PKA-dependent potentiation of glucose-stimulated insulin secretion by Epac activator 8-pCPT-2′-<i>O</i>-Me-cAMP-AM in human islets of Langerhans

Chepurny, Oleg G.; Kelley, Grant G.; Dzhura, Igor; Leech, Colin A.; Roe, Michael W.; Dzhura, Elvira; Li, Xiangquan; Schwede, Frank; Genieser, Hans‐Gottfried; Holz, George G.

doi:10.1152/ajpendo.00630.2009

Cited by 70 publications

(94 citation statements)

References 74 publications

(109 reference statements)

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“…The kinase inhibitor H-89 had been reported not to alter the effect of glucose in intact islets [32]. However, recent pharmacological data from mouse [2], rat [33] and human islets [28,34] are in line with our observations. Previously reported differences may be due to the timing of drug addition as they are less evident once glucoseinduced effects are installed [2].…”

Section: Discussionsupporting

confidence: 91%

Multilevel control of glucose homeostasis by adenylyl cyclase 8

et al. 2014

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Aims/hypothesis Nutrient homeostasis requires integration of signals generated by glucose metabolism and hormones. Expression of the calcium-stimulated adenylyl cyclase ADCY8 is regulated by glucose and the enzyme is capable of integrating signals from multiple pathways. It may thus have an important role in glucose-induced signalling and glucose homeostasis. Methods We used pharmacological and genetic approaches in beta cells to determine secretion and calcium metabolism. Furthermore, Adcy8 knockout mice were characterised. Results In clonal beta cells, inhibitors of adenylyl cyclases or their downstream targets reduced the glucose-induced increase in cytosolic calcium and insulin secretion. This was reproduced by knock-down of ADCY8, but not of ADCY1. These agents also inhibited glucose-induced increase in cytosolic calcium and electrical activity in primary beta cells and similar effects were observed after ADCY8 knock-down.Moreover, insulin secretion was diminished in islets from Adcy8 knockout mice. These mice were glucose intolerant after oral or intraperitoneal administration of glucose whereas their levels of glucagon-like peptide-1 remained unaltered. Finally, we knocked down ADCY8 in the ventromedial hypothalamus to evaluate the need for ADCY8 in the central regulation of glucose homeostasis. Whereas mice fed a standard diet had normal glucose levels, high-fat diet exacerbated glucose intolerance and knock-down mice were incapable of raising their plasma insulin levels. Finally we confirmed that ADCY8 is expressed in human islets. Conclusions/interpretations Collectively, our findings demonstrate that ADCY8 is required for the physiological activation of glucose-induced signalling pathways in beta cells, for glucose tolerance and for hypothalamic adaptation to a high-fat diet via regulation of islet insulin secretion.Electronic supplementary material The online version of this article (doi:10.1007/s00125-014-3445-z) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

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Section: Discussionsupporting

confidence: 91%

Multilevel control of glucose homeostasis by adenylyl cyclase 8

et al. 2014

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“…The excitation light was delivered at 435/9 nm (455 nm cutoff), and the emitted light was detected at 485/15 nm (CFP) or 535/15 nm (YFP). For live-cell imaging of FRET in single cells of the HEK cell clones reported here, the methods of analyses were identical to those reported by Chepurny et al in prior reports (40,41).…”

Section: Methodsmentioning

confidence: 93%

Isoform-specific antagonists of exchange proteins directly activated by cAMP

Tsalkova

Mei

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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127

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The major physiological effects of cAMP in mammalian cells are transduced by two ubiquitously expressed intracellular cAMP receptors, protein kinase A (PKA) and exchange protein directly activated by cAMP (EPAC), as well as cyclic nucleotide-gated ion channels in certain tissues. Although a large number of PKA inhibitors are available, there are no reported EPAC-specific antagonists, despite extensive research efforts. Here we report the identification and characterization of noncyclic nucleotide EPAC antagonists that are exclusively specific for the EPAC2 isoform. These EAPC2-specific antagonists, designated as ESI-05 and ESI-07, inhibit Rap1 activation mediated by EAPC2, but not EPAC1, with high potency in vitro. Moreover, ESI-05 and ESI-07 are capable of suppressing the cAMP-mediated activation of EPAC2, but not EPAC1 and PKA, as monitored in living cells through the use of EPAC-and PKA-based FRET reporters, or by the use of Rap1-GTP pull-down assays. Deuterium exchange mass spectroscopy analysis further reveals that EPAC2-specific inhibitors exert their isoform selectivity through a unique mechanism by binding to a previously undescribed allosteric site: the interface of the two cAMP binding domains, which is not present in the EPAC1 isoform. Isoform-specific EPAC pharmacological probes are highly desired and will be valuable tools for dissecting the biological functions of EPAC proteins and their roles in various disease states.cAMP-regulated guanine nucleotide exchange factor | high-throughput screening | hydrogen/deuterium exchange mass spectrometry

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“…8-pCPT-acetoxymethyl (8-pCPT-AM), an activator of EPAC with little activation effect on PKA at lower micromolar doses (25), enhanced the NSCC current increase at 2.8 mmol/L glucose more potently than ex-4 (Fig. 6C).…”

Section: Attenuation Of Glucose- Ex-4- and Gip-elicited Increase Inmentioning

confidence: 99%

Involvement of cAMP/EPAC/TRPM2 Activation in Glucose- and Incretin-Induced Insulin Secretion

et al. 2014

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In pancreatic b-cells, closure of the ATP-sensitive K + (K ATP ) channel is an initial process triggering glucosestimulated insulin secretion. In addition, constitutive opening of background nonselective cation channels (NSCCs) is essentially required to effectively evoke depolarization as a consequence of K ATP channel closure. Thus, it is hypothesized that further opening of NSCC facilitates membrane excitability. We identified a class of NSCC that was activated by exendin (ex)-4, GLP-1, and its analog liraglutide at picomolar levels. This NSCC was also activated by increasing the glucose concentration. NSCC activation by glucose and GLP-1 was a consequence of the activated cAMP/EPAC-mediated pathway and was attenuated in TRPM2-deficient mice. The NSCC was not activated by protein kinase A (PKA) activators and was activated by ex-4 in the presence of PKA inhibitors. These results suggest that glucose-and incretinactivated NSCC (TRPM2) works in concert with closure of the K ATP channel to effectively induce membrane depolarization to initiate insulin secretion. The current study reveals a new mechanism for regulating electrical excitability in b-cells and for mediating the action of glucose and incretin to evoke insulin secretion, thereby providing an innovative target for the treatment of type 2 diabetes.It has been long proposed that glucose-stimulated insulin secretion in pancreatic b-cells is initiated by closure of the ATP-sensitive K + (K ATP ) channel, followed by membrane depolarization (1). In theory, closure of the K ATP channel is an important process but is not sufficient to induce the shift of the membrane potential toward a threshold level, as membrane potential is determined by the overall balance of outward and inward currents. Modest constitutive opening of background inward current through nonselective cation channels (NSCCs) is crucial to facilitate depolarization after K ATP channel closure (2). This idea suggests that further regulated opening of a class of NSCCs may bring about greater depolarization. However, whether glucose metabolism regulates NSCC activity remains unclear.The incretin hormones, GLP-1 and glucose-dependent insulinotropic polypeptide (GIP), potentiate insulin secretion in association with increased intracellular Ca 2+ concentrations at insulin-secreting glucose concentrations (3-5). GLP-1 fails to increase insulin secretion from the islets of mice deficient in transmembrane receptor potential (TRP) melastatin 2 (TRPM2) (6,7), a type of NSCC, suggesting that the TRPM2 channel is essential for GLP-1-induced potentiation of glucose-stimulated insulin secretion (8). GLP-1 depolarizes the plasma membrane by the opening of NSCC in b-cells (2). Several types of NSCC (TRPs) are expressed in insulin-secreting cells (9). The aims of the current study were to determine 1) the type of NSCC activation (through TRPM2 or other TRPs) that is crucial for signaling after stimulation of the incretin receptor, 2) whether the NSCC is modulated by glucose

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PKA-dependent potentiation of glucose-stimulated insulin secretion by Epac activator 8-pCPT-2′-O-Me-cAMP-AM in human islets of Langerhans

Cited by 70 publications

References 74 publications

Multilevel control of glucose homeostasis by adenylyl cyclase 8

Multilevel control of glucose homeostasis by adenylyl cyclase 8

Isoform-specific antagonists of exchange proteins directly activated by cAMP

Involvement of cAMP/EPAC/TRPM2 Activation in Glucose- and Incretin-Induced Insulin Secretion

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