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The Cross-Talk between GABA Signalling and Metabolic Hormones in the Brain and Pancreatic Islets
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiology.
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

GABA is a well-known neurotransmitter that can be synthesized in the central nervous system (CNS) and, interestingly, also in pancreatic islets. Once released, GABA activates GABA-A channels tonically or transiently resulting in different physiological functions. The pancreatic islets are important micro-organs composed of mainly α, β and δ cells secreting the metabolic hormones, namely insulin, glucagon and somatosatin, respectively. When insulin is secreted from pancreatic β cells, it can enter the blood and travel to the target tissues including the brain where the insulin receptor is prominently expressed such as in the hippocampus. It has been suggested that insulin regulates hippocampal function and, thereby, possibly modulates cognition. However, how this comes about is not understood. On the other hand, GABA secreted from the pancreatic β cells can regulate the islet cells via the para or autocrine loop. Nevertheless, in order to elucidate the details of GABA effects on cellular function, more insight into the pharmacological characteristics of GABA-A receptors, the physiological concentration of GABA and activation types of the GABA-receptors are required. We, therefore, used the whole-cell and single-channel patch-clamp technique to record from cells in the hippocampal slice and pancreatic islets for studying the function of GABA-A receptors and how they are modified by hormones, GABA or drugs. RT-qPCR was utilized to profile the expression of GABA-A receptors in the intact tissues. We also initiated the patch-clamp combined single-cell RT-PCR in the intact rat and human islets to investigate the cell-specific function of GABA-A receptors.

We have shown in acute rat hippocampal slices that 1 nM insulin “turns on” extrasynaptic GABA-A receptors in CA1 pyramidal neurons resulting in decreased frequency of action potential firing. The single-channel current amplitude is related to the GABA concentration resulting in a single-channel GABA affinity in the pM range. The benzodiazepines, flumazenil and zolpidem, are inverse agonists. The results demonstrated an unexpected hormonal control of the inhibitory channel subtype expressed and excitability of hippocampal neurons.

In the intact rat islets, the GABA-evoked tonic currents were present in the α cells and may contribute to keeping the resting membrane potential of α cells population at hyperpolarized membrane potential and, thereby, making it more difficult to depolarize the cells. In the human, the GABA signaling system was compromised in islets from type 2 diabetic individuals, where the expression of genes encoding the α1, α2, β2 and β3 GABA-A receptor subunits were down-regulated. GABA originating within the islets evoked tonic currents in the α, β and δ cells. However, transient current was observed only in δ cells, which implies a rapid regulation of somatostatin secretion by GABA. The effects of SR95531 on hormone release revealed that activation of GABA-A receptors decreased both insulin and glucagon secretion. The data is important for understanding the mechanism underlying GABA regulation of hormones secretion in human islets.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2013. , 40 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 947
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-209532ISBN: 978-91-554-8795-9 (print)OAI: oai:DiVA.org:uu-209532DiVA: diva2:658368
Public defence
2013-12-12, lecture hall A1:111a, BMC, Husargatan 3, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2013-11-21 Created: 2013-10-21 Last updated: 2014-01-23
List of papers
1. Insulin reduces neuronal excitability by turning on GABA(A) channels that generate tonic current
Open this publication in new window or tab >>Insulin reduces neuronal excitability by turning on GABA(A) channels that generate tonic current
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2011 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 6, no 1, e16188- p.Article in journal (Refereed) Published
Abstract [en]

Insulin signaling to the brain is important not only for metabolic homeostasis but also for higher brain functions such as cognition. GABA (γ-aminobutyric acid) decreases neuronal excitability by activating GABA(A) channels that generate phasic and tonic currents. The level of tonic inhibition in neurons varies. In the hippocampus, interneurons and dentate gyrus granule cells normally have significant tonic currents under basal conditions in contrast to the CA1 pyramidal neurons where it is minimal. Here we show in acute rat hippocampal slices that insulin (1 nM) "turns on" new extrasynaptic GABA(A) channels in CA1 pyramidal neurons resulting in decreased frequency of action potential firing. The channels are activated by more than million times lower GABA concentrations than synaptic channels, generate tonic currents and show outward rectification. The single-channel current amplitude is related to the GABA concentration resulting in a single-channel GABA affinity (EC(50)) in intact CA1 neurons of 17 pM with the maximal current amplitude reached with 1 nM GABA. They are inhibited by GABA(A) antagonists but have novel pharmacology as the benzodiazepine flumazenil and zolpidem are inverse agonists. The results show that tonic rather than synaptic conductances regulate basal neuronal excitability when significant tonic conductance is expressed and demonstrate an unexpected hormonal control of the inhibitory channel subtypes and excitability of hippocampal neurons. The insulin-induced new channels provide a specific target for rescuing cognition in health and disease.

National Category
Medical and Health Sciences
Research subject
Physiology
Identifiers
urn:nbn:se:uu:diva-148240 (URN)10.1371/journal.pone.0016188 (DOI)000286516500036 ()21264261 (PubMedID)
Available from: 2011-03-03 Created: 2011-03-03 Last updated: 2017-12-11Bibliographically approved
2. In Intact Islets Interstitial GABA Activates GABA(A) Receptors That Generate Tonic Currents in alpha-Cells
Open this publication in new window or tab >>In Intact Islets Interstitial GABA Activates GABA(A) Receptors That Generate Tonic Currents in alpha-Cells
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2013 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 6, e67228- p.Article in journal (Refereed) Published
Abstract [en]

In the rat islets γ-aminobutyric acid (GABA) is produced by the β-cells and, at least, the α-cells express the GABAA receptors (GABAA channels). In this study, we examined in intact islets if the interstitial GABA activated the GABAA receptors. We used the patch-clamp technique to record whole-cell and single-channel currents and single-cell RT-PCR to identify the cell-type we recorded from, in the intact rat islets. We further identified which GABAA receptor subunits were expressed. We determined the cell-type of 43 cells we recorded from and of these 49%, 28% and 7% were α, β and δ-cells, respectively. In the remaining 16% of the cells, mRNA transcripts of more than one hormone gene were detected. The results show that in rat islets interstitial GABA activates tonic current in the α-cells but not in the β-cells. Seventeen different GABAA receptor subunits are expressed with high expression of α1, α2, α4, α6, β3, γ1, δ, ρ1, ρ2 and ρ3 subunits whereas no expression was detected for α5 or ε subunits. The abundance of the GABAA receptor subunits detected suggests that a number of GABAA receptor subtypes are formed in the islets. The single-channel and tonic currents were enhanced by pentobarbital and inhibited by the GABAA receptor antagonist SR-95531. The single-channel conductance ranged from 24 to 105 pS. Whether the single-channel conductance is related to subtypes of the GABAA receptor or variable interstitial GABA concentrations remains to be determined. Our results reveal that GABA is an extracellular signaling molecule in rat pancreatic islets and reaches concentration levels that activate GABAA receptors on the glucagon-releasing α-cells.

National Category
Physiology
Identifiers
urn:nbn:se:uu:diva-203228 (URN)10.1371/journal.pone.0067228 (DOI)000321738400122 ()23826240 (PubMedID)
Available from: 2013-07-05 Created: 2013-07-05 Last updated: 2017-12-06Bibliographically approved
3. gamma-Aminobutyric acid (GABA) signalling in human pancreatic islets is altered in type 2 diabetes
Open this publication in new window or tab >>gamma-Aminobutyric acid (GABA) signalling in human pancreatic islets is altered in type 2 diabetes
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2012 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 55, no 7, 1985-1994 p.Article in journal (Refereed) Published
Abstract [en]

gamma-Aminobutyric acid (GABA) is a signalling molecule in the interstitial space in pancreatic islets. We examined the expression and function of the GABA signalling system components in human pancreatic islets from normoglycaemic and type 2 diabetic individuals. Expression of GABA signalling system components was studied by microarray, quantitative PCR analysis, immunohistochemistry and patch-clamp experiments on cells in intact islets. Hormone release was measured from intact islets. The GABA signalling system was compromised in islets from type 2 diabetic individuals, where the expression of the genes encoding the alpha 1, alpha 2, beta 2 and beta 3 GABA(A) channel subunits was downregulated. GABA originating within the islets evoked tonic currents in the cells. The currents were enhanced by pentobarbital and inhibited by the GABA(A) receptor antagonist, SR95531. The effects of SR95531 on hormone release revealed that activation of GABA(A) channels (GABA(A) receptors) decreased both insulin and glucagon secretion. The GABA(B) receptor antagonist, CPG55845, increased insulin release in islets (16.7 mmol/l glucose) from normoglycaemic and type 2 diabetic individuals. Interstitial GABA activates GABA(A) channels and GABA(B) receptors and effectively modulates hormone release in islets from type 2 diabetic and normoglycaemic individuals.

Keyword
gamma-Aminobutyric acid, Gene expression, Human islets, Type 2 diabetes
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-178541 (URN)10.1007/s00125-012-2548-7 (DOI)000305215200017 ()
Note

De tre första författarna delar förstaförfattarskapet.

Available from: 2012-08-01 Created: 2012-07-31 Last updated: 2017-12-07Bibliographically approved
4. In a cell-type specific manner, high-affinity GABA-A receptors participate in autocrine and paracrine GABA signaling in human pancreatic islets
Open this publication in new window or tab >>In a cell-type specific manner, high-affinity GABA-A receptors participate in autocrine and paracrine GABA signaling in human pancreatic islets
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

γ-Aminobutyric acid (GABA), best known as the classical inhibitory neurotransmitter, is also produced and released by pancreatic islet cells. The hormone secreting α, β and δ- cells in human islets express GABA-A receptors that are activated by GABA. GABA signaling in the islets is thought to regulate hormone secretion but how it comes about is unclear. To-date the interstitial GABA concentration and cell-type specific GABA-A receptors have not been characterized. As a consequence, it is not clear how the interstitial GABA in the intact human islet regulates the specific cell-types. We have set- up single-cell RT-PCR combined whole-cell patch-clamp to investigate the functional role of GABA-A receptors in identified cell within intact human islets. GABA-activated tonic current is present in all α, β and δ-cells whereas only the δ-cells respond to GABA with large, transient currents. High-affinity GABA-A receptors activated with interstitial concentrations lower than 10 nM GABA are expressed in both α and β-cells. In the β- cells different subtypes of GABA-A receptors were identified based on single-channel kinetics, current-voltage relation and pharmacology. The data provides insight into the mechanisms underlying GABA regulation of different cell-types in intact human islet.

National Category
Medical and Health Sciences
Research subject
Physiology
Identifiers
urn:nbn:se:uu:diva-209530 (URN)
Available from: 2013-10-21 Created: 2013-10-21 Last updated: 2013-11-22Bibliographically approved

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