NMDA receptors mediate leptin signaling and regulate potassium channel trafficking in pancreatic β-cells

Abstract

NMDA receptors (NMDARs) are Ca-permeant, ligand-gated ion channels activated by the excitatory neurotransmitter glutamate and have well-characterized roles in the nervous system. The expression and function of NMDARs in pancreatic β-cells, by contrast, are poorly understood. Here, we report a novel function of NMDARs in β-cells. Using a combination of biochemistry, electrophysiology, and imaging techniques, we now show that NMDARs have a key role in mediating the effect of leptin to modulate β-cell electrical activity by promoting AMP-activated protein kinase (AMPK)-dependent trafficking of K and Kv2.1 channels to the plasma membrane. Blocking NMDAR activity inhibited the ability of leptin to activate AMPK, induce K and Kv2.1 channel trafficking, and promote membrane hyperpolarization. Conversely, activation of NMDARs mimicked the effect of leptin, causing Ca influx, AMPK activation, and increased trafficking of K and Kv2.1 channels to the plasma membrane, and triggered membrane hyperpolarization. Moreover, leptin potentiated NMDAR currents and triggered NMDAR-dependent Ca influx. Importantly, NMDAR-mediated signaling was observed in rat insulinoma 832/13 cells and in human β-cells, indicating that this pathway is conserved across species. The ability of NMDARs to regulate potassium channel surface expression and thus, β-cell excitability provides mechanistic insight into the recently reported insulinotropic effects of NMDAR antagonists and therefore highlights the therapeutic potential of these drugs in managing type 2 diabetes.

Leptin modulates pancreatic β-cell membrane potential through Src kinase–mediated phosphorylation of NMDA receptors

“…In the absence of leptin, repeated puff applications of 1mM NMDA at one minute intervals elicited NMDAR currents of similar amplitudes. Bath application of 10nM leptin potentiated NMDA-evoked currents within 5 min of leptin treatment from a baseline of 14.8 ± 8.0 pA to 25.8 ± 13.3 pA (p<0.05 by Friedman's test; Fig.3), as we reported previously (19). Subsequent co-application of 10M AZD0530 with leptin abolished the effect of leptin, with averaged NMDA currents of 15.5 ± 8.7 pA comparable to baseline values (Fig.3).…”

“…These experiments were performed using cell-attached current-clamp recording, which provides a robust assessment of changes in membrane potential while maintaining cell integrity and preventing dialysis of soluble factors that may be important for intracellular signaling (31,32). Bath application of leptin alone (10nM) induced a mean membrane hyperpolarization of −46.8 ± 8.1 mV, similar to that reported previously (9,19). However, co-application of the GluN2A selective antagonist TCN-201 (50µM) with leptin only induced a mean hyperpolarization of -14.4 ± 3.9 mV, significantly less than that observed in cells treated with leptin alone (Fig.1E, F).…”

“…In rodent pancreatic β-cell lines as well as primary human β-cells, leptin induces membrane hyperpolarization at glucose concentrations that depolarize β-cell membrane potential (3,(7)(8)(9)19). Recently, we showed that, both in rat insulinoma INS-1 832/13 cells and human β-cells, this effect is mediated by NMDARs (19). Leptin stimulation increased NMDA currents, leading to increased Ca 2+ influx and increased surface density of KATP and Kv2.1 channels (19).…”

“…Recently, we showed that, both in rat insulinoma INS-1 832/13 cells and human β-cells, this effect is mediated by NMDARs (19). Leptin stimulation increased NMDA currents, leading to increased Ca 2+ influx and increased surface density of KATP and Kv2.1 channels (19). To determine the mechanism by which leptin increases NMDA currents, we began by characterizing the NMDARs that are expressed in β-cells.…”

“…NMDARs are calcium-permeant ionotropic glutamate receptors that are highly expressed in the brain and are important for learning and memory (16). Although they are extensively studied in the central nervous system, there is growing evidence that they are expressed in pancreatic β-cells and play a role in regulating insulin secretion (17)(18)(19). Functional NMDARs generally form as heterotetramers of two obligatory glycine-binding GluN1 subunits (also known as NR1) and two glutamate-binding GluN2 (GluN2A-D, or NR2A-D) subunits (20).…”