TRPM7 is a crucial regulator of pancreatic endocrine development and high-fat-diet-induced ß-cell proliferation.

The melastatin subfamily of the transient receptor potential channels (TRPM) are regulators of pancreatic ß-cell function. TRPM7 is the most abundant islet TRPM channel; however, the role of TRPM7 in ß-cell function has not been determined. Here, we used various spatiotemporal transgenic mouse models to investigate how TRPM7 knockout influences pancreatic endocrine development, proliferation and function. Ablation of TRPM7 within pancreatic progenitors reduced pancreatic size, and α-cell and ß-cell mass. This resulted in modestly impaired glucose tolerance. However, TRPM7 ablation following endocrine specification or in adult mice did not impact endocrine expansion or glucose tolerance. As TRPM7 regulates cell proliferation, we assessed how TRPM7 influences ß-cell hyperplasia under insulin-resistant conditions. ß-Cell proliferation induced by high-fat diet was significantly decreased in TRPM7-deficient ß-cells. The endocrine roles of TRPM7 may be influenced by cation flux through the channel, and indeed we found that TRPM7 ablation altered ß-cell Mg2+ and reduced the magnitude of elevation in ß-cell Mg2+ during proliferation. Together, these findings revealed that TRPM7 controls pancreatic development and ß-cell proliferation, which is likely due to regulation of Mg2+ homeostasis.

Liraglutide increases islet Ca2+ oscillation frequency and insulin secretion by activating hyperpolarization-activated cyclic nucleotide-gated channels.

AIM: To determine whether hyperpolarization-activated cyclic nucleotide-gated (HCN) channels impact glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) modulation of islet Ca2+ handling and insulin secretion. METHODS: The impact of liraglutide (GLP-1 analogue) on islet Ca2+ handling, HCN currents and insulin secretion was monitored with fluorescence microscopy, electrophysiology and enzyme immunoassays, respectively. Furthermore, liraglutide-mediated ß-to-δ-cell cross-communication was assessed following selective ablation of either mouse islet δ or ß cells. RESULTS: Liraglutide increased ß-cell Ca2+ oscillation frequency in mouse and human islets under stimulatory glucose conditions. This was dependent in part on liraglutide activation of HCN channels, which also enhanced insulin secretion. Similarly, liraglutide activation of HCN channels also increased ß-cell Ca2+ oscillation frequency in islets from rodents exposed to a diabetogenic diet. Interestingly, liraglutide accelerated Ca2+ oscillations in a majority of islet δ cells, which showed synchronized Ca2+ oscillations equivalent to ß cells; therefore, we assessed if either cell type was driving this liraglutide-mediated islet Ca2+ response. Although δ-cell loss did not impact liraglutide-mediated increase in ß-cell Ca2+ oscillation frequency, ß-cell ablation attenuated liraglutide-facilitated acceleration of δ-cell Ca2+ oscillations. CONCLUSION: The data presented here show that liraglutide-induced stimulation of islet HCN channels augments Ca2+ oscillation frequency. As insulin secretion oscillates with ß-cell Ca2+ , these findings have important implications for pulsatile insulin secretion that is probably enhanced by liraglutide activation of HCN channels and therapeutics that target GLP-1Rs for treating diabetes. Furthermore, these studies suggest that liraglutide as well as GLP-1-based therapies enhance δ-cell Ca2+ oscillation frequency and somatostatin secretion kinetics in a ß-cell-dependent manner.

Tetraspanin-7 regulation of L-type voltage-dependent calcium channels controls pancreatic ß-cell insulin secretion.

KEY POINTS: Tetraspanin (TSPAN) proteins regulate many biological processes, including intracellular calcium (Ca2+ ) handling. TSPAN-7 is enriched in pancreatic islet cells; however, the function of islet TSPAN-7 has not been identified. Here, we characterize how ß-cell TSPAN-7 regulates Ca2+ handling and hormone secretion. We find that TSPAN-7 reduces ß-cell glucose-stimulated Ca2+ entry, slows Ca2+ oscillation frequency and decreases glucose-stimulated insulin secretion. TSPAN-7 controls ß-cell function through a direct interaction with L-type voltage-dependent Ca2+ channels (CaV 1.2 and CaV 1.3), which reduces channel Ca2+ conductance. TSPAN-7 slows activation of CaV 1.2 and accelerates recovery from voltage-dependent inactivation; TSPAN-7 also slows CaV 1.3 inactivation kinetics. These findings strongly implicate TSPAN-7 as a key regulator in determining the set-point of glucose-stimulated Ca2+ influx and insulin secretion. ABSTRACT: Glucose-stimulated insulin secretion (GSIS) is regulated by calcium (Ca2+ ) entry into pancreatic ß-cells through voltage-dependent Ca2+ (CaV ) channels. Tetraspanin (TSPAN) transmembrane proteins control Ca2+ handling, and thus they may also modulate GSIS. TSPAN-7 is the most abundant islet TSPAN and immunostaining of mouse and human pancreatic slices shows that TSPAN-7 is highly expressed in ß- and α-cells; however, the function of islet TSPAN-7 has not been determined. Here, we show that TSPAN-7 knockdown (KD) increases glucose-stimulated Ca2+ influx into mouse and human ß-cells. Additionally, mouse ß-cell Ca2+ oscillation frequency was accelerated by TSPAN-7 KD. Because TSPAN-7 KD also enhanced Ca2+ entry when membrane potential was clamped with depolarization, the effect of TSPAN-7 on CaV channel activity was examined. TSPAN-7 KD enhanced L-type CaV currents in mouse and human ß-cells. Conversely, heterologous expression of TSPAN-7 with CaV 1.2 and CaV 1.3 L-type CaV channels decreased CaV currents and reduced Ca2+ influx through both channels. This was presumably the result of a direct interaction of TSPAN-7 and L-type CaV channels because TSPAN-7 coimmunoprecipitated with both CaV 1.2 and CaV 1.3 from primary human ß-cells and from a heterologous expression system. Finally, TSPAN-7 KD in human ß-cells increased basal (5.6 mM glucose) and stimulated (45 mM KCl + 14 mM glucose) insulin secretion. These findings strongly suggest that TSPAN-7 modulation of ß-cell L-type CaV channels is a key determinant of ß-cell glucose-stimulated Ca2+ entry and thus the set-point of GSIS.

Gi/o protein-coupled receptor inhibition of beta-cell electrical excitability and insulin secretion depends on Na+/K+ ATPase activation.

Gi/o-coupled somatostatin or α2-adrenergic receptor activation stimulated ß-cell NKA activity, resulting in islet Ca2+ fluctuations. Furthermore, intra-islet paracrine activation of ß-cell Gi/o-GPCRs and NKAs by δ-cell somatostatin secretion slowed Ca2+ oscillations, which decreased insulin secretion. ß-cell membrane potential hyperpolarization resulting from Gi/o-GPCR activation was dependent on NKA phosphorylation by Src tyrosine kinases. Whereas, ß-cell NKA function was inhibited by cAMP-dependent PKA activity. These data reveal that NKA-mediated ß-cell membrane potential hyperpolarization is the primary and conserved mechanism for Gi/o-GPCR control of electrical excitability, Ca2+ handling, and insulin secretion.

A KCNK16 mutation causing TALK-1 gain of function is associated with maturity-onset diabetes of the young.

Maturity-onset diabetes of the young (MODY) is a heterogeneous group of monogenic disorders of impaired pancreatic ß cell function. The mechanisms underlying MODY include ß cell KATP channel dysfunction (e.g., KCNJ11 [MODY13] or ABCC8 [MODY12] mutations); however, no other ß cell channelopathies have been associated with MODY to date. Here, we have identified a nonsynonymous coding variant in KCNK16 (NM_001135105: c.341T>C, p.Leu114Pro) segregating with MODY. KCNK16 is the most abundant and ß cell-restricted K+ channel transcript, encoding the two-pore-domain K+ channel TALK-1. Whole-cell K+ currents demonstrated a large gain of function with TALK-1 Leu114Pro compared with TALK-1 WT, due to greater single-channel activity. Glucose-stimulated membrane potential depolarization and Ca2+ influx were inhibited in mouse islets expressing TALK-1 Leu114Pro with less endoplasmic reticulum Ca2+ storage. TALK-1 Leu114Pro significantly blunted glucose-stimulated insulin secretion compared with TALK-1 WT in mouse and human islets. These data suggest that KCNK16 is a previously unreported gene for MODY.

Lactate activation of α-cell KATP channels inhibits glucagon secretion by hyperpolarizing the membrane potential and reducing Ca2+ entry.

OBJECTIVE: Elevations in pancreatic α-cell intracellular Ca2+ ([Ca2+]i) lead to glucagon (GCG) secretion. Although glucose inhibits GCG secretion, how lactate and pyruvate control α-cell Ca2+ handling is unknown. Lactate enters cells through monocarboxylate transporters (MCTs) and is also produced during glycolysis by lactate dehydrogenase A (LDHA), an enzyme expressed in α-cells. As lactate activates ATP-sensitive K+ (KATP) channels in cardiomyocytes, lactate may also modulate α-cell KATP. Therefore, this study investigated how lactate signaling controls α-cell Ca2+ handling and GCG secretion. METHODS: Mouse and human islets were used in combination with confocal microscopy, electrophysiology, GCG immunoassays, and fluorescent thallium flux assays to assess α-cell Ca2+ handling, Vm, KATP currents, and GCG secretion. RESULTS: Lactate-inhibited mouse (75 ± 25%) and human (47 ± 9%) α-cell [Ca2+]i fluctuations only under low-glucose conditions (1 mM) but had no effect on ß- or δ-cells [Ca2+]i. Glyburide inhibition of KATP channels restored α-cell [Ca2+]i fluctuations in the presence of lactate. Lactate transport into α-cells via MCTs hyperpolarized mouse (14 ± 1 mV) and human (12 ± 1 mV) α-cell Vm and activated KATP channels. Interestingly, pyruvate showed a similar KATP activation profile and α-cell [Ca2+]i inhibition as lactate. Lactate-induced inhibition of α-cell [Ca2+]i influx resulted in reduced GCG secretion in mouse (62 ± 6%) and human (43 ± 13%) islets. CONCLUSIONS: These data demonstrate for the first time that lactate entry into α-cells through MCTs results in KATP activation, Vm hyperpolarization, reduced [Ca2+]i, and inhibition of GCG secretion. Thus, taken together, these data indicate that lactate either within α-cells and/or elevated in serum could serve as important modulators of α-cell function.