Brown adipose tissue (BAT) plays a critical role in cold- and diet-induced thermogenesis. Although BAT is densely innervated by the sympathetic nervous system (SNS), little is known about the central nervous system (CNS) origins of this innervation. The purpose of the present experiment was to determine the neuroanatomic chain of functionally connected neurons from the CNS to BAT. A transneuronal viral tract tracer, Bartha’s K strain of the pseudorabies virus (PRV), was injected into the interscapular BAT of Siberian hamsters. The animals were killed 4 and 6 days postinjection, and the infected neurons were visualized by immunocytochemistry. PRV-infected neurons were found in the spinal cord, brain stem, midbrain, and forebrain. The intensity of labeled neurons in the forebrain varied from heavy infections in the medial preoptic area and paraventricular hypothalamic nucleus to few infections in the ventromedial hypothalamic nucleus, with moderate infections in the suprachiasmatic and lateral hypothalamic nuclei. These results define the SNS outflow from the brain to BAT for the first time in any species.

The innervation of brown adipose tissue (BAT) by the sympathetic nervous system (SNS) is incontrovertible and, with its activation, functions as the principal, if not exclusive, stimulator of BAT thermogenesis. The parasympathetic innervation of BAT only appears in two minor BAT depots, but not in the major interscapular BAT (IBAT) depot. BAT thermogenesis is triggered by the release of norepinephrine from its sympathetic nerve terminals, stimulating β3-adrenoceptors that turns on a cascade of intracellular events ending in activation of uncoupling protein-1 (UCP-1). BAT also has sensory innervation that may function to monitor BAT lipolysis, a response necessary for activation of UCP-1 by fatty acids, or perhaps responding in a feedback manner to BAT temperature changes. The central sympathetic outflow circuits ultimately terminating in BAT have been revealed by injecting the retrograde viral transneuronal tract tracer, pseudorabies virus, into the tissue; moreover, there is a high degree of colocalization of melanocortin 4-receptor mRNA on these neurons across the neural axis. The necessary and sufficient central BAT SNS outflow sites that are activated by various thermogenic stimuli are not precisely known. In a chronic decerebration procedure, IBAT UCP-1 gene expression can be triggered by fourth ventricular injections of melanotan II, the melanocortin 3/4 receptor agonist, suggesting that there is sufficient hindbrain neural circuitry to generate thermogenic responses with this stimulation. The recent recognition of BAT in normal adult humans suggests a potential target for stimulation of energy expenditure by BAT to help mitigate increased body fat storage.

Summary Circulating factors are typically invoked to explain bidirectional communication between the CNS and white adipose tissue (WAT). Thus, initiation of lipolysis has been relegated primarily to adrenal medullary secreted catecholamines and the inhibition of lipolysis primarily to pancreatic insulin, whereas signals of body fat levels to the brain have been ascribed to adipokines such as leptin. By contrast, evidence is given for bidirectional communication between brain and WAT occurring via the sympathetic nervous system (SNS) and sensory innervation of this tissue. Using retrograde transneuronal viral tract tracers, the SNS outflow from brain to WAT has been defined. Functionally, sympathetic denervation of WAT blocks lipolysis to a variety of lipolytic stimuli. Using anterograde transneuronal viral tract tracers, the sensory input from WAT to brain has been defined. Functionally, these WAT sensory nerves respond electrophysiologically to increases in WAT SNS drive suggesting a possible neural negative feedback loop to regulate lipolysis.

. Sympathetic innervation of white adipose tissue and its regulation of fat cell number. Am J Physiol Regul Integr Comp Physiol 286: R1167-R1175, 2004 10.1152/ajpregu.00558.2003.-White adipose tissue (WAT) is innervated by the sympathetic nervous system (SNS), and the central origins of this innervation have been demonstrated for inguinal and epididymal WAT (iWAT and eWAT, respectively) using a viral transneuronal tract tracer, the pseudorabies virus (PRV). Although the more established role of this sympathetic innervation of WAT is as a major stimulator of lipid mobilization, this innervation also inhibits WAT fat cell number (FCN); thus, local denervation of WAT leads to marked increases in WAT mass and FCN. The purpose of this study was to extend our understanding of the SNS regulation of FCN using neuroanatomical and functional analyses. Therefore, we injected PRV into retroperitoneal WAT (rWAT) to compare the SNS outflow to this pad from what already is known for iWAT and eWAT. In addition, we tested the ability of local unilateral denervation of rWAT or iWAT to promote increases in WAT mass and FCN vs. their contralateral neurally intact counterparts. Although the overall pattern of innervation was more similar than different for rWAT vs. iWAT or eWAT, its SNS outflow appeared to involve more neurons in the suprachiasmatic and solitary tract nuclei. Denervation produced significant increases in WAT mass and FCN for both iWAT and rWAT, but FCN was increased significantly more in iWAT than in rWAT. These data suggest differences in origins of the sympathetic outflow to WAT and functional differences in the WAT SNS innervation that could contribute to the differential propensity for fat cell proliferation across WAT depots in vivo. body fat; cellularity; hyperplasia; pseudorabies virus; tract tracing; white fat ENERGY BALANCE IS MAINTAINED through alterations in energy intake and energy expenditure. Lipid is the primary form of stored energy in mammals and is largely found in white adipose tissue (WAT) as triacylglycerols. Prolonged positive energy balance promotes obesity that in humans is associated with a number of health risks (34). The deposition of lipid initially results in increases in fat cell size (FCS), but soon triggers increases in fat cell number (FCN; for review, see Ref. 16). The processes underlying this hypercellularity, a hallmark sign of obesity, are not well understood (for review, see Ref. 21). Furthermore, the accretion of lipid in and of itself is not necessarily associated with these health risks (38); rather it is the distribution of body fat that is critical (19). That is, visceral obesity is closely associated with development of the "metabolic syndrome" and type II diabetes (for review, see Ref. 44). Thus understanding how body fat is preferentially deposited and mobilized is necessary for the prevention and treatment of obesity, respectively.One mechanism that may underlie the differential accumulation of lipid by WAT depots, as well as the differential mobilization of lipid, i...

Song CK, Vaughan CH, Keen-Rhinehart E, Harris RB, Richard D, Bartness TJ. Melanocortin-4 receptor mRNA expressed in sympathetic outflow neurons to brown adipose tissue: neuroanatomical and functional evidence. Am J Physiol Regul Integr Comp Physiol 295: R417-R428, 2008. First published June 11, 2008 doi:10.1152/ajpregu.00174.2008.-A precise understanding of neural circuits controlling lipid mobilization and thermogenesis remains to be determined. We have been studying the sympathetic nervous system (SNS) contributions to white adipose tissue (WAT) lipolysis largely in Siberian hamsters. Central melanocortins are implicated in the control of the sympathetic outflow to WAT, and, moreover, the melanocortin 4 receptors (MC4-R) appear to be principally involved. We previously found that acute third ventricular melanotan II (MTII; an MC3/4-R agonist) injections increase sympathetic drive (norepinephrine turnover) to interscapular brown adipose tissue (IBAT) and IBAT temperature. Here we tested whether MC4-R mRNA is expressed in IBAT SNS outflow neurons using in situ hybridization for the former and injections of the transneuronal viral retrograde tract tracer, pseudorabies virus (PRV) into IBAT, for the latter. Significant numbers of double-labeled cells for PRV and MC4-R mRNA were found across the neuroaxis (mean of all brain sites ϳ60%), including the hypothalamic paraventricular nucleus (PVH; ϳ80%). Acute parenchymal MTII microinjections into the PVH of awake, freely-moving hamsters, using doses below those able to increase IBAT temperature when injected into the third ventricle, increased IBAT temperature for as long as 4 h, as measured by temperature transponders implanted below the tissue. Collectively, these data add significant support to the view that central melanocortins are important in controlling IBAT thermogenesis via the SNS innervation of this tissue, likely through the MC4-Rs.Siberian hamsters; in situ hybridization; pseudorabies virus; tract tracing; melanocortins A COMPLETE UNDERSTANDING OF the neural circuits controlling the sympathetic nervous system (SNS) mobilization of lipid from white adipose tissue (WAT) and those triggering thermogenesis in brown adipose tissue (BAT) remain to be determined, although our knowledge has increased markedly in the past decade (for review, see Refs. 6 and 16). We have been studying the SNS contributions to the reversal of the naturally-occurring seasonal obesity of Siberian hamsters (Phodopus sungorus; for a review, see Ref. 5) in WAT, and to a lesser extent BAT (4,11,12,18,49). Of the many remaining unknown details of the SNS outflow circuitry to WAT and BAT is the identification of their neurochemical phenotype, although some progress has been made for both tissues (e.g., 8,39,48,51,53,56). One of the neurochemicals strongly implicated in the sympathetic control of these adipose tissues is the melanocortins (for review, see Refs. 6 and 45). The most important members of the melanocortin family for the control of energy balance is ␣-melanocyte stimulating hor...

Melanocortins are implicated in the control of energy intake/expenditure. Centrally administered melanotan II (MTII), a synthetic melanocortin 3/4-receptor agonist, decreases adiposity beyond that accountable by food intake decreases. Melanocortin-4 receptor (MC4-R) mRNA is expressed on sympathetic nervous system (SNS) outflow neurons to white adipose tissue (WAT) in Siberian hamsters, suggesting a role in lipid mobilization. Therefore, we tested whether third ventricular injections of MTII increased sympathetic drive to WAT and interscapular brown adipose tissue (IBAT) using norepinephrine turnover (NETO) as a measure of sympathetic drive. We also tested for MTII-induced changes in lipolysis-related WAT gene expression (beta3-adrenoceptors, hormone sensitive lipase) and IBAT thermogenesis (beta3-adrenoceptor, uncoupling protein-1). Finally, we tested whether third ventricularly injected MTII, a highly selective MC4-R agonist (cyclo[beta-Ala-His-D-Phe-Arg-Trp-Glu]NH2) increased or agouti-related protein decreased IBAT temperature in hamsters implanted with sc IBAT temperature transponders. Centrally administered MTII provoked differential sympathetic drives to WAT and IBAT (increased inguinal WAT, dorsosubcutaneous WAT and IBAT NETO, but not epididymal WAT and retroperitoneal WAT NETO). MTII also increased circulating concentrations of the lipolytic products free fatty acids and glycerol but not plasma catecholamines, suggesting lipid mobilization via WAT SNS innervation and not via adrenal medullary catecholamines. WAT or IBAT gene expression was largely unaffected by acute MTII treatment, but IBAT temperature was increased by MTII and the MC4-R agonist and decreased by agouti-related protein. Collectively, this is the first demonstration of central melanocortin agonist stimulation of WAT lipolysis through the SNS and confirms melanocortin-induced changes in BAT thermogenesis.

Converging evidence indicates that white adipose tissue (WAT) is innervated by the sympathetic nervous system (SNS) based on immunohistochemical labeling of a SNS marker (tyrosine hydroxylase [TH]), tract tracing of WAT sympathetic postganglionic innervation, pseudorabies virus (PRV) transneuronal labeling of WAT SNS outflow neurons, and functional evidence from denervation studies. Recently, WAT para-SNS (PSNS) innervation was suggested because local surgical WAT sympathectomy (sparing hypothesized parasympathetic innervation) followed by PRV injection yielded infected cells in the vagal dorsomotor nucleus (DMV), a traditionally-recognized PSNS brain stem site. In addition, local surgical PSNS WAT denervation triggered WAT catabolic responses. We tested histologically whether WAT was parasympathetically innervated by searching for PSNS markers in rat, and normal (C57BL) and obese (ob/ob) mouse WAT. Vesicular acetylcholine transporter, vasoactive intestinal peptide and neuronal nitric oxide synthase immunoreactivities were absent in WAT pads (retroperitoneal, epididymal, inguinal subcutaneous) from all animals. Nearly all nerves innervating WAT vasculature and parenchyma that were labeled with protein gene product 9.5 (PGP9.5; pan-nerve marker) also contained TH, attesting to pervasive SNS innervation. When Siberian hamster inguinal WAT was sympathetically denervated via local injections of catecholaminergic toxin 6-hydroxydopamine (sparing putative parasympathetic nerves), subsequent PRV injection resulted in no central nervous system (CNS) or sympathetic chain infections suggesting no PSNS innervation. By contrast, vehicle-injected WAT subsequently inoculated with PRV had typical CNS/sympathetic chain viral infection patterns. Collectively, these data indicate no parasympathetic nerve markers in WAT of several species, with sparse DMV innervation and question the claim of PSNS WAT innervation as well as its functional significance.

Functional and histological evidence for the sympathetic nervous system (SNS) innervation of white adipose tissue (WAT) exists for several species; however, its sensory innervation has only been shown in laboratory rats, and its function is unclear. We tested the effects of sensory and SNS innervation of Siberian hamster epididymal and inguinal WAT (EWAT and IWAT) by assessing calcitonin gene-related peptide (CGRP)- and tyrosine hydroxylase-immunoreactivity (ir), respectively. Next, we tested the role of the sensory innervation of WAT on growth and cellularity because WAT surgical denervation increases pad mass via selective increases in fat cell number, an effect ascribed to SNS denervation but that could be due to the accompanying surgical disruption of WAT sensory innervation. Sensory denervation was accomplished via multiple local microinjections of capsaicin into WAT, and its effects were compared with those of surgical denervation. Surgically denervated IWAT and EWAT showed significantly decreased tyrosine hydroxylase-ir and CGRP-ir, whereas capsaicin-treated WAT had only significantly decreased CGRP-ir. Surgically denervated pad masses were significantly increased; this was accompanied by increased total fat cell number in IWAT, with no change in fat cell size. EWAT only showed a significant increase in the number of small- to medium-sized adipocytes (75-125 mum diameter). By contrast, sensory-denervated pad masses were unchanged, but IWAT showed significantly increased average fat cell size. Collectively, these data provide immunohistochemical evidence for sensory and SNS innervation of WAT in Siberian hamsters and differential control of WAT cellularity by these innervations, as well as the ability of locally applied capsaicin to selectively reduce WAT sensory innervation.