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Studies of cAMP and Ca2+ signaling in pancreatic islet cells
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
2018 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

The blood glucose-lowering and -elevating hormones insulin and glucagon are released from the pancreatic islet β- and α-cells, respectively. The intracellular messengers Ca2+ and cAMP have central roles in controlling the secretion of both hormones, but the underlying mechanisms are incompletely understood. A powerful approach to gain further insight is to study the messengers in individual cells within pancreatic islets, provided that each cell can be identified. To facilitate such studies, adenoviral vectors were generated for expression of fluorescent proteins controlled by the insulin and preproglucagon promoters, as well as the somatostatin and pancreatic polypeptide promoters that identify the other two major islet cell types, δ- and PP-cells. Recordings of cAMP and Ca2+ concentration changes with fluorescent reporters demonstrated that cells expressing identification markers responded as expected to well-known stimuli and modulators of the two messengers. Glucose-induced Ca2+ oscillations in β-cells were found to be synchronized with those in δ-cells, and two subpopulations of α-cells with different Ca2+ regulation by glucose were identified. Mouse and human β-cells responded to the insulinotropic hormones glucagon, GIP and GLP-1 with elevations of cAMP. Most α-cells reacted similarly to GIP, whereas only a subpopulation – larger among human than mouse α-cells - responded to glucagon and GLP-1. The GLP-1-receptor antagonist exendin-(9-39) suppressed both GLP-1- and glucagon-induced cAMP elevations in β-cells. Since exendin-(9-39) did not antagonize glucagon receptors, glucagon apparently activates GLP-1 receptors in β-cells. Even in the absence of glucagon/GLP-1, exendin-(9-39) reduced cAMP increases obtained by glucose stimulation or elevation of Ca2+. This effect was attributable to constitutive GLP-1-receptor activity rather than paracrine effects. Exendin-(9-39) also inhibited glucose-induced insulin release, highlighting the importance of cAMP formation in nutrient-stimulated secretion. Simultaneous recordings of cAMP and Ca2+ showed a complex and variable interrelationship between the messengers and the cAMP precursor ATP in β-cells. Depolarization-induced Ca2+ increases inhibited forskolin-, IBMX- and GLP-1-induced cAMP elevations. This cAMP lowering in part reflected suppression of the Ca2+-sensitive activity of adenylyl cyclases AC5 and 6, but also autocrine signaling induced by Ca2+-triggered exocytosis of insulin and adenine nucleotides, whose receptors activate phosphodiesterases and inhibit adenylyl cyclases, respectively.

sted, utgiver, år, opplag, sider
Uppsala: Acta Universitatis Upsaliensis, 2018. , s. 53
Serie
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1521
Emneord [en]
pancreatic islet, insulin, glucagon, somatostatin, pancreatic polypeptide, exendin-(9-39), GLP-1, GIP, ATP, adenosine, cAMP, Ca2+, β-cell, α-cell, δ-cell, PP-cell
HSV kategori
Identifikatorer
URN: urn:nbn:se:uu:diva-368612ISBN: 978-91-513-0526-4 (tryckt)OAI: oai:DiVA.org:uu-368612DiVA, id: diva2:1269609
Disputas
2019-02-12, B41, Biomedical Centre, Husargatan 3, Uppsala, 09:15 (engelsk)
Opponent
Veileder
Merknad

Ca2+ in the keywords is Ca2+, please help me to change it. Thanks!

Tilgjengelig fra: 2019-01-16 Laget: 2018-12-11 Sist oppdatert: 2019-01-21
Delarbeid
1. Fluorescent protein vectors for pancreatic islet cell identification in live-cell imaging
Åpne denne publikasjonen i ny fane eller vindu >>Fluorescent protein vectors for pancreatic islet cell identification in live-cell imaging
Vise andre…
2016 (engelsk)Inngår i: Pflügers Archiv: European Journal of Physiology, ISSN 0031-6768, E-ISSN 1432-2013, Vol. 468, nr 10, s. 1765-1777Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

The islets of Langerhans contain different types of endocrine cells, which are crucial for glucose homeostasis. beta- and alpha-cells that release insulin and glucagon, respectively, are most abundant, whereas somatostatin-producing delta-cells and particularly pancreatic polypeptide-releasing PP-cells are more scarce. Studies of islet cell function are hampered by difficulties to identify the different cell types, especially in live-cell imaging experiments when immunostaining is unsuitable. The aim of the present study was to create a set of vectors for fluorescent protein expression with cell-type-specific promoters and evaluate their applicability in functional islet imaging. We constructed six adenoviral vectors for expression of red and green fluorescent proteins controlled by the insulin, preproglucagon, somatostatin, or pancreatic polypeptide promoters. After transduction of mouse and human islets or dispersed islet cells, a majority of the fluorescent cells also immunostained for the appropriate hormone. Recordings of the sub-plasma membrane Ca2+ and cAMP concentrations with a fluorescent indicator and a protein biosensor, respectively, showed that labeled cells respond to glucose and other modulators of secretion and revealed a striking variability in Ca2+ signaling among alpha-cells. The measurements allowed comparison of the phase relationship of Ca2+ oscillations between different types of cells within intact islets. We conclude that the fluorescent protein vectors allow easy identification of specific islet cell types and can be used in live-cell imaging together with organic dyes and genetically encoded biosensors. This approach will facilitate studies of normal islet physiology and help to clarify molecular defects and disturbed cell interactions in diabetic islets.

Emneord
Islets, alpha-cell, beta-cell, delta-cell, PP-cell, Insulin, Glucagon, Somatostatin, Pancreatic polypeptide, Ca2+, cAMP
HSV kategori
Identifikatorer
urn:nbn:se:uu:diva-306746 (URN)10.1007/s00424-016-1864-z (DOI)000384425500011 ()27539300 (PubMedID)
Forskningsfinansiär
Swedish Research Council, 325-2012-6778, 55X-06240Swedish Diabetes AssociationNovo NordiskEXODIAB - Excellence of Diabetes Research in Sweden
Tilgjengelig fra: 2016-11-09 Laget: 2016-11-03 Sist oppdatert: 2018-12-11bibliografisk kontrollert
2. Constitutive GLP-1-receptor signaling contributes to basal and glucose-stimulated cAMP formation in β-cells: Constitutive GLP-1 receptor signaling in β-cells
Åpne denne publikasjonen i ny fane eller vindu >>Constitutive GLP-1-receptor signaling contributes to basal and glucose-stimulated cAMP formation in β-cells: Constitutive GLP-1 receptor signaling in β-cells
(engelsk)Inngår i: Artikkel i tidsskrift (Annet (populærvitenskap, debatt, mm)) Submitted
HSV kategori
Identifikatorer
urn:nbn:se:uu:diva-368723 (URN)
Tilgjengelig fra: 2018-12-11 Laget: 2018-12-11 Sist oppdatert: 2018-12-11
3. Effects of Ca2+ and autocrine signals on cAMP dynamics in β-cells
Åpne denne publikasjonen i ny fane eller vindu >>Effects of Ca2+ and autocrine signals on cAMP dynamics in β-cells
(engelsk)Manuskript (preprint) (Annet (populærvitenskap, debatt, mm))
HSV kategori
Identifikatorer
urn:nbn:se:uu:diva-368721 (URN)
Tilgjengelig fra: 2018-12-11 Laget: 2018-12-11 Sist oppdatert: 2018-12-11
4. Glucagon and GLP-1 evoke cAMP elevations in sub-populations of mouse and human α-cells
Åpne denne publikasjonen i ny fane eller vindu >>Glucagon and GLP-1 evoke cAMP elevations in sub-populations of mouse and human α-cells
(engelsk)Manuskript (preprint) (Annet (populærvitenskap, debatt, mm))
HSV kategori
Identifikatorer
urn:nbn:se:uu:diva-368722 (URN)
Tilgjengelig fra: 2018-12-11 Laget: 2018-12-11 Sist oppdatert: 2018-12-20

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