Atrial natriuretic peptide (ANP) influences glucose homeostasis and possibly acts as a link between the cardiovascular system and metabolism, especially in metabolic disorders like diabetes. The current study evaluated effects of ANP on β-cell function by the use of a β-cell-specific knockout of the ANP receptor with guanylate cyclase activity (βGC-A-KO). ANP augmented insulin secretion at the threshold glucose concentration of 6 mmol/L and decreased K single-channel activity in β-cells of control mice but not of βGC-A-KO mice. In wild-type β-cells but not β-cells lacking functional K channels (SUR1-KO), ANP increased electrical activity, suggesting no involvement of other ion channels. At 6 mmol/L glucose, ANP readily elicited Ca influx in control β-cells. This effect was blunted in β-cells of βGC-A-KO mice, and the maximal cytosolic Ca concentration was lower. Experiments with inhibitors of protein kinase G (PKG), protein kinase A (PKA), phosphodiesterase 3B (PDE3B), and a membrane-permeable cyclic guanosine monophosphate (cGMP) analog on K channel activity and insulin secretion point to participation of the cGMP/PKG and cAMP/PKA/Epac (exchange protein directly activated by cAMP) directly activated by cAMP Epac pathways in the effects of ANP on β-cell function; the latter seems to prevail. Moreover, ANP potentiated the effect of glucagon-like peptide 1 (GLP-1) on glucose-induced insulin secretion, which could be caused by a cGMP-mediated inhibition of PDE3B, which in turn reduces cAMP degradation.

Edited by Norma M. AllewellNeuroendocrine-type ATP-sensitive K ؉ (K ATP ) channels are metabolite sensors coupling membrane potential with metabolism, thereby linking insulin secretion to plasma glucose levels. They are octameric complexes, (SUR1/Kir6.2) 4 , comprising sulfonylurea receptor 1 (SUR1 or ABCC8) and a K ؉ -selective inward rectifier (Kir6.2 or KCNJ11). Interactions between nucleotide-, agonist-, and antagonist-binding sites affect channel activity allosterically. Although it is hypothesized that opening these channels requires SUR1-mediated MgATP hydrolysis, we show here that ATP binding to SUR1, without hydrolysis, opens channels when nucleotide antagonism on Kir6.2 is minimized and SUR1 mutants with increased ATP affinities are used. We found that ATP binding is sufficient to switch SUR1 alone between inward-or outward-facing conformations with low or high dissociation constant, K D , values for the conformation-sensitive channel antagonist [ 3 H]glibenclamide ([ 3 H]GBM), indicating that ATP can act as a pure agonist. Assembly with Kir6.2 reduced SUR1's K D for [ 3 H]GBM.ThisreductionrequiredtheKirNterminus(KNtp), consistent with KNtp occupying a "transport cavity," thus positioning it to link ATP-induced SUR1 conformational changes to channel gating. Moreover, ATP/GBM site coupling was constrained in WT SUR1/WT Kir6.2 channels; ATP-bound channels had a lower K D for [ 3 H]GBM than ATP-bound SUR1. This constraint was largely eliminated by the Q1179R neonatal diabetes-associated mutation in helix 15, suggesting that a "swapped" helix pair, 15 and 16, is part of a structural pathway connecting the ATP/GBM sites. Our results suggest that ATP binding to SUR1 biases K ATP channels toward open states, consistent with SUR1 variants with lower K D values causing neonatal diabetes, whereas increased K D values cause congenital hyperinsulinism.Neuroendocrine-type ATP-sensitive K ϩ (K ATP ) 2 channels comprise an ATP-binding cassette (ABC) protein (1), ABCC8/ SUR1, and a K ϩ -selective inward rectifier (2), KCNJ11/Kir6.2, assembled as heterooctamers (3-5), (SUR1/Kir6.2) 4 . In pancreatic ␤-cells, these channels are metabolite sensors that couple cellular metabolism with membrane electrical activity to link insulin secretion with blood glucose levels. This coupling is critical for normal physiology; loss of channel function is a cause of congenital hyperinsulinism (Ref. 6; for reviews, see Refs. 7 and 8), whereas gain-of-function mutations in Kir6.2 (9) and SUR1 (10) cause neonatal diabetes (ND) (for reviews, see Refs. 11 and 12). Gain of function is one cause of mature-onset diabetes of the young (13), whereas a ABCC8/SUR1 polymorphism, the Ala amino acid allele at p.A1369S, is associated with an increased risk for type 2 diabetes (14).Channel activity is regulated positively by ATP and ADP binding to SUR1 and negatively by nucleotide binding to . Additionally, multiple metabolites, including phosphoinositides (21-23) and long-chain acyl-CoA esters (24 -26), and phosphorylation (27-29) positively modulate ch...

The role of liver X receptor (LXR) in pancreatic β-cell physiology and pathophysiology is still unclear. It has been postulated that chronic LXR activation in β-cells induces lipotoxicity, a key step in the development of β-cell dysfunction, which accompanies type 2 diabetes mellitus. In most of these studies, the LXR ligand T0901317 has been administered chronically in the micromolar range to study the significance of LXR activation. In the current study, we have evaluated acute effects of T0901317 on stimulus-secretion coupling of β-cells. We found that 10 µM T0901317 completely suppressed oscillations of the cytosolic Ca2+ concentration induced by 15 mM glucose. Obviously, this effect was due to inhibition of mitochondrial metabolism. T0901317 markedly depolarized the mitochondrial membrane potential, thus inhibiting adenosine triphosphate (ATP) production and reducing the cytosolic ATP concentration. This led in turn to a huge increase in KATP current and hyperpolarization of the cell membrane potential. Eventually, T0901317 inhibited glucose-induced insulin secretion. These effects were rapid in on-set and not compatible with the activation of a nuclear receptor. In vivo, T0901317 acutely increased the blood glucose concentration after intraperitoneal application. In summary, these data clearly demonstrate that T0901317 exerts acute effects on stimulus-secretion coupling. This observation questions the chronic use of T0901317 and limits the interpretation of results obtained under these experimental conditions.

Objective: Congenital hyperinsulinism (CHI) is a rare disease characterized by persistent hypoglycemia as a result of inappropriate insulin secretion, which can lead to irreversible neurological defects in infants. Poor efficacy and strong adverse effects of the current medications impede successful treatment. The aim of the study was to investigate new approaches to silence b-cells and thus attenuate insulin secretion. Research Design and Methods: In the scope of our research, we tested substances more selective and more potent than the gold standard diazoxide that also interact with neuroendocrine ATP-sensitive K + (K ATP) channels. Additionally, K ATP channel-independent targets as Ca 2+-activated K + channels of intermediate conductance (K Ca 3.1) and L-type Ca 2+ channels were investigated. Experiments were performed using human islet cell clusters isolated from tissue of CHI patients (histologically classified as pathological) and islet cell clusters obtained from C57BL/6N (WT) or SUR1 knockout (SUR1-/-) mice. The cytosolic Ca 2+ concentration ([Ca 2+ ] c) was used as a parameter for the pathway regulated by electrical activity and was determined by fura-2 fluorescence. The mitochondrial membrane potential (DY) was determined by rhodamine 123 fluorescence and single channel currents were measured by the patch-clamp technique. Results: The selective K ATP channel opener NN414 (5 µM) diminished [Ca 2+ ] c in isolated human CHI islet cell clusters and WT mouse islet cell clusters stimulated with 10 mM glucose. In islet cell clusters lacking functional K ATP channels (SUR1-/-) the drug was without effect. VU0071063 (30 µM), another K ATP channel opener considered to be selective, lowered [Ca 2+ ] c in human CHI islet cell clusters. The compound was also effective in islet cell clusters from SUR1-/mice, showing that [Ca 2+ ] c is influenced by additional effects besides K ATP channels. Contrasting to NN414, the drug depolarized DY in murine islet cell clusters pointing to severe interference with mitochondrial metabolism. An opener of K Ca 3.1 channels, DCEBIO (100 µM), significantly decreased [Ca 2+ ] c in SUR1-/and human CHI islet cell clusters. To target L-type Ca 2+ channels we tested two

It is suggested that extracellular ATP which is co-secreted with insulin in a pulsatile manner during glucose-stimulated exocytosis provides a negative feedback signal driving β-cell oscillations in co-operation with Ca and other signals.