Immaturity of insulin secretion by pancreatic islets isolated from one human neonate.

Human β-cells are functionally mature by the age of 1 year. The timeline and mechanisms of this maturation are unknown owing to the exceptional availability of testable tissue. Here, we report the first in vitro study of insulin secretion by islets from a 5-day-old newborn.

Pharmacological approach to understanding the control of insulin secretion in human islets.

Aims: To understand better the control of insulin secretion by human β cells and to identify similarities to and differences from rodent models.

Methods: Dynamic insulin secretion was measured in perifused human islets treated with pharmacological agents of known modes of action.

Results: Glucokinase activation (Ro28-1675) lowered the glucose threshold for stimulation of insulin secretion to 1 mmol/L (G1), augmented the response to G3-G5 but not to G8-G15, whereas tolbutamide remained active in G20, which indicates that not all K channels were closed by high glucose concentrations.

Dynamics and Regulation of Insulin Secretion in Pancreatic Islets from Normal Young Children.

Insulin secretion has only exceptionally been investigated in pancreatic islets from healthy young children. It remains unclear whether those islets behave like adult islets despite substantial differences in cellular composition and higher β-cell replication rates. Islets were isolated from 5 infants/toddlers (11-36 month-old) and perifused to characterize their dynamics of insulin secretion when subjected to various stimuli and inhibitors.

Sulphonylurea receptor-1, sulphonylureas and amplification of insulin secretion by Epac activation in β cells.

Amplification of insulin secretion by cyclic AMP involves activation of protein kinase A (PKA) and Epac2 in pancreatic β cells. Recent hypotheses suggest that sulphonylurea receptor-1 (SUR1), the regulatory subunit of ATP-sensitive potassium channels, is implicated in Epac2 effects and that direct activation of Epac2 by hypoglycaemic sulphonylureas contributes to the stimulation of insulin secretion by these drugs. In the present experiments, using islets from Sur1KO mice, we show that dibutyryl-cAMP and membrane-permeant selective activators of Epac or PKA normally amplify insulin secretion in β cells lacking SUR1.

Dynamics of glucose-induced insulin secretion in normal human islets.

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.

Human Insulinomas Show Distinct Patterns of Insulin Secretion In Vitro.

Insulinomas are β-cell tumors that cause hypoglycemia through inappropriate secretion of insulin. Characterization of the in vitro dynamics of insulin secretion by perifused fragments of 10 human insulinomas permitted their subdivision into three functional groups with similar insulin content. Group A (four patients with fasting and/or postprandial hypoglycemic episodes) showed qualitatively normal responses to glucose, leucine, diazoxide, tolbutamide, and extracellular CaCl2 omission or excess.

Activators of PKA and Epac distinctly influence insulin secretion and cytosolic Ca2+ in female mouse islets stimulated by glucose and tolbutamide.

Amplification of insulin secretion by cAMP is mediated by protein kinase A (PKA) and exchange protein directly activated by cAMP (Epac). Using selective activators, we determined how each effector influences the cytosolic free Ca(2+) concentration ([Ca(2+)]c) and insulin secretion in mouse islets. Alone PKA activator amplified glucose- and tolbutamide-induced insulin secretion, with a greater impact on second than first phase.

Congenital hyperinsulinism caused by hexokinase I expression or glucokinase-activating mutation in a subset of β-cells.

Congenital hyperinsulinism causes persistent hypoglycemia in neonates and infants. Most often, uncontrolled insulin secretion (IS) results from a lack of functional K(ATP) channels in all β-cells or only in β-cells within a resectable focal lesion. In more rare cases, without K(ATP) channel mutations, hyperfunctional islets are confined within few lobules, whereas hypofunctional islets are present throughout the pancreas.

Amplification of insulin secretion by acetylcholine or phorbol ester is independent of β-cell microfilaments and distinct from metabolic amplification.

Insulin secretion (IS) triggered by β-cell [Ca(2+)](c) is amplified by metabolic and receptor-generated signals. Diacylglycerol largely mediates acetylcholine (ACh) effects through protein-kinase C and other effectors, which can be directly activated by phorbol-ester (PMA). Using mouse islets, we investigated the possible role of microfilaments in ACh/PMA-mediated amplification of IS.

cAMP-mediated and metabolic amplification of insulin secretion are distinct pathways sharing independence of β-cell microfilaments.

Insulin secretion is triggered by an increase in the cytosolic calcium concentration ([Ca(2+)](c)) in β-cells. Ca(2+)-induced exocytosis of insulin granules can be augmented by metabolic amplification (unknown signals generated through glucose metabolism) or neurohormonal amplification (in particular cAMP mediated). Functional actin microfilaments are not required for metabolic amplification, but their possible role in cAMP-mediated amplification is unknown.

Disruption and stabilization of β-cell actin microfilaments differently influence insulin secretion triggered by intracellular Ca2+ mobilization or store-operated Ca2+ entry.

Latrunculin depolymerizes and jasplakinolide polymerizes β-cell actin microfilaments. Both increase insulin secretion when Ca(2+) enters β-cells during depolarization by glucose, sulfonylureas or potassium. Mouse islets were held hyperpolarized with diazoxide, and stimulated with acetylcholine to test the role of microfilaments in insulin secretion triggered by intracellular Ca(2+) mobilization and store-operated Ca(2+) entry (SOCE).

In vitro insulin secretion by pancreatic tissue from infants with diazoxide-resistant congenital hyperinsulinism deviates from model predictions.

Congenital hyperinsulinism (CHI) is the major cause of persistent neonatal hypoglycemia. CHI most often occurs due to mutations in the ABCC8 (which encodes sulfonylurea receptor 1) or KCNJ11 (which encodes the potassium channel Kir6.2) gene, which result in a lack of functional KATP channels in pancreatic β cells.

Metabolic amplification of insulin secretion by glucose is independent of β-cell microtubules.

Glucose-induced insulin secretion (IS) by β-cells is controlled by two pathways. The triggering pathway involves ATP-sensitive potassium (K(ATP)) channel-dependent depolarization, Ca(2+) influx, and rise in the cytosolic Ca(2+) concentration ([Ca(2+)](c)), which triggers exocytosis of insulin granules. The metabolic amplifying pathway augments IS without further increasing [Ca(2+)](c).

Metabolic amplifying pathway increases both phases of insulin secretion independently of beta-cell actin microfilaments.

Two pathways control glucose-induced insulin secretion (IS) by beta-cells. The triggering pathway involves ATP-sensitive potassium (K(ATP)) channel-dependent depolarization, Ca(2+) influx, and a rise in the cytosolic Ca(2+) concentration ([Ca(2+)](c)), which triggers exocytosis of insulin granules. The metabolic amplifying pathway augments IS without further increasing [Ca(2+)](c).

Pharmacological stimulation and inhibition of insulin secretion in mouse islets lacking ATP-sensitive K+ channels.

Background And Purpose: ATP-sensitive potassium channels (K(ATP) channels) in beta cells are a major target for insulinotropic drugs. Here, we studied the effects of selected stimulatory and inhibitory pharmacological agents in islets lacking K(ATP) channels.

Experimental Approach: We compared insulin secretion (IS) and cytosolic calcium ([Ca(2+)](c)) changes in islets isolated from control mice and mice lacking sulphonylurea receptor1 (SUR1), and thus K(ATP) channels in their beta cells (Sur1KO).

Shortcomings of current models of glucose-induced insulin secretion.

Glucose-induced insulin secretion by pancreatic beta-cells is generally schematized by a 'consensus model' that involves the following sequence of events: acceleration of glucose metabolism, closure of ATP-sensitive potassium channels (K(ATP) channels) in the plasma membrane, depolarization, influx of Ca(2+) through voltage-dependent calcium channels and a rise in cytosolic-free Ca(2+) concentration that induces exocytosis of insulin-containing granules. This model adequately depicts the essential triggering pathway but is incomplete. In this article, we first make a case for a model of dual regulation in which a metabolic amplifying pathway is also activated by glucose and augments the secretory response to the triggering Ca(2+) signal under physiological conditions.

Glucose controls cytosolic Ca2+ and insulin secretion in mouse islets lacking adenosine triphosphate-sensitive K+ channels owing to a knockout of the pore-forming subunit Kir6.2.

Glucose-induced insulin secretion is classically attributed to the cooperation of an ATP-sensitive potassium (K ATP) channel-dependent Ca2+ influx with a subsequent increase of the cytosolic free Ca2+ concentration ([Ca2+]c) (triggering pathway) and a K ATP channel-independent augmentation of secretion without further increase of [Ca2+]c (amplifying pathway). Here, we characterized the effects of glucose in beta-cells lacking K ATP channels because of a knockout (KO) of the pore-forming subunit Kir6.2.

Insulin secretion in islets from mice with a double knockout for the dense core vesicle proteins islet antigen-2 (IA-2) and IA-2beta.

Islet antigen-2 (IA-2 or ICA 512) and IA-2beta (or phogrin) are major autoantigens in type 1 diabetes. They are located in dense core secretory vesicles including insulin granules, but their role in beta-cell function is unclear. Targeted disruption of either IA-2 or IA-2beta, or both, impaired glucose tolerance, an effect attributed to diminution of insulin secretion.

Glucose-induced cytosolic pH changes in beta-cells and insulin secretion are not causally related: studies in islets lacking the Na+/H+ exchangeR NHE1.

The contribution of Na(+)/H(+) exchange (achieved by NHE proteins) to the regulation of beta-cell cytosolic pH(c), and the role of pH(c) changes in glucose-induced insulin secretion are disputed and were examined here. Using real-time PCR, we identified plasmalemmal NHE1 and intracellular NHE7 as the two most abundant NHE isoforms in mouse islets. We, therefore, compared insulin secretion, cytosolic free Ca(2+) ([Ca(2+)](c)) and pH(c) in islets from normal mice and mice bearing an inactivating mutation of NHE1 (Slc9A1-swe/swe).

Nutrient control of insulin secretion in perifused adult pig islets.

Objectives: Xenotransplantation of pig islets is a potential solution to the shortage of human islets, but our knowledge of how these islets secrete insulin in response to nutrients is still fragmentary. This was the question addressed in the present study.

Methods: After 24 h culture adult pig islets were perifused to characterize the dynamics of insulin secretion.

Overnight culture unmasks glucose-induced insulin secretion in mouse islets lacking ATP-sensitive K+ channels by improving the triggering Ca2+ signal.

A current model ascribes glucose-induced insulin secretion to the interaction of a triggering pathway (K(ATP) channel-dependent Ca(2+) influx and rise in cytosolic [Ca(2+)](c)) and an amplifying pathway (K(ATP) channel-independent augmentation of secretion without further increase of [Ca(2+)](c)). However, several studies of sulfonylurea receptor 1 null mice (Sur1KO) failed to measure significant effects of glucose in their islets lacking K(ATP) channels. We addressed this issue that challenges the model.

Glucose stimulates Ca2+ influx and insulin secretion in 2-week-old beta-cells lacking ATP-sensitive K+ channels.

In adult beta-cells glucose-induced insulin secretion involves two mechanisms (a) a K(ATP) channel-dependent Ca(2+) influx and rise of cytosolic [Ca(2+)](c) and (b) a K(ATP) channel-independent amplification of secretion without further increase of [Ca(2+)](c). Mice lacking the high affinity sulfonylurea receptor (Sur1KO), and thus K(ATP) channels, have been developed as a model of congenital hyperinsulinism. Here, we compared [Ca(2+)](c) and insulin secretion in overnight cultured islets from 2-week-old normal and Sur1KO mice.

Nutrient control of insulin secretion in isolated normal human islets.

Pancreatic islets were isolated from 16 nondiabetic organ donors and, after culture for approximately 2 days in 5 mmol/l glucose, were perifused to characterize nutrient-induced insulin secretion in human islets. Stepwise increases from 0 to 30 mmol/l glucose (eight 30-min steps) evoked concentration-dependent insulin secretion with a threshold at 3-4 mmol/l glucose, K(m) at 6.5 mmol/l glucose, and V(max) at 15 mmol/l glucose.

In vivo and in vitro glucose-induced biphasic insulin secretion in the mouse: pattern and role of cytoplasmic Ca2+ and amplification signals in beta-cells.

The mechanisms underlying biphasic insulin secretion have not been completely elucidated. We compared the pattern of plasma insulin changes during hyperglycemic clamps in mice to that of glucose-induced insulin secretion and cytosolic calcium concentration ([Ca(2+)](c)) changes in perifused mouse islets. Anesthetized mice were infused with glucose to clamp blood glucose at 8.