Mind affects matter: Hindbrain GLP1 neurons link stress, physiology and behaviour

The brain responds rapidly to stressful stimuli by increasing sympathetic outflow, activating the hypothalamic–pituitary–adrenal axis and eliciting avoidance behaviours to limit risks to safety. Stress responses are adaptive and essential but can become maladaptive when the stress is chronic, causing autonomic imbalance, hypothalamic–pituitary–adrenal axis hyper-reactivity and a state of hypervigilance. Ultimately, this contributes to the development of cardiovascular disease and affective disorders, including major depression and anxiety. Stress responses are often thought to be driven mainly by forebrain areas; however, the brainstem nucleus of the solitary tract (NTS) is ideally located to control both autonomic outflow and behaviour in response to stress. Here, I review the preclinical evidence that the NTS and its resident glucagon-like peptide-1 (GLP1)-expressing neurons are prominent mediators of stress responses. This Lecture introduces the reader to the idea of good and bad stress and outlines the types of stress that engage the NTS and GLP1 neurons. I describe in particular detail the recent studies by myself and others aimed at mapping sources of synaptic inputs to GLP1 neurons and consider the implications for our understanding of the role of GLP1 neurons in stress. This is followed by a discussion of the contribution of brain GLP1 and GLP1 neurons to behavioural and physiological stress responses. The evidence reviewed highlights a potentially prominent role for GLP1 neurons in the response of the brain to acute stress and reveals important unanswered questions regarding their role in chronic stress

The physiological role of the brain GLP-1 system in stress

Glucagon-like peptide-1 (GLP-1) within the brain is a potent regulator of food intake and most studies have investigated the anorexic effects of central GLP-1. A range of brain regions have now been found to be involved in GLP-1 mediated anorexia, including some which are not traditionally associated with appetite regulation. However, a change in food intake can be indicative of not only reduced energy demand, but also changes in the organism's motivation to eat following stressful stimuli. In fact, acute stress is well-known to reduce food intake. Recently, more research has focused on the role of GLP-1 in stress and the central GLP-1 system has been found to be activated in response to stressful stimuli. The source of GLP-1 within the brain, the preproglucagon (PPG) neurons, are ideally situated in the brainstem to receive and relay signals of stress and our recent data on the projection pattern of the PPG neurons to the spinal cord suggest a potential strong link with the sympathetic nervous system. We review here the role of central GLP-1 in the regulation of stress responses and discuss the potential involvement of the endogenous source of GLP-1 within the brain, the PPG neurons

The role of nucleus of the solitary tract glucagon-like peptide-1 and prolactin-releasing peptide neurons in stress: anatomy, physiology and cellular interactions

Neuroendocrine, behavioural and autonomic responses to stressful stimuli are orchestrated by complex neural circuits. The caudal nucleus of the solitary tract (cNTS) in the dorsomedial hindbrain is uniquely positioned to integrate signals of both interoceptive and psychogenic stress. Within the cNTS, glucagon-like peptide-1 (GLP-1) and prolactin-releasing peptide (PrRP) neurons play crucial roles in organising neural responses to a broad range of stressors. In this review we discuss the anatomical and functional overlap between PrRP and GLP-1 neurons. We outline their co-activation in response to stressful stimuli and their importance as mediators of behavioural and physiological stress responses. Finally, we review evidence that PrRP neurons are downstream of GLP-1 neurons and outline unexplored areas of the research field. Based on the current state-of-knowledge, PrRP and GLP-1 neurons may be compelling targets in the treatment of stress-related disorders

Distribution and characterisation of Glucagon-like peptide-1 receptor expressing cells in the mouse brain.

© 2015 The Authors.Objective: Although Glucagon-like peptide 1 is a key regulator of energy metabolism and food intake, the precise location of GLP-1 receptors and the physiological relevance of certain populations is debatable. This study investigated the novel GLP-1R-Cre mouse as a functional tool to address this question. Methods: Mice expressing Cre-recombinase under the Glp1r promoter were crossed with either a ROSA26 eYFP or tdRFP reporter strain to identify GLP-1R expressing cells. Patch-clamp recordings were performed on tdRFP-positive neurons in acute coronal brain slices from adult mice and selective targeting of GLP-1R cells in vivo was achieved using viral gene delivery. Results: Large numbers of eYFP or tdRFP immunoreactive cells were found in the circumventricular organs, amygdala, hypothalamic nuclei and the ventrolateral medulla. Smaller numbers were observed in the nucleus of the solitary tract and the thalamic paraventricular nucleus. However, tdRFP positive neurons were also found in areas without preproglucagon-neuronal projections like hippocampus and cortex. GLP-1R cells were not immunoreactive for GFAP or parvalbumin although some were catecholaminergic. GLP-1R expression was confirmed in whole-cell recordings from BNST, hippocampus and PVN, where 100 nM GLP-1 elicited a reversible inward current or depolarisation. Additionally, a unilateral stereotaxic injection of a cre-dependent AAV into the PVN demonstrated that tdRFP-positive cells express cre-recombinase facilitating virally-mediated eYFP expression. Conclusions: This study is a comprehensive description and phenotypic analysis of GLP-1R expression in the mouse CNS. We demonstrate the power of combining the GLP-1R-CRE mouse with a virus to generate a selective molecular handle enabling future in vivo investigation as to their physiological importance

PI3K/mTORC2 regulates TGF-β/Activin signalling by modulating Smad2/3 activity via linker phosphorylation

Crosstalk between the phosphatidylinositol 3-kinase (PI3K) and the transforming growth factor-β signalling pathways play an important role in regulating many cellular functions. However, the molecular mechanisms underpinning this crosstalk remain unclear. Here, we report that PI3K signalling antagonizes the Activin-induced definitive endoderm (DE) differentiation of human embryonic stem cells by attenuating the duration of Smad2/3 activation via the mechanistic target of rapamycin complex 2 (mTORC2). Activation of mTORC2 regulates the phosphorylation of the Smad2/3-T220/T179 linker residue independent of Akt, CDK and Erk activity. This phosphorylation primes receptor-activated Smad2/3 for recruitment of the E3 ubiquitin ligase Nedd4L, which in turn leads to their degradation. Inhibition of PI3K/mTORC2 reduces this phosphorylation and increases the duration of Smad2/3 activity, promoting a more robust mesendoderm and endoderm differentiation. These findings present a new and direct crosstalk mechanism between these two pathways in which mTORC2 functions as a novel and critical mediator

Preproglucagon neurons in the hindbrain have IL-6 Receptor α (IL-6Rα) and show Ca 2+ influx in response to IL-6

Neuronal circuits in the hypothalamus and hindbrain are of importance for control of food intake, energy expenditure, and fat mass. We have recently shown that treatment with exendin-4 (Ex-4), an analog of the proglucagon-derived molecule glucagon-like peptide 1 (GLP-1), markedly increases mRNA expression of the cytokine interleukin-6 (IL-6) in the hypothalamus and hindbrain and that this increase partly mediates the suppression of food intake and body weight by Ex-4. Endogenous GLP-1 in the central nervous system (CNS) is produced by preproglucagon (PPG) neurons of the nucleus of the solitary tract (NTS) in the hindbrain. These neurons project to various parts of the brain, including the hypothalamus. Outside the brain, IL-6 stimulates GLP-1 secretion from the gut and pancreas. In this study, we aim to investigate whether IL-6 can affect GLP-1-producing PPG neurons in the nucleus of the solitary tract (NTS) in mouse hindbrain via the ligand binding part of the IL-6 receptor, IL-6 receptor-α (IL-6Rα). Using immunohistochemistry, we found that IL-6Rα was localized on PPG neurons of the NTS. Recordings of these neurons in GCaMP3/GLP-1 reporter mice showed that IL-6 enhances cytosolic Ca2+ concentration in neurons capable of expressing PPG. We also show that the Ca2+ increase originates from the extracellular space. Furthermore, we found that IL-6Rα was localized on cells in the caudal hindbrain expressing immunoreactive NeuN (a neuronal marker) or CNP:ase (an oligodendrocyte marker). In summary, IL-6Rα is present on PPG neurons in the NTS, and IL-6 can stimulate these cells by increasing influx of Ca2+ to the cytosol from the extracellular space

Serotonergic modulation of the activity of GLP-1 producing neurons in the nucleus of the solitary tract in mouse.

OBJECTIVE: Glucagon-like peptide-1 (GLP-1) and 5-HT are potent regulators of food intake within the brain. GLP-1 is expressed by preproglucagon (PPG) neurons in the nucleus tractus solitarius (NTS). We have previously shown that PPG neurons innervate 5-HT neurons in the ventral brainstem. Here, we investigate whether PPG neurons receive serotonergic input and respond to 5-HT. METHODS: We employed immunohistochemistry to reveal serotonergic innervation of PPG neurons. We investigated the responsiveness of PPG neurons to 5-HT using in vitro Ca²⁺ imaging in brainstem slices from transgenic mice expressing the Ca²⁺ indicator, GCaMP3, in PPG neurons, and cell-attached patch-clamp recordings. RESULTS: Close appositions from 5-HT-immunoreactive axons occurred on many PPG neurons. Application of 20 μM 5-HT produced robust Ca²⁺ responses in NTS PPG dendrites but little change in somata. Dendritic Ca²⁺ spikes were concentration-dependent (2, 20, and 200 μM) and unaffected by blockade of glutamatergic transmission, suggesting 5-HT receptors on PPG neurons. Neither activation nor blockade of 5-HT₃ receptors affected [Ca²⁺]i. In contrast, inhibition of 5-HT₃ receptors attenuated increases in intracellular Ca²⁺ and 5-HT₂c receptor activation produced Ca²⁺ spikes. Patch-clamp recordings revealed that 44% of cells decreased their firing rate under 5-HT, an effect blocked by 5-HT₁ᴀ receptor antagonism. CONCLUSIONS: PPG neurons respond directly to 5-HT with a 5-HT₂c receptor-dependent increase in dendritic [Ca²⁺]i. Electrical responses to 5-HT revealed additional inhibitory effects due to somatic 5-HT₁ᴀ receptors. Reciprocal innervation between 5-HT and PPG neurons suggests that the coordinated activity of these brainstem neurons may play a role in the regulation of food intake.This study was supported by grants MR/J013293/2 from the MRC, UK (ST) and Project Grant #1025031 from NHMRC Australia (ILS). MKH holds a UCL Graduate Research Scholarship. FR and FMG are supported by the Wellcome Trust (106262/Z/14/Z, 106263/Z/14/Z) and the MRC (MRC_MC_UU_12012/3)

Preproglucagon Neurons in the Nucleus of the Solitary Tract are the Main Source of Brain GLP-1, Mediate Stress-Induced Hypophagia, and Limit Unusually Large Intakes of Food

Centrally administered glucagon-like peptide-1 (GLP-1) supresses food intake. Here we demonstrate that GLP-1-producing (PPG) neurons in the nucleus tractus solitarii (NTS) are the predominant source of endogenous GLP-1 within the brain. Selective ablation of NTS PPG neurons by viral expression of diphtheria toxin subunit A (DTA) substantially reduced active GLP-1 concentrations in brain and spinal cord. Contrary to expectations, this loss of central GLP-1 had no significant effect on ad libitum feeding of mice, affecting neither daily chow intake nor body weight or glucose tolerance. Only after bigger challenges to homeostasis were PPG neurons necessary for food intake control. PPG-ablated mice increased food intake following a prolonged fast and after a liquid diet preload. Consistent with our ablation data, acute inhibition of hM4Di-expressing PPG neurons did not affect ad libitum feeding, however, it increased post-fast refeeding intake and blocked stress-induced hypophagia. Additionally, chemogenetic PPG neuron activation through hM3Dq caused a strong acute anorectic effect. We conclude that PPG neurons are not involved in primary intake regulation, but form part of a secondary satiation/satiety circuit, activated by both psychogenic stress and large meals. Given their hypophagic capacity, PPG neurons might be an attractive drug target in obesity treatment

Central and peripheral GLP-1 systems independently suppress eating

The anorexigenic peptide glucagon-like peptide-1 (GLP-1) is secreted from gut enteroendocrine cells and brain preproglucagon (PPG) neurons, which, respectively, define the peripheral and central GLP-1 systems. PPG neurons in the nucleus tractus solitarii (NTS) are widely assumed to link the peripheral and central GLP-1 systems in a unified gut–brain satiation circuit. However, direct evidence for this hypothesis is lacking, and the necessary circuitry remains to be demonstrated. Here we show that PPGNTS neurons encode satiation in mice, consistent with vagal signalling of gastrointestinal distension. However, PPGNTS neurons predominantly receive vagal input from oxytocin-receptor-expressing vagal neurons, rather than those expressing GLP-1 receptors. PPGNTS neurons are not necessary for eating suppression by GLP-1 receptor agonists, and concurrent PPGNTS neuron activation suppresses eating more potently than semaglutide alone. We conclude that central and peripheral GLP-1 systems suppress eating via independent gut–brain circuits, providing a rationale for pharmacological activation of PPGNTS neurons in combination with GLP-1 receptor agonists as an obesity treatment strategy

Conducting Health Research in Korean American Churches: Perspectives from Church Leaders

Korean Americans experience many challenges to obtaining adequate health care coverage and access to needed services. Because a large proportion of Korean Americans attend churches on a regular basis, churches may be a promising venue where health programs can be delivered. In order to gain an in-depth understanding of Korean American churches with respect to conducting future health intervention research, we conducted exploratory interviews and focus groups with 58 leaders from 23 Korean American churches and three community organizations. From these interviews and focus groups, we found that Korean churches and church leaders seek to meet a variety of social and health needs of their congregation and their surrounding community. Several leaders have stated that assisting with social and medical needs of their members is an important component of their current ministry. They described profound health needs of their congregations and have suggested various ways in which the university can partner with the local churches to help address these needs through research. Additionally, they described various resources churches can provide to researchers such as: their personal assistance, church volunteer base, church facility, and church network and contacts. Our findings suggest that Korean churches have a high potential to serve an important role in the health of Korean Americans. On the basis of the promising results of the present study, we are planning to conduct a cross sectional survey of Korean church leaders and members in Los Angeles County to substantiate our findings in a larger representative sample