Shi Z, Cassaglia PA, Pelletier NE, Brooks VL.
PMID: PMID: 30628058 | DOI: DOI:10.1113/JP277517
KEY POINTS: ICV insulin increased SNA and baroreflex control of SNA and HR dramatically more in obese male rats; in obese females, the responses were abolished. In obese males, the enhanced LSNA responses were associated with reduced tonic inhibition of LSNA by NPY in the PVN. Yet, PVN NPY injection decreased LSNA similarly in OP/OR/CON rats. Collectively, these results suggest that NPY inputs were decreased. In obese females, NPY inhibition in the PVN was maintained. Moreover, NPY neurons in the ArcN became resistant to the inhibitory effects of insulin. A HFD did not alter arcuate NPY neuronal InsR expression in males or females. Obesity-induced "selective sensitization" of the brain to the sympathoexcitatory effects of insulin and leptin may contribute to elevated basal SNA, and therefore hypertension development, in males with obesity. These data may explain in part why obesity increases SNA less in women compared to men. ABSTRACT: Obesity increases sympathetic nerve activity (SNA) in men, but not women; however, the mechanisms are unknown. We tested if intracerebroventricular insulin infusion increases SNA more in obese male than female rats and if sex differences are mediated by changes in tonic inhibition of SNA by Neuropeptide Y (NPY) in the paraventricular nucleus (PVN). When consuming a high fat diet, obesity prone (OP) rats accrued excess fat, whereas obesity resistant (OR) rats maintained adiposity as in rats eating a control (CON) diet. Insulin increased lumbar SNA (LSNA) similarly in CON/OR males and females under urethane-anesthesia. The LSNA response was magnified in OP males, but abolished in OP females. In males, blockade of PVN NPY Y1 receptors with BIBO3304 increased LSNA in CON/OR rats, but not OP rats. Yet, PVN nanoinjections of NPY decreased LSNA similarly between groups. Thus, tonic PVN NPY inhibition of LSNA may be lost in obese males, due to a decrease in NPY inputs. In contrast, in females, PVN BIBO3304 increased LSNA similarly in OP, OR and CON rats. After insulin, PVN BIBO3304 failed to increase LSNA in CON/OR females, but increased LSNA in OP females, suggesting that with obesity NPY neurons become resistant to the inhibitory effects of insulin. These sex differences were not associated with changes in arcuate NPY neuronal insulin receptor expression. Collectively, these data reveal a marked sex difference in the impact of obesity on insulin's sympathoexcitatory actions and implicate sexually dimorphic changes in NPY inhibition of SNA in the PVN as one mechanism.
Zhang N, Zhang HY, Bi SA, Moran TH and Bi S
PMID: 30902570 | DOI: 10.1016/j.neulet.2019.03.030
Thyrotropin-releasing hormone (TRH) plays an important role in the regulation of energy balance. While the regulation of TRH in the paraventricular nucleus (PVN) in response to changes of energy balance has been well studied, how TRH is regulated in the dorsomedial hypothalamus (DMH) in maintaining energy homeostasis remains unclear. Here, we assessed the effects of food restriction and exercise on hypothalamic Trh expression using Otsuka Long-Evens Tokushima Fatty (OLETF) rats. Sedentary ad lib fed OLETF rats (OLETF-SED) became hyperphagic and obese. These alterations were prevented in OLETF rats with running wheel access (OLETF-RW) or food restriction in which their food was pair-fed (OLETF-PF) to the intake of lean control rats (LETO-SED). Evaluation of hypothalamic gene expression revealed that Trh mRNA expression was increased in the PVN of OLETF-SED rats and normalized in OLETF-RW and OLETF-PF rats compared to LETO-SED rats. In contrast, the expression of Trh in the DMH was decreased in OLETF-SED rats relative to LETO-SED rats. This alteration was reversed in OLETF-RW rats as seen in LETO-SED rats, but food restriction resulted in a significant increase in DMH Trh expression in OLETF-PF rats compared to LETO-SED rats. Strikingly, while Trh mRNA expression was decreased in the PVN of intact rats in response to acute food deprivation, food deprivation resulted in increased expression of Trh in the DMH. Together, these results demonstrate the differential regulation of Trh expression in the PVN and DMH in OLETF rats and suggest that DMH TRH also contributes to hypothalamic regulation of energy balance.
Kume, M;Ahmad, A;DeFea, KA;Vagner, J;Dussor, G;Boitano, S;Price, TJ;
PMID: 37315729 | DOI: 10.1016/j.jpain.2023.06.006
Chemotherapy-Induced Peripheral Neuropathy (CIPN) is a common, dose-limiting side effect of cancer therapy. Protease-activated receptor 2 (PAR2) is implicated in a variety of pathologies, including CIPN. In this study, we demonstrate the role of PAR2 expressed in sensory neurons in a paclitaxel (PTX)-induced model of CIPN in mice. PAR2 knockout/WT mice and mice with PAR2 ablated in sensory neurons were treated with paclitaxel administered via intraperitoneal injection. In vivo behavioral studies were done in mice using von Frey filaments and the Mouse Grimace Scale. We then examined immunohistochemical staining of dorsal root ganglion (DRG) and hind paw skin samples from CIPN mice to measure satellite cell gliosis and intra-epidermal nerve fiber (IENF) density. Pharmacological reversal of CIPN pain was tested with the PAR2 antagonist C781. Mechanical allodynia caused by paclitaxel treatment was alleviated in PAR2 knockout mice of both sexes. In the PAR2 sensory neuronal conditional knockout (cKO) mice, both mechanical allodynia and facial grimacing were attenuated in mice of both sexes. In the dorsal root ganglion of the paclitaxel-treated PAR2 cKO mice, satellite glial cell activation was reduced compared to control mice. IENF density analysis of the skin showed that the paclitaxel-treated control mice have a reduction in nerve fiber density while the PAR2 cKO mice had a comparable skin innervation as the vehicle-treated animals. Similar results were seen with satellite cell gliosis in the DRG where gliosis induced by PTX was absent in PAR cKO mice. Finally, C781 was able to transiently reverse established PTX-evoked mechanical allodynia. PERSPECTIVE: Our work demonstrates that PAR2 expressed in sensory neurons plays a key role in paclitaxel-induced mechanical allodynia, spontaneous pain and signs of neuropathy, suggesting PAR2 as a possible therapeutic target in multiple aspects of paclitaxel CIPN.
Haidar M, Tin K, Zhang C, Nategh M, Covita J, Wykes AD, Rogers J and Gundlach AL
PMID: 30906254 | DOI: 10.3389/fnana.2019.00030
Relaxin-3 is a highly conserved neuropeptide abundantly expressed in neurons of the nucleus incertus (NI), which project to nodes of the septohippocampal system (SHS) including the medial septum/diagonal band of Broca (MS/DB) and dorsal hippocampus, as well as to limbic circuits. High densities of the Gi/o-protein-coupled receptor for relaxin-3, known as relaxin-family peptide-3 receptor (RXFP3) are expressed throughout the SHS, further suggesting a role for relaxin-3/RXFP3 signaling in modulating learning and memory processes that occur within these networks. Therefore, this study sought to gain further anatomical and functional insights into relaxin-3/RXFP3 signaling in the mouse MS/DB. Using Cre/LoxP recombination methods, we assessed locomotion, exploratory behavior, and spatial learning and long-term reference memory in adult C57BL/6J Rxfp3 (loxP/loxP) mice with targeted depletion of Rxfp3 in the MS/DB. Following prior injection of an AAV((1/2))-Cre-IRES-eGFP vector into the MS/DB to delete/deplete Rxfp3 mRNA/RXFP3 protein, mice tested in a Morris water maze (MWM) displayed an impairment in allocentric spatial learning during acquisition, as well as an impairment in long-term reference memory on probe day. However, RXFP3-depleted and control mice displayed similar motor activity in a locomotor cell and exploratory behavior in a large open-field (LOF) test. A quantitative characterization using multiplex, fluorescent in situ hybridization (ISH) identified a high level of co-localization of Rxfp3 mRNA and vesicular GABA transporter (vGAT) mRNA in MS and DB neurons (~87% and ~95% co-expression, respectively). Rxfp3 mRNA was also detected, to a correspondingly lesser extent, in vesicular glutamate transporter 2 (vGlut2) mRNA-containing neurons in MS and DB (~13% and ~5% co-expression, respectively). Similarly, a qualitative assessment of the MS/DB region, identified Rxfp3 mRNA in neurons that expressed parvalbumin (PV) mRNA (reflecting hippocampally-projecting GABA neurons), whereas choline acetyltransferase mRNA-positive (acetylcholine) neurons lacked Rxfp3 mRNA. These data are consistent with a qualitative immunohistochemical analysis that revealed relaxin-3-immunoreactive nerve fibers in close apposition with PV-immunoreactive neurons in the MS/DB. Together these studies suggest relaxin-3/RXFP3 signaling in the MS/DB plays a role in modulating specific learning and long-term memory associated behaviors in adult mice via effects on GABAergic neuron populations known for their involvement in modulating hippocampal theta rhythm and associated cognitive processes.
Lemos JC, Shin JH, Alvarez VA.
PMID: 31109960 | DOI: 10.1523/JNEUROSCI.0479-19.2019
Cholinergic interneurons (CINs) are critical regulators of striatal network activity and output. Changes in CIN activity are thought to encode salient changes in the environment and stimulus-response-outcome associations. Here we report that the stress-associated neuropeptide corticotropin releasing factor (CRF) produces a profound and reliable increase in the spontaneous firing of CINs in both dorsal striatum and nucleus accumbens (NAc) through activation of CRF type 1 receptors, production of cAMP and reduction in spike accommodation in male mice. The increase of CIN firing by CRF results in the activation muscarinic acetylcholine receptors type 5, which mediate potentiation of dopamine transmission in the striatum. This study provides critical mechanistic insight into how CRF modulates striatal activity and dopamine transmission in the NAc to likely account for CRF facilitation of appetitive behaviors.SIGNIFICANCE STATEMENT Although the presence of CRF receptors in the dorsal and ventral striatum has been acknowledged, the cellular identity and the functional consequences of receptor activation is unknown. Here we report that striatal cholinergic interneurons express CRF-R1 receptors and are acutely activated by the neuropeptide CRF that is released in response to salient environmental stimuli. Cholinergic interneurons make <1% of the cells in the striatum but are critical regulators of the striatal circuitry and its output. CRF's fast and potent activation of cholinergic interneurons could have far reaching behavioral implications across motivated behaviors controlled by the striatum.
Proc Natl Acad Sci U S A.
Labouesse MA, Sartori AM, Weinmann O, Simpson EH, Kellendonk C, Weber-Stadlbauer U.
PMID: 30254156 | DOI: 10.1073/pnas.1800171115
Dopaminergic signaling in the striatum, particularly at dopamine 2 receptors (D2R), has been a topic of active investigation in obesity research in the past decades. However, it still remains unclear whether variations in striatal D2Rs modulate the risk for obesity and if so in which direction. Human studies have yielded contradictory findings that likely reflect a complex nonlinear relationship, possibly involving a combination of causal effects and compensatory changes. Animal work indicates that although chronic obesogenic diets reduce striatal D2R function, striatal D2R down-regulation does not lead to obesity. In this study, we evaluated the consequences of striatal D2R up-regulation on body-weight gain susceptibility and energy balance in mice. We used a mouse model of D2R overexpression (D2R-OE) in which D2Rs were selectively up-regulated in striatal medium spiny neurons. We uncover a pathological mechanism by which striatal D2R-OE leads to reduced brown adipose tissue thermogenesis, reduced energy expenditure, and accelerated obesity despite reduced eating. We also show that D2R-OE restricted to development is sufficient to promote obesity and to induce energy-balance deficits. Together, our findings indicate that striatal D2R-OE during development persistently increases the propensity for obesity by reducing energy output in mice. This suggests that early alterations in the striatal dopamine system could represent a key predisposition factor toward obesity.
Cannabidiol produces distinct U-shaped dose-response effects on cocaine conditioned place preference and associated recruitment of prelimbic neurons in male rats
Biological Psychiatry Global Open Science
Nedelescu, H;Wagner, G;De Ness, G;Carrol, A;Kerr, T;Wang, J;Zhang, S;Chang, S;Than, A;Emerson, N;Suto, N;Weiss, F;
| DOI: 10.1016/j.bpsgos.2021.06.014
Background Cannabidiol (CBD) has received attention for the treatment of Substance Use Disorders. In preclinical models of relapse, CBD attenuates drug seeking across several drugs of abuse, including cocaine. However, in these models, CBD has not been consistently effective. This inconsistency in CBD effects may be related to presently insufficient information on the full spectrum of CBD dose effects on drug-related behaviors. Methods We address this issue by establishing a full dose-response profile of CBD’s actions using expression of cocaine-induced conditioned place preference (CPP) as a model for drug motivated behavior in male rats, and by concurrently identifying dose-dependent effects of CBD on underlying neuronal activation as well as distinct neuronal phenotypes showing dose-dependent activation changes. Additionally, CBD levels in plasma and brain were established. Results CBD produced linear increases in CBD brain/plasma concentrations but suppressed CPP in a distinct U-shaped manner. In parallel with its behavioral effects, CBD produced U-shaped suppressant effects on neuronal activation in the prelimbic but not infralimbic cortex or nucleus accumbens core and shell. RNAscope in situ hybridization identified suppression of glutamatergic and GABAergic signaling in the prelimbic cortex as a possible cellular mechanism for the attenuation of cocaine CPP by CBD. Conclusions The findings extend previous evidence on the potential of CBD in preventing drug motivated behavior. However, CBD’s dose-response profile may have important dosing implications for future clinical applications and may contribute to the understanding of discrepant CBD effects on drug seeking in the literature.
Zhang, L;Koller, J;Gopalasingam, G;Qi, Y;Herzog, H;
PMID: 35691527 | DOI: 10.1016/j.molmet.2022.101525
Neuropeptide FF (NPFF) group peptides belong to the evolutionary conserved RF-amide peptide family. While they have been assigned a role as pain modulators, their roles in other aspects of physiology have received much less attention. NPFF peptides and their receptor NPFFR2 have strong and localized expression within the dorsal vagal complex that has emerged as the key centre for regulating glucose homeostasis. Therefore, we investigated the role of the NPFF system in the control of glucose metabolism and the histochemical and molecular identities of NPFF and NPFFR2 neurons.We examined glucose metabolism in Npff-/- and wild type (WT) mice using intraperitoneal (i.p.) glucose tolerance and insulin tolerance tests. Body composition and glucose tolerance was further examined in mice after 1-week and 3-week of high-fat diet (HFD). Using RNAScope double ISH, we investigated the neurochemical identity of NPFF and NPFFR2 neurons in the caudal brainstem, and the expression of receptors for peripheral factors in NPFF neurons.Lack of NPFF signalling in mice leads to improved glucose tolerance without significant impact on insulin excursion after the i.p. glucose challenge. In response to an i.p. bolus of insulin, Npff-/- mice have lower glucose excursions than WT mice, indicating an enhanced insulin action. Moreover, while HFD has rapid and potent detrimental effects on glucose tolerance, this diet-induced glucose intolerance is ameliorated in mice lacking NPFF signalling. This occurs in the absence of any significant impact of NPFF deletion on lean or fat masses, suggesting a direct effect of NPFF signalling on glucose metabolism. We further reveal that NPFF neurons in the subpostrema area (SubP) co-express receptors for peripheral factors involved in glucose homeostasis regulation such as insulin and GLP1. Furthermore, Npffr2 is expressed in the glutamatergic NPFF neurons in the SubP, and in cholinergic neurons of the dorsal motor nucleus of the vagus (DMV), indicating that central NPFF signalling is likely modulating vagal output to innervated peripheral tissues including those important for glucose metabolic control.NPFF signalling plays an important role in the regulation of glucose metabolism. NPFF neurons in the SubP are likely to receive peripheral signals and mediate the control of whole-body glucose homeostasis via centrally vagal pathways. Targeting NPFF and NPFFR2 signalling may provide a new avenue for treating type 2 diabetes and obesity.
The Journal of neuroscience : the official journal of the Society for Neuroscience
Ambler, M;Hitrec, T;Wilson, A;Cerri, M;Pickering, A;
PMID: 35440490 | DOI: 10.1523/JNEUROSCI.2102-21.2022
Torpor is a naturally occurring, hypometabolic, hypothermic state engaged by a wide range of animals in response to imbalance between the supply and demand for nutrients. Recent work has identified some of the key neuronal populations involved in daily torpor induction in mice, in particular projections from the preoptic area of the hypothalamus (POA) to the dorsomedial hypothalamus (DMH). The DMH plays a role in thermoregulation, control of energy expenditure, and circadian rhythms, making it well positioned to contribute to the expression of torpor. We used activity dependent genetic TRAPing techniques to target DMH neurons that were active during natural torpor bouts in female mice. Chemogenetic reactivation of torpor-TRAPed DMH neurons in calorie-restricted mice promoted torpor, resulting in longer and deeper torpor bouts. Chemogenetic inhibition of torpor-TRAPed DMH neurons did not block torpor entry, suggesting a modulatory role for the DMH in the control of torpor. This work adds to the evidence that the POA and the DMH form part of a circuit within the mouse hypothalamus that controls entry into daily torpor.SIGNIFICANCEDaily heterotherms such as mice employ torpor to cope with environments in which the supply of metabolic fuel is not sufficient for the maintenance of normothermia. Daily torpor involves reductions in body temperature, as well as active suppression of heart rate and metabolism. How the central nervous system controls this profound deviation from normal homeostasis is not known, but a projection from the preoptic area to the dorsomedial hypothalamus has recently been implicated. We demonstrate that the dorsomedial hypothalamus contains neurons that are active during torpor. Activity in these neurons promotes torpor entry and maintenance, but their activation alone does not appear to be sufficient for torpor entry.
Journal of cellular physiology
Zhang, CL;Lin, YZ;Wu, Q;Yan, C;Wong, MW;Zeng, F;Zhu, P;Bowes, K;Lee, K;Zhang, X;Song, ZY;Lin, S;Shi, YC;
PMID: 35312067 | DOI: 10.1002/jcp.30719
Chronic high salt intake is one of the leading causes of hypertension. Salt activates the release of the key neurotransmitters in the hypothalamus such as vasopressin to increase blood pressure, and neuropepetide Y (NPY) has been implicated in the modulation of vasopressin levels. NPY in the hypothalamic arcuate nucleus (Arc) is best known for its control in appetite and energy homeostasis, but it is unclear whether it is also involved in the development of salt-induced hypertension. Here, we demonstrate that wild-type mice given 2% NaCl salt water for 8 weeks developed hypertension which was associated with marked downregulation of NPY expression in the hypothalamic Arc as demonstrated in NPY-GFP reporter mice as well as by in situ hybridization analysis. Furthermore, salt intake activates neurons in the hypothalamic paraventricular nucleus (PVN) where mRNA expression of brain-derived neurotrophic factor (BDNF) and vasopressin was found to be upregulated, leading to elevated serum vasopressin levels. This finding suggests an inverse correlation between the Arc NPY level and expression of vasopressin and BDNF in the PVN. Specific restoration of NPY by injecting AAV-Cre recombinase into the Arc only of the NPY-targeted mutant mice carrying a loxP-flanked STOP cassette reversed effects of salt intake on vasopressin and BDNF expression, leading to a normalization of salt-dependent blood pressure. In summary, our study uncovers an important Arc NPY-originated neuronal circuitry that could sense and respond to peripheral electrolyte signals and thereby regulate hypertension via vasopressin and BDNF in the PVN.
Harris NA, Isaac AT, Günther A, Merkel K, Melchior J, Xu M, Eguakun E, Perez R, Nabit BP, Flavin S, Gilsbach R, Shonesy B, Hein L, Abel T, Baumann A, Matthews R, Centanni SW, Winder DG.
PMID: 30150361 | DOI: 10.1523/JNEUROSCI.0963-18.2018
Stress is a precipitating agent in neuropsychiatric disease and initiates relapse to drug-seeking behavior in addicted patients. Targeting the stress system in protracted abstinence from drugs of abuse with anxiolytics may be an effective treatment modality for substance use disorders. α2A-adrenergic receptors (α2A-ARs) in extended amygdala structures play key roles in dampening stress responses. Contrary to early thinking, α2A-ARs are expressed at non-noradrenergic sites in the brain. These non-noradrenergic α2A-ARs play important roles in stress-responses, but their cellular mechanisms of action are unclear. In humans, the α2A-AR agonist guanfacine reduces overall craving and uncouples craving from stress yet minimally affects relapse, potentially due to competing actions in the brain. Here we show that heteroceptor α2A-ARs postsynaptically enhance dorsal BNST (dBNST) neuronal activity in mice of both sexes. This effect is mediated by hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels, as inhibition of these channels is necessary and sufficient for excitatory actions. Finally, this excitatory action is mimicked by clozapine-N-oxide activation of the Gi-coupled DREADD hM4Di in dBNST neurons, and its activation elicits anxiety-like behavior in the elevated plus maze. Together, this data provides a framework for elucidating cell-specific actions of GPCR signaling and provides a potential mechanism whereby competing anxiogenic and anxiolytic actions of guanfacine may affect its clinical utility in the treatment of addiction.SIGNIFICANCE STATEMENTStress impacts the development of neuropsychiatric disorders including anxiety and addiction. Guanfacine is an α2A-adrenergic receptor (α2A-AR) agonist with actions in the bed nucleus of the stria terminalis (BNST) that produces antidepressant actions and uncouples stress from reward-related behaviors. Here we show that guanfacine increases dBNST neuronal activity through actions at postsynaptic α2A-ARs via a mechanism that involves hyperpolarization-activated cyclic nucleotide gated cation (HCN) channels. This action is mimicked by activation of the designer receptor hM4Di expressed in the BNST, which also induces anxiety-like behaviors. Together, these data suggest 1) that postsynaptic α2A-ARs in BNST have excitatory actions on BNST neurons, and 2) these actions can be phenocopied by the so-called "inhibitory" DREADDs, suggesting care must be taken regarding interpretation of data obtained with these tools.
The Journal of physiology
Shi, Y;Sobrinho, CR;Soto-Perez, J;Milla, BM;Stornetta, DS;Stornetta, RL;Takakura, AC;Mulkey, DK;Moreira, TS;Bayliss, DA;
PMID: 35385139 | DOI: 10.1113/JP282279
A brainstem homeostatic system senses CO2 /H+ to regulate ventilation, blood gases and acid-base balance. Neurons of the retrotrapezoid nucleus (RTN) and medullary raphe are both implicated in this mechanism as respiratory chemosensors, but recent pharmacological work suggested that the CO2 /H+ sensitivity of RTN neurons is mediated indirectly, by raphe-derived serotonin acting on 5-HT7 receptors. To investigate this further, we characterized Htr7 transcript expression in phenotypically identified RTN neurons using multiplex single cell qRT-PCR and RNAscope. Although present in multiple neurons in the parafacial region of the ventrolateral medulla, Htr7 expression was undetectable in most RTN neurons (Nmb+ /Phox2b+ ) concentrated in the densely packed cell group ventrolateral to the facial nucleus. Where detected, Htr7 expression was modest and often associated with RTN neurons that extend dorsolaterally to partially encircle the facial nucleus. These dorsolateral Nmb+ /Htr7+ neurons tended to express Nmb at high levels and the intrinsic RTN proton detectors Gpr4 and Kcnk5 at low levels. In mouse brainstem slices, CO2 -stimulated firing in RTN neurons was mostly unaffected by a 5-HT7 receptor antagonist, SB269970 (n = 11/13). At the whole animal level, microinjection of SB269970 into the RTN of conscious mice blocked respiratory stimulation by co-injected LP-44, a 5-HT7 receptor agonist, but had no effect on CO2 -stimulated breathing in those same mice. We conclude that Htr7 is expressed by a minor subset of RTN neurons with a molecular profile distinct from the established chemoreceptors and that 5-HT7 receptors have negligible effects on CO2 -evoked firing activity in RTN neurons or on CO2 -stimulated breathing in mice. KEY POINTS: Neurons of the retrotrapezoid nucleus (RTN) are intrinsic CO2 /H+ chemosensors and serve as an integrative excitatory hub for control of breathing. Serotonin can activate RTN neurons, in part via 5-HT7 receptors, and those effects have been implicated in conferring an indirect CO2 sensitivity. Multiple single cell molecular approaches revealed low levels of 5-HT7 receptor transcript expression restricted to a limited population of RTN neurons. Pharmacological experiments showed that 5-HT7 receptors in RTN are not required for CO2 /H+ -stimulation of RTN neuronal activity or CO2 -stimulated breathing. These data do not support a role for 5-HT7 receptors in respiratory chemosensitivity mediated by RTN neurons.
