Use of diphenylhydantoin and diazoxide to investigate insulin secretory mechanisms

In the isolated, perfused rat pancreas, we contrasted effects of diphenylhydantoin (DPH) and diazoxide on glucose-induced biphasic insulin secretion. Either drug partially inhibited the first phase. However, DPH completely inhibited the second phase, whereas diazoxide produced inhibition, then escape and post-inhibitory overshoot. Exposure to DPH prior to glucose further inhibited the first phase, and increasing the dose had no additional effects, whereas only raising the diazoxide dose intensified inhibition of early release. DPH sequentially suppressed early response to a series of two, short, glucose pulses. In contrast, no additional effects of diazoxide were noted after its initial inhibition of the first pulse. A computer analysis was programmed from hypotheses based on these experiments. It suggests that DPH inhibits release from a labile compartment and provision of insulin to that compartment, whereas diazoxide divides the labile compartment into two sequential subcompartments. Further, the computer analysis indicates that, with diazoxide, insulin (or substances on which secretion depends) accumulates not at the final release step but at a proximal portion of the labile compartment.

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Dynamics of glucose-induced insulin secretion in normal human islets

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00251.2015 ◽

2015 ◽

Vol 309 (7) ◽

pp. E640-E650 ◽

Cited By ~ 40

Author(s):

Jean-Claude Henquin ◽

Denis Dufrane ◽

Julie Kerr-Conte ◽

Myriam Nenquin

Keyword(s):

Insulin Secretion ◽

Stimulation Index ◽

Human Islets ◽

Diabetic Patients ◽

Second Phase ◽

Biphasic Insulin ◽

Biphasic Insulin Secretion ◽

Normal Human ◽

Second Phases

The biphasic pattern of glucose-induced insulin secretion is altered in type 2 diabetes. Impairment of the first phase is an early sign of β-cell dysfunction, but the underlying mechanisms are still unknown. Their identification through in vitro comparisons of islets from diabetic and control subjects requires characterization and quantification of the dynamics of insulin secretion by normal islets. When perifused normal human islets were stimulated with 15 mmol/l glucose (G15), the proinsulin/insulin ratio in secretory products rapidly and reversibly decreased (∼50%) and did not reaugment with time. Switching from prestimulatory G3 to G6–G30 induced biphasic insulin secretion with flat but sustained (2 h) second phases. Stimulation index reached 6.7- and 3.6-fold for the first and second phases induced by G10. Concentration dependency was similar for both phases, with half-maximal and maximal responses at G6.5 and G15, respectively. First-phase response to G15–G30 was diminished by short (30–60 min) prestimulation in G6 (vs. G3) and abolished by prestimulation in G8, whereas the second phase was unaffected. After 1–2 days of culture in G8 (instead of G5), islets were virtually unresponsive to G15. In both settings, a brief return to G3–G5 or transient omission of CaCl2 restored biphasic insulin secretion. Strikingly, tolbutamide and arginine evoked immediate insulin secretion in islets refractory to glucose. In conclusion, we quantitatively characterized the dynamics of glucose-induced insulin secretion in normal human islets and showed that slight elevation of prestimulatory glucose reversibly impairs the first phase, which supports the view that the similar impairment in type 2 diabetic patients might partially be a secondary phenomenon.

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Assessment of the Metabolic Pathways Associated With Glucose-Stimulated Biphasic Insulin Secretion

Endocrinology ◽

10.1210/en.2013-1805 ◽

2014 ◽

Vol 155 (5) ◽

pp. 1653-1666 ◽

Cited By ~ 23

Author(s):

Mei Huang ◽

Jamie W. Joseph

Keyword(s):

Insulin Secretion ◽

Time Course ◽

Tricarboxylic Acid Cycle ◽

Global Analysis ◽

Tricarboxylic Acid ◽

Second Phase ◽

Data Set ◽

Negatively Associated ◽

Biphasic Insulin ◽

Biphasic Insulin Secretion

Biphasic glucose-stimulated insulin secretion involves a rapid first phase followed by a prolonged second phase of insulin secretion. The biochemical pathways that control these 2 phases of insulin secretion are poorly defined. In this study, we used a gas chromatography mass spectroscopy-based metabolomics approach to perform a global analysis of cellular metabolism during biphasic insulin secretion. A time course metabolomic analysis of the clonal β-cell line 832/13 cells showed that glycolytic, tricarboxylic acid, pentose phosphate pathway, and several amino acids were strongly correlated to biphasic insulin secretion. Interestingly, first-phase insulin secretion was negatively associated with l-valine, trans-4-hydroxy-l-proline, trans-3-hydroxy-l-proline, dl-3-aminoisobutyric acid, l-glutamine, sarcosine, l-lysine, and thymine and positively with l-glutamic acid, flavin adenine dinucleotide, caprylic acid, uridine 5′-monophosphate, phosphoglycerate, myristic acid, capric acid, oleic acid, linoleic acid, and palmitoleic acid. Tricarboxylic acid cycle intermediates pyruvate, α-ketoglutarate, and succinate were positively associated with second-phase insulin secretion. Other metabolites such as myo-inositol, cholesterol, dl-3-aminobutyric acid, and l-norleucine were negatively associated metabolites with the second-phase of insulin secretion. These studies provide a detailed analysis of key metabolites that are either negatively or positively associated with biphasic insulin secretion. The insights provided by these data set create a framework for planning future studies in the assessment of the metabolic regulation of biphasic insulin secretion.

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Minimal modeling of insulin secretion in the perfused rat pancreas: a drug effect case study

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00603.2013 ◽

2014 ◽

Vol 306 (6) ◽

pp. E627-E634 ◽

Cited By ~ 8

Author(s):

Michela Riz ◽

Morten Gram Pedersen ◽

Gianna Maria Toffolo ◽

Guido Haschke ◽

Hans-Christoph Schneider ◽

...

Keyword(s):

Insulin Secretion ◽

Minimal Model ◽

Cell Function ◽

Fold Increase ◽

Second Phase ◽

Good Precision ◽

Β Cell ◽

Perfused Rat Pancreas ◽

Rat Pancreas ◽

Β Cell Function

The experimental protocol of the perfused rat pancreas is commonly used to evaluate β-cell function. In this context, mathematical models become useful tools through the determination of indexes that allow the assessment of β-cell function in different experimental groups and the quantification of the effects of antidiabetic drugs, secretagogues, or treatments. However, a minimal model applicable to the isolated perfused rat pancreas has so far been unavailable. In this work, we adapt the C-peptide minimal model applied previously to the intravenous glucose tolerance test to obtain a specific model for the experimental settings of the perfused pancreas. Using the model, it is possible to estimate indexes describing β-cell responsivity for first (ΦD) and second phase (ΦS, T) of insulin secretion. The model was initially applied to untreated pancreata and afterward used for the assessment of pharmacologically relevant agents (the gut hormone GLP-1, the potent GLP-1 receptor agonist lixisenatide, and a GPR40/FFAR1 agonist, SAR1) to quantify and differentiate their effect on insulin secretion. Model fit was satisfactory, and parameters were estimated with good precision for both untreated and treated pancreata. Model application showed that lixisenatide reaches improvement of β-cell function similarly to GLP-1 (11.7- vs. 13.1-fold increase in ΦD and 2.3- vs. 2.8-fold increase in ΦS) and demonstrated that SAR1 leads to an additional improvement of β-cell function in the presence of postprandial GLP-1 levels.

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Calcium signaling and secretory granule pool dynamics underlie biphasic insulin secretion and its amplification by glucose: experiments and modeling

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00380.2018 ◽

2019 ◽

Vol 316 (3) ◽

pp. E475-E486 ◽

Cited By ~ 4

Author(s):

Morten Gram Pedersen ◽

Alessia Tagliavini ◽

Jean-Claude Henquin

Keyword(s):

Insulin Secretion ◽

Glucose Concentration ◽

Calcium Influx ◽

Secretory Granules ◽

Cytosolic Calcium ◽

Second Phase ◽

Calcium Dynamics ◽

Β Cells ◽

Biphasic Insulin ◽

Biphasic Insulin Secretion

Glucose-stimulated insulin secretion from pancreatic β-cells is controlled by a triggering pathway that culminates in calcium influx and regulated exocytosis of secretory granules, and by a less understood amplifying pathway that augments calcium-induced exocytosis. In response to an abrupt increase in glucose concentration, insulin secretion exhibits a first peak followed by a lower sustained second phase. This biphasic secretion pattern is disturbed in diabetes. It has been attributed to depletion and subsequent refilling of a readily releasable pool of granules or to the phasic cytosolic calcium dynamics induced by glucose. Here, we apply mathematical modeling to experimental data from mouse islets to investigate how calcium and granule pool dynamics interact to control dynamic insulin secretion. Experimental calcium traces are used as inputs in three increasingly complex models of pool dynamics, which are fitted to insulin secretory patterns obtained using a set of protocols of glucose and tolbutamide stimulation. New calcium and secretion data for so-called staircase protocols, in which the glucose concentration is progressively increased, are presented. These data can be reproduced without assuming any heterogeneity in the model, in contrast to previous modeling, because of nontrivial calcium dynamics. We find that amplification by glucose can be explained by increased mobilization and priming of granules. Overall, our results indicate that calcium dynamics contribute substantially to shaping insulin secretion kinetics, which implies that better insight into the events creating phasic calcium changes in human β-cells is needed to understand the cellular mechanisms that disturb biphasic insulin secretion in diabetes.

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The Cell Physiology of Biphasic Insulin Secretion

Physiology ◽

10.1152/physiologyonline.2000.15.2.72 ◽

2000 ◽

Vol 15 (2) ◽

pp. 72-77 ◽

Cited By ~ 29

Author(s):

Patrik Rorsman ◽

Lena Eliasson ◽

Erik Renström ◽

Jesper Gromada ◽

Sebastian Barg ◽

...

Keyword(s):

Insulin Secretion ◽

Secretory Granules ◽

Diabetes Type ◽

Cell Physiology ◽

Second Phase ◽

Type Ii ◽

Diabetes Type Ii ◽

Biphasic Insulin ◽

Biphasic Insulin Secretion ◽

Glucose Stimulated Insulin Secretion

Glucose-stimulated insulin secretion consists of a transient first phase followed by a sustained second phase. Diabetes (type II) is associated with abnormalities in this release pattern. Here we review the evidence that biphasic insulin secretion reflects exocytosis of two functional subsets of secretory granules and the implications for diabetes.

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Glucose and cyclic AMP as stimulators of somatostatin and insulin secretion from the isolated, perfused rat pancreas: a quantitative study

Diabetes ◽

10.2337/diabetes.30.1.40 ◽

1981 ◽

Vol 30 (1) ◽

pp. 40-44 ◽

Cited By ~ 5

Author(s):

P. P. Gerber ◽

E. R. Trimble ◽

C. B. Wollheim ◽

A. E. Renold ◽

R. E. Miller

Keyword(s):

Insulin Secretion ◽

Cyclic Amp ◽

Quantitative Study ◽

Perfused Rat Pancreas ◽

Rat Pancreas ◽

Isolated Perfused Rat Pancreas

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Effect of somatostatin-28 on dynamics of insulin secretion in perfused rat pancreas

Diabetes ◽

10.2337/diabetes.40.9.1163 ◽

1991 ◽

Vol 40 (9) ◽

pp. 1163-1169 ◽

Cited By ~ 5

Author(s):

J. W. Ensinck ◽

E. C. Laschansky ◽

R. E. Vogel ◽

D. A. D'Alessio

Keyword(s):

Insulin Secretion ◽

Perfused Rat Pancreas ◽

Rat Pancreas ◽

Dynamics Of Insulin Secretion

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Demonstration that the vitamin D metabolite 1,25(OH)2-vitamin D3 and not 24R,25(OH)2-vitamin D3 is essential for normal insulin secretion in the perfused rat pancreas

Diabetes ◽

10.2337/diabetes.34.4.315 ◽

1985 ◽

Vol 34 (4) ◽

pp. 315-320 ◽

Cited By ~ 13

Author(s):

S. Kadowaki ◽

A. W. Norman

Keyword(s):

Vitamin D ◽

Insulin Secretion ◽

Vitamin D3 ◽

Perfused Rat Pancreas ◽

Rat Pancreas

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Effects of PHI on hormonal secretion from perfused rat pancreas

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1983.245.4.e313 ◽

1983 ◽

Vol 245 (4) ◽

pp. E313-E317

Author(s):

J. Szecowka ◽

D. Tendler ◽

S. Efendic

Keyword(s):

Amino Acids ◽

Insulin Secretion ◽

Glucagon Release ◽

The Novel ◽

Perfused Rat Pancreas ◽

Rat Pancreas ◽

Isolated Perfused Rat Pancreas ◽

Hormonal Secretion ◽

D Cells

Effects of the novel gastrointestinal polypeptide PHI with N-terminal histidine, C-terminal isoleucine amide, and 27 amino acids have been studied in isolated perfused rat pancreas. PHI increased the release of insulin, glucagon, and somatostatin. The amounts of these hormones released were strictly dependent on the prevailing glucose concentrations. In the absence of glucose, PHI (1 nmol/liter) stimulated glucagon release. In the presence of 4.4 and 6.7 mmol/liter glucose, the same dose of this peptide stimulated insulin and somatostatin release. In the presence of 16.7 mmol/liter glucose, only insulin secretion was increased by PHI. When arginine was used as a secretagogue, PHI (10 nmol/liter) potentiated secretion of insulin, glucagon, and somatostatin. Thus, PHI may take part in the regulation of the function of the pancreatic A, B, and D cells.

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