Arcuate NPY Neurons Research Articles

Metabolic insights from a GHSR-A203E mutant mouse model

ObjectiveBinding of ghrelin to its receptor, growth hormone secretagogue receptor (GHSR), stimulates GH release, induces eating, and increases blood glucose. These processes may also be influenced by constitutive (ghrelin-independent) GHSR activity, as suggested by findings in short people with naturally occurring GHSR-A204E mutations and reduced food intake and blood glucose in rodents administered GHSR inverse agonists, both of which impair constitutive GHSR activity. In this study, we aimed to more fully determine the physiologic relevance of constitutive GHSR activity. MethodsWe generated mice with a GHSR mutation that replaces alanine at position 203 with glutamate (GHSR-A203E), which corresponds to the previously described human GHSR-A204E mutation, and used them to conduct ex vivo neuronal electrophysiology and in vivo metabolic assessments. We also measured signaling within COS-7 and HEK293T cells transfected with wild-type GHSR (GHSR-WT) or GHSR-A203E constructs. ResultsIn COS-7 cells, GHSR-A203E resulted in lower baseline IP3 accumulation than GHSR-WT; ghrelin-induced IP3 accumulation was observed in both constructs. In HEK293T cells co-transfected with voltage-gated CaV2.2 calcium channel complex, GHSR-A203E had no effect on basal CaV2.2 current density while GHSR-WT did; both GHSR-A203E and GHSR-WT inhibited CaV2.2 current in the presence of ghrelin. In cultured hypothalamic neurons from GHSR-A203E and GHSR-deficient mice, native calcium currents were greater than those in neurons from wild-type mice; ghrelin inhibited calcium currents in cultured hypothalamic neurons from both GHSR-A203E and wild-type mice. In brain slices, resting membrane potentials of arcuate NPY neurons from GHSR-A203E mice were hyperpolarized compared to those from wild-type mice; the same percentage of arcuate NPY neurons from GHSR-A203E and wild-type mice depolarized upon ghrelin exposure. The GHSR-A203E mutation did not significantly affect body weight, body length, or femur length in the first ∼6 months of life, yet these parameters were lower in GHSR-A203E mice after 1 year of age. During a 7-d 60% caloric restriction regimen, GHSR-A203E mice lacked the usual marked rise in plasma GH and demonstrated an exaggerated drop in blood glucose. Administered ghrelin also exhibited reduced orexigenic and GH secretagogue efficacies in GHSR-A203E mice. ConclusionsOur data suggest that the A203E mutation ablates constitutive GHSR activity and that constitutive GHSR activity contributes to the native depolarizing conductance of GHSR-expressing arcuate NPY neurons. Although the A203E mutation does not block ghrelin-evoked signaling as assessed using in vitro and ex vivo models, GHSR-A203E mice lack the usual acute food intake response to administered ghrelin in vivo. The GHSR-A203E mutation also blunts GH release, and in aged mice leads to reduced body length and femur length, which are consistent with the short stature of human carriers of the GHSR-A204E mutation.

Salmon calcitonin distributes into the arcuate nucleus to a subset of NPY neurons in mice

The amylin receptor (AMY) and calcitonin receptor (CTR) agonists induce acute suppression of food intake in rodents by binding to receptors in the area postrema (AP) and potentially by targeting arcuate (ARC) neurons directly. Salmon calcitonin (sCT) induces more potent, longer lasting anorectic effects compared to amylin. We thus aimed to investigate whether AMY/CTR agonists target key neuronal populations in the ARC, and whether differing brain distribution patterns could mediate the observed differences in efficacy with sCT and amylin treatment. Brains were examined by whole brain 3D imaging and confocal microscopy following subcutaneous administration of fluorescently labelled peptides to mice. We found that sCT, but not amylin, internalizes into a subset of ARC NPY neurons, along with an unknown subset of ARC, AP and dorsal vagal motor nucleus cells. ARC POMC neurons were not targeted. Furthermore, amylin and sCT displayed similar distribution patterns binding to receptors in the AP, the organum vasculosum of the lamina terminalis (OVLT) and the ARC. Amylin distributed within the median eminence with only specs of sCT being present in this region, however amylin was only detectable 10 minutes after injection while sCT displayed a residence time of up to 2 hours post injection. We conclude that AMY/CTR agonists bind to receptors in a subset of ARC NPY neurons and in circumventricular organs. Furthermore, the more sustained and greater anorectic efficacy of sCT compared to rat amylin is not attributable to differences in brain distribution patterns but may more likely be explained by greater potency at both the CTR and AMY.

FGF19: How gut talks to brain to keep your sugar down

FGF19: How gut talks to brain to keep your sugar down

The arcuate nucleus as a conduit for diverse signals relevant to energy homeostasis.

Arcuate nucleus neurons are known to be responsive to a wide array of hormones and nutrients, including leptin, insulin, gonadal steroids and glucose. In addition to potential transport mechanisms, peripheral substances may access these neurons via arcuate cell bodies in and projections to the median eminence, a region considered to be a circumventricular organ. The arcuate is a potent site of leptin action, probably mediating a component of leptin's effects via arcuate neuropeptide Y/agouti-related peptide (NPY/AgRP) and pro-opiomelanocortin (POMC) neurons, and implicating this structure in the long-term control of energy stores. However, ghrelin, the endogenous ligand of the growth hormone secretagogue receptor, may also stimulate feeding and weight gain, in part through action on receptors in arcuate NPY neurons. Since ghrelin is secreted by the stomach upon content depletion, with a half-life of no more than an hour, the arcuate nucleus may also be important in sensing and responding to acute changes in nutrients. We have developed a system for recording from arcuate POMC neurons using a mouse containing a transgene in which the POMC promoter is driving expression of the green fluorescent protein (GFP). In these mice, 99% of the beta-endorphin positive neurons express GFP, making whole cell patch clamp recordings from the sparsely distributed POMC neurons facile. All of the POMC neurons appear to be activated by leptin, via two different mechanisms, while approximately 30-50% of the neurons appear to be inhibited by a gamma-melanocyte stimulating hormone (MSH) specific agonist. The latter result suggests that the melanocortin-3 receptor (MC3-R) may act as an autoinhibitory receptor on some POMC neurons. This hypothalamic slice preparation also confirms the responsiveness of arcuate POMC neurons to a wide variety of nutrients and hormones. Thus the arcuate melanocortin system is best described as a conduit of many diverse signals involved in energy homeostasis, with leptin acting tonically to regulate the responsiveness of the circuit to a wide variety of hormones and nutrients.

Ultrastructural Localization of Neuropeptide Y in the Hypothalamus

Immunoreactive NPY neurons are widely distributed in the hypothalamus of several mammalian species. In the rat, dense NPY fiber networks are found in the paraventricular, suprachiasmatic and arcuate nuclei. NPY-containing cell bodies are mostly found in the arcuate nucleus. Studies performed at the electron microscope level clearly indicate that NPY is concentrated in dense core vesicles in the cytoplasm of cell bodies as well as in terminals. Only a small percentage (about 20%) of the NPY endings are making synaptic contacts with nerve processes, especially dendrites. These ultrastructural data suggest that NPY might play a neurotransmitter/neuromodulator role. NPY has been shown, when injected into hypothalamic areas, to exert a variety of effects, including modifications in food intake, energy balance and pituitary secretion. In an attempt to define the exact role of NPY in hypothalamic functions, we have designed experiments to study the interactions of NPY with other neurotransmitter systems. In the suprachiasmatic nucleus, both NPY and 5-HT terminals have been shown to establish synaptic junctions sometimes with the same neurons. Occasionally, axoaxonic junctions between these two types of endings have been observed. These results suggested that both 5-HT and NPY might be involved in the complex regulation of circadian rythms. In the arcuate nucleus, nonsynaptic appositions between 5-HT nerve endings and NPY-containing neurons were demonstrated. In this nucleus, direct appositions between TH- and NPY-containing neurons were also detected. These appositions were of axosomatic, axodendritic or axoaxonic types. Since it has been demonstrated that arcuate NPY neurons are projected to other hypothalamic areas, such as the paraventricular and dorsomedial nuclei, it might be speculated that arcuate 5-HT/NPY and catecholamines/NPY interactions might be involved in regulation of behavior and neuroendocrine functions.

On the Role of Neuropeptide Y in Information Handling in the Central Nervous System in Normal and Physiopathological States

The NPY neurons play an important role in information handling in the CNS by their ability to interact in both wiring and volume transmission at the network, local circuit and synaptic level. The importance of NPY/alpha 2 receptor-receptor interactions in cardiovascular, neuroendocrine and vigilance control is emphasized. Alterations in these receptor-receptor interactions take place in the spontaneously hypertensive rats as well as in the ischemic brain, which may have profound consequences for the information handling and contribute to the functional alterations found in these pathophysiological states. Finally, in the aging brain there appears to exist a marked reduction in NPY transmission line, which may affect higher brain functions, such as learning and memory retrieval. The most impressive result is, however, the indications of a role for NPY in volume transmission, where NPY appears to produce syndromic actions via its conversion into biologically active fragments, which may have preferential actions at Y2 NPY receptors. These syndromic pathways may be altered in the spontaneously hypertensive rat and may be controlled by gonadal steroids and glucocorticoids. Glucocorticoid receptors have been demonstrated in all arcuate NPY neurons and all NA/NPY and A/NPY costoring neurons.