K+ (KATP) channels play a key role in stimulus-secretion coupling in pancreatic beta-cells. Recent studies have shown that the beta-cell KATP channel comprises two subunits: a novel member of the inwardly rectifying K+ channel family, designated BIR and expressed at highest levels in pancreatic islets, and the sulfonylurea receptor (SUR). Moreover, the genes encoding these two proteins are adjacent to one another on human chromosome 11. Genetic factors contribute to the development of NIDDM, and it seems likely that mutations in genes encoding proteins involved in insulin secretion or action may contribute to NIDDM susceptibility. The present study examined the contribution of the linked BIR and SUR genes to the development of NIDDM. These genes were localized to the same yeast artificial chromosome as two microsatellite DNA polymorphisms, D11S902 and D11S921. These microsatellite DNA polymorphisms were typed in 140 Japanese NIDDM-affected sib pairs. There was no evidence for linkage between these markers and NIDDM, suggesting that genetic variation in the BIR and SUR genes does not play a major role in susceptibility to NIDDM in Japanese.

ATP-Binding Cassette Transporters↗
PubMed1995

Reconstitution of IKATP: an inward rectifier subunit plus the sulfonylurea receptor.

A member of the inwardly rectifying potassium channel family was cloned here. The channel, called BIR (Kir6.2), was expressed in large amounts in rat pancreatic islets and glucose-responsive insulin-secreting cell lines. Coexpression with the sulfonylurea receptor SUR reconstituted an inwardly rectifying potassium conductance of 76 picosiemens that was sensitive to adenosine triphosphate (ATP) (IKATP) and was inhibited by sulfonylureas and activated by diazoxide. The data indicate that these pancreatic beta cell potassium channels are a complex composed of at least two subunits--BIR, a member of the inward rectifier potassium channel family, and SUR, a member of the ATP-binding cassette superfamily. Gene mapping data show that these two potassium channel subunit genes are clustered on human chromosome 11 at position 11p15.1.

ATP-Binding Cassette Transporters↗
PubMed1995

cDNA sequence, gene structure, and chromosomal localization of the human ATP-sensitive potassium channel, uKATP-1, gene (KCNJ8).

ATP-sensitive K+ (KATP) channels play a crucial role in coupling metabolic energy to the membrane potential of cells. Recently, we have isolated a KATP channel cDNA (uKATP-1) that is expressed ubiquitously in rat tissues including pancreatic islets, pituitary, skeletal muscle, and heart. Here, we report cloning of the human cDNA and gene encoding uKATP-1. Human uKATP-1 is a protein of 424 amino acids exhibiting 98% identity with rat uKATP-1. The human gene encoding uKATP-1, designated KCNJ8, is approximately 9.7 kb in length and is composed of three exons. KCNJ8 was mapped to chromosome 12p11.23 using fluorescence in situ hybridization.

ATP-Binding Cassette Transporters↗
PubMed1995

Human gastric inhibitory polypeptide receptor: cloning of the gene (GIPR) and cDNA.

Gastric inhibitory polypeptide (GIP), which is released from the gastrointestinal tract, stimulates insulin secretion from pancreatic beta cells and plays a crucial role in the regulation of insulin secretion during the postprandial phase. We have isolated the human gene (GIPR) and cDNA encoding the GIP receptor by a combination of the conventional screening and polymerase chain reaction procedures. Human GIP receptor cDNA encodes a protein of 466 amino acids that is 81.5 and 81.2% identical to the previously cloned hamster and rat GIP receptor, respectively. Hydropathic analysis shows the presence of a signal peptide and seven potential transmembrane domains, a feature characteristic of the VIP/glucagon/secretin receptor family of G protein-coupled receptors. The human GIPR gene is about 13.8 kb long, consists of 14 exons, and carries 17 Alu repeats.

Amino Acid Sequence↗
PubMed1995

The structures of the human calcium channel alpha 1 subunit (CACNL1A2) and beta subunit (CACNLB3) genes.

Calcium influx in pancreatic beta-cells is regulated mainly by L-type voltage-dependent calcium channels (VDCCs) and triggers insulin secretion. The alpha 1 subunit (CACN4) and the beta subunit (beta 3) of VDCCs, both of which are expressed in pancreatic islets, are major components for the VDCC activity, and so they may play a critical role in the regulation of insulin secretion. We have determined the structures of the human CACN4 (CACNL1A2) and the human beta 3 (CACNLB3) genes. The CACNL1A2 gene spans more than 155 kb and has 49 exons. Most of the positions interrupted by introns are well conserved between the CACNL1A2 gene and the previously reported L-type VDCC alpha 1 subunit, CACNL1A1, gene. On the other hand, the CACNLB3 gene distributes in approximately 8 kb and comprises 13 exons, most of which are located together within approximately 5 kb. Comparisons of the genomic sequences of CACNL1A2 with the previously reported cDNA sequences indicate that there are a number of polymorphisms in the human CACNL1A2 gene. In addition, the PCR-SSCP procedure of exon 1 of CACNL1A2 revealed a change from 7 to 8 ATG trinucleotide repeats in a patient with non-insulin-dependent diabetes mellitus (NIDDM), resulting in an addition of methionine at the amino-terminus of CACN4. The determination of the structures of the human CACNL1A2 and CACNLB3 genes should facilitate study of the role of these genes in the development of NIDDM and also other genetic diseases such as long QT syndrome.

Base Sequence↗
PubMed1995

Cloning and sequencing of a human endothelin converting enzyme in renal adenocarcinoma (ACHN) cells producing endothelin-2.

Endothelin (ET)-2 is a 21 residue vasoactive peptide which is biosynthesized from big ET-2(1-38) by a specific cleavage at Trp21-Val22 with an ET converting enzyme (ECE). To identify an ECE in ACHN (human renal adenocarcinoma) cells which produce ET-2, we have cloned and sequenced a novel cDNA encoding a human ECE in ACHN (hAECE). It encodes a 770 amino acid protein with a zinc-binding motif and a single membrane spanning region. The sequences of nucleic acids and amino acids from Leu45 to Trp770 of hAECE are identical to those from Leu33 to Trp758 of a human ECE in HUVEC (hHECE). The sequences in the amino-terminal moiety are divergent between hAECE and hHECE. Based on the difference of the amino-terminal amino acid sequences, ECEs reported so far, can be classified into two isoforms. These results strongly suggest that an alternative splicing might occur in the 5'-terminal region of the ECE pre-mRNA.

Adenocarcinoma↗
PubMed1995

Cloning and functional characterization of a novel ATP-sensitive potassium channel ubiquitously expressed in rat tissues, including pancreatic islets, pituitary, skeletal muscle, and heart.

ATP-sensitive K+ (KATP) channels play a crucial role in coupling metabolic energy to the membrane potential of cells. We have isolated a cDNA encoding a novel member (uKATP-1) of the inward rectifier K+ channel family from a rat pancreatic islet cDNA library. Rat uKATP-1 is a 424-amino acid residue protein (M(r) = 47,960). Electrophysiological studies of uKATP-1 expressed in Xenopus laevis oocytes show that uKATP-1 is a weak rectifier and is blocked with Ba2+ ions. Single-channel patch clamp study of clonal human kidney epithelial cells (HEK293) transfected with uKATP-1 cDNA reveals that uKATP-1 closes in response to 1 mM ATP and has a single channel conductance of 70 +/- 2 picosiemens (n = 6), indicating that uKATP-1 is an ATP-sensitive inward rectifier K+ channel. In addition, uKATP-1 is activated by the KATP channel opener, diazoxide. RNA blot analysis shows that uKATP-1 mRNA is expressed ubiquitously in rat tissues, including pancreatic islets, pituitary, skeletal muscle, and heart, suggesting that uKATP-1 may play a physiological role as a link between the metabolic state and membrane K+ permeability of cells in almost every normal tissue. Since uKATP-1 shares only 43-46% amino acid identity with members of previously reported inward rectifier K+ channel subfamilies, including ROMK1, IRK1, GIRK1, and cKATP-1, uKATP-1 is not an isoform of these subfamilies and, therefore, represents a new subfamily of the inward rectifier K+ channel family having two transmembrane segments.

ATP-Binding Cassette Transporters↗
PubMed1995

Expression and complex formation of soluble N-ethyl-maleimide-sensitive factor attachment protein (SNAP) receptors in clonal rat endocrine cells.

Syntaxin, synaptosome-associated protein 25 (SNAP-25) and synaptobrevin are neuronal receptors of N-ethylmaleimide-sensitive factor (NSF) and soluble NSF attachment protein (SNAP), cytosolic proteins essential for membrane transport. The expression of these SNAP receptors was investigated by immunoblotting in clonal endocrine cells; catecholamine-secreting adrenal medullary cells (PC12), growth hormone-secreting pituitary cells (GH3), and insulin-secreting pancreatic beta-cells (RINr and RINm5F). All of these receptors as well as NSF and SNAP are expressed in these cells and, co-immunoprecipitation experiments indicate that syntaxin is associated with SNAP-25 and synaptobrevin after solubilization. These results suggest that syntaxin, SNAP-25, and synaptobrevin form a complex and are involved in the secretion of hormones from endocrine cells by exocytosis.

Amino Acid Sequence↗
PubMed1995

Inositol 1,4,5-trisphosphate receptors, secretory granules and secretion in endocrine and neuroendocrine cells.

Recent studies have revealed the presence of inositol 1,4,5-trisphosphate receptors in the secretory granules of endocrine and neuroendocrine cells. This distribution suggests that inositol 1,4,5-trisphosphate-regulated release of granule stores of Ca2+ might facilitate the secretory process. In addition, inositol 1,4,5-trisphosphate receptors might participate directly in the biogenesis of secretory granules. The presence of inositol 1,4,5-trisphosphate receptors in synaptic nerve terminals raises the possibility that they might also be involved in the control of neurotransmitter release.

Animals↗
PubMed1995

CACN4, the major alpha 1 subunit isoform of voltage-dependent calcium channels in pancreatic beta-cells: a minireview of current progress.

Calcium influx through L-type voltage-dependent calcium channels (VDCCs) triggers insulin secretion. Until recently, the structure of VDCCs in pancreatic beta-cells and their regulation in altered metabolic states were not known. Study of the VDCC protein in skeletal muscle has shown that the alpha 1 subunit is functionally the most important subunit among the five subunits (alpha 1, alpha 2, beta, gamma and delta), acting as a voltage sensor and an ion-conducting pore. Molecular cloning of a novel alpha 1 subunit (beta-cell/neuroendocrine type, CACN4) of VDCCs from pancreatic islets and insulinoma have made it possible to study the electrophysiological and pharmacogical properties, regulation, and genetics of the VDCCs expressed in beta-cells. The CACN4 is structurally related to other members of the VDCC alpha 1 subunit family, including skeletal muscle, cardiac, and brain types. In situ hybridization experiments reveal that CACN4 mRNAs are expressed in beta-cells in the islets. Heterologous expression studies show that CACN4 in the presence of the beta subunit elicits L-type VDCC currents, although expression of CACN4 alone is not sufficient for VDCC activity. Studies of animal models with chronic hyperglycemia and starvation have indicated that the reduced CACN4 mRNA levels in pancreatic islets are associated with impaired insulin responses to stimuli in both hyperglycemic and fasting states. These studies demonstrate that CACN4 is the major component of VDCCs in pancreatic beta-cells and suggest that it plays a crucial role in the regulation of insulin secretion in normal and altered metabolic states.

Animals↗
PubMed1995

Molecular diversity and functional characterization of voltage-dependent calcium channels (CACN4) expressed in pancreatic beta-cells.

Dihydropyridine-sensitive voltage-dependent calcium channels (VDCC) play a crucial role in insulin secretion. We recently have cloned a human alpha 1-subunit of the VDCC expressed in pancreatic beta-cells, designated CACN4. In this study we have isolated complementary DNAs encoding two forms of rat CACN4 (rCACN4A and rCACN4B) from a rat insulinoma RINm5F complementary DNA library. Rat CACN4A is a protein of 2203 amino acids and is the rat homolog of human CACN4, whereas rCACN4B lacks 535 amino acids in the carboxyl-terminal region, probably due to alternative splicing. We have found two additional variations, one in the intracellular loop between repeats I and II and the other in the extracellular region between the third and fourth segments of repeat IV. Reverse transcriptase-polymerase chain reaction analysis of rat pancreatic islet messenger RNA reveals that these variants are present in pancreatic islets. In addition, whole-cell voltage-clamp recordings of Chinese hamster ovary cells stably expressing the alpha 1-subunit (rCACN4A or rCACN4B) with or without the calcium channel beta 2-subunit show that coexpression of rCACN4A with the beta 2-subunit or rCACN4B with the beta 2-subunit elicits L-type VDCC currents, whereas expression of the alpha 1-subunit alone does not, indicating that CACN4 can associate functionally with the beta 2-subunit and that the beta-subunit is essential for functional expression of CACN4. These results suggest that there are various subtypes of CACN4 expressed in pancreatic beta-cells, and that both rCACN4A and rCACN4B can function as VDCC. Furthermore, the present study suggests that the expression of the beta-subunit as well as the alpha 1-subunit may participate in the regulation of insulin secretion.

Amino Acid Sequence↗
PubMed1995

A monitor of secretion from single pancreatic beta-cells.

ATP is known to be coreleased with insulin from pancreatic beta-cells. To monitor insulin secretion from single beta-cells, a single beta-cell was surrounded in culture by Fura 2-loaded calf pulmonary artery endothelium (CPAE) cells, which can detect the ATP. CPAE cells did not respond with an elevation in cytoplasmic calcium concentration ([Ca2+]i) to either tolbutamide (100 mumol/l) or kainate (1 mmol/l) but did respond with an elevation in [Ca2+]i to ATP (0.1-10 mumol/l) without desensitization and in a dose-dependent manner. A brief application of tolbutamide (10 mumol/l) increased [Ca2+]i in both the beta-cell and the adjacent CPAE cells in co-culture. Suramin (100 mumol/l), an ATP-receptor blocker, inhibited the tolbutamide-induced elevation in [Ca2+]i in the CPAE cells but did not inhibit the elevation in [Ca2+]i in the beta-cell, confirming that the insulin secretagogue-induced Ca2+ response in CPAE cells in co-culture is mediated by ATP released from the beta-cell. When co-culture of the beta-cell and CPAE cells was stimulated by kainate (1 mmol/l) and then tolbutamide (10 mumol/l), the CPAE cells showed elevations in [Ca2+]i in response to kainate and tolbutamide during elevation in [Ca2+]i in the beta-cell. This strongly suggests that insulin secretion as well as an increase in [Ca2+]i in response to different agents, i.e., kainate and tolbutamide, occurs in a single beta-cell. A long exposure of tolbutamide (100 mumol/l, 4 min) resulted in a long-lasting elevation in [Ca2+]i in the beta-cell.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1995

Cellular localization of synaptotagmin I, II, and III mRNAs in the central nervous system and pituitary and adrenal glands of the rat.

Three isoforms of synaptotagmin, a synaptic vesicle protein involved in neurotransmitter release, have been characterized in the rat, although functional differences between these isoforms have not been reported. In situ hybridization was used to define the localization of synaptotagmin I, II, and III transcripts in the rat CNS and pituitary and adrenal glands. Each of the three synaptotagmin genes has a unique expression pattern. The synaptotagmin III gene is expressed in most neurons, but transcripts are much less abundant than the products of the synaptotagmin I and II genes. A majority of neurons in the forebrain expressed both synaptotagmin I and III mRNAs while synaptotagmin II gene expression was confined to subsets of neurons in layers IV-VI of the cerebral cortex, in the dentate granule cell region, the hilus, and the CA1-CA3 areas of the hippocampus. In the cerebellum, all three transcripts were visualized in the granule cell layer. Furthermore, synaptotagmin I probes revealed striking differences between distinct populations of neurons, as in addition to moderate labeling of granule cells, much more prominent hybridization signals were detected on scattered cell bodies likely to be Golgi interneurons. In the most caudal part of the brain, synaptotagmin II transcripts were abundant and were coexpressed with synaptotagmin III mRNAs. This pattern was found in putative motoneurons of the spinal cord, suggesting that the two isoforms might be involved in exocytosis at the neuromuscular junction. Only synaptotagmin I mRNAs were detected in the anterior and intermediate pituitary and in adrenal medullary cells. These data reveal an unexpectedly subtle segregation of the expression of synaptotagmin genes and the existence of multiple combinations of synaptotagmin isoforms which may provide diversity in the regulation of neurosecretion.

Adrenal Glands↗
PubMed1995

Expression and role of ionotropic glutamate receptors in pancreatic islet cells.

Although the excitatory amino acid glutamate and its receptors play crucial roles in many functions of the central nervous system (CNS), their presence in the peripheral tissues has remained unclear. In the present study, we have identified kainate, alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA), and N-methyl-D-aspartate (NMDA) receptor subtype mRNAs in pancreatic islets, using reverse transcriptase polymerase chain reaction (RT-PCR). Intracellular calcium ([Ca2+]i) measurements and electrophysiological recordings indicate that kainate, AMPA, and NMDA all elicit increases of [Ca2+]i in single pancreatic beta-cells and depolarize them. In addition, kainate and AMPA stimulate insulin secretion from isolated pancreatic islets, whereas NMDA does not. Also, immunocytochemical study shows the presence of intense glutaminase immunoreactivity in pancreatic alpha-cells and intrapancreatic ganglia, a finding compatible with the possibility that glutamate is released from alpha-cells as well as from neurons. Because the inhibitory amino acid gamma-amino butyric acid (GABA) is present in beta-cells as well as in neurons and inhibits glucagon secretion from alpha-cells, the present study suggests that glutamate and GABA are coordinated in the regulation of hormone secretion in pancreatic islets.

Animals↗
PubMed1994

Hamster gastric inhibitory polypeptide receptor expressed in pancreatic islets and clonal insulin-secreting cells: its structure and functional properties.

We have isolated a cDNA encoding a gastric inhibitory polypeptide (GIP) receptor from a hamster insulinoma (HIT-T15) cDNA library. The hamster GIP receptor is a 462 amino acid protein having seven transmembrane segments. Expression of recombinant of hamster GIP receptors in Chinese hamster ovary (CHO) cells shows that it binds specifically to GIP with high affinity (IC50 = 9.6 nM) and is positively coupled to adenylate cyclase, but not to phospholipase C. RNA blot analysis reveals that a 3.8-kb GIP receptor mRNA is expressed at high levels in rat pancreatic islets as well as in HIT-T15 cells.

Amino Acid Sequence↗