The thermogenic activity of interscapular brown adipose tissue (IBAT) in response to physiologic stimuli, such as cold exposure, is controlled by its sympathetic innervation. To determine which brain regions might be involved in the regulation of cold-evoked increases in sympathetic outflow to IBAT, the present study compared central nervous system (CNS) areas activated by cold exposure with brain regions anatomically linked to the sympathetic innervation of IBAT. Immunocytochemical localization of Fos was examined in the brains of rats exposed to 4 degrees C for 4 hours. In a separate group of rats, the neural circuit involved in IBAT control, including the location of sympathetic preganglionic neurons in the spinal cord, was characterized with pseudorabies virus, a retrograde transynaptic tracer. Central noradrenergic and serotonergic groups related to the sympathetic outflow to IBAT also were identified. Localization of viral antigens at different survival times (66-96 hours) revealed infection in circumscribed CNS populations, but only a subset of the regions comprising this circuitry showed cold-evoked Fos expression. The raphe pallidus and the ventromedial parvicellular subdivision of the paraventricular hypothalamic nucleus (PVH), both infected at early survival times, were the main areas containing sympathetic premotor neurons activated by cold exposure. Major cold-sensitive areas projecting to spinal interneurons or to regions containing sympathetic premotor neurons, which became infected at intermediate intervals, included lateral hypothalamic, perifornical, and retrochiasmatic areas, anterior and posterior PVH, ventrolateral periaqueductal gray, and Barrington's nucleus. Areas infected later, most likely related to reception of cold-related signals, comprised the lateral preoptic area, parastrial nucleus, dorsomedial hypothalamic nucleus, lateral parabrachial nucleus, and nucleus of the solitary tract. These interconnected areas, identified by combining functional and retrograde anatomic approaches, likely constitute the central circuitry responsible for the increase in sympathetic outflow to IBAT during cold-evoked thermogenesis.

A good night's sleep is one of life's most satisfying experiences, while sleeplessness is stressful and causes cognitive impairment. Yet the mechanisms that regulate the ability to sleep have only recently been subjected to detailed investigation. New studies show that the control of wake and sleep emerges from the interaction of cell groups that cause arousal with other nuclei that induce sleep such as the ventrolateral preoptic nucleus (VLPO). The VLPO inhibits the ascending arousal regions and is in turn inhibited by them, thus forming a mutually inhibitory system resembling what electrical engineers call a "flip-flop switch." This switch may help produce sharp transitions between discrete behavioral states, but it is not necessarily stable. The orexin neurons in the lateral hypothalamus may help stabilize this system by exciting arousal regions during wakefulness, preventing unwanted transitions between wakefulness and sleep. The importance of this stabilizing role is apparent in narcolepsy, in which an absence of the orexin neurons causes numerous, unintended transitions in and out of sleep and allows fragments of REM sleep to intrude into wakefulness. These influences on the sleep/wake system by homeostatic and circadian drives, as well as emotional inputs, are reviewed. Understanding the pathways that underlie the regulation of sleep and wakefulness may provide important insights into how the cognitive and emotional systems interact with basic homeostatic and circadian drives for sleep.

Orexin (hypocretin) neurons, located exclusively in the PeF-LH, which includes the perifornical area (PeF), the lateral hypothalamus (LH) and lateral portions of the medial hypothalamus, have widespread projections and influence many physiological functions, including the autonomic regulation of body temperature and energy metabolism. Narcolepsy is characterized by the loss of orexin neurons and by disrupted sleep, but also by dysregulation of body temperature and by a strong tendency for obesity. Heat production (thermogenesis) in brown adipose tissue (BAT) contributes to the maintenance of body temperature and, through energy consumption, to body weight regulation. We identified a neural substrate for the influence of orexin neurons on BAT thermogenesis in rat. Nanoinjection of orexin-A (12 pmol) into the rostral raphe pallidus (rRPa), the site of BAT sympathetic premotor neurons, produced large, sustained increases in BAT sympathetic outflow and in BAT thermogenesis. Activation of neurons in the PeF-LH also enhanced BAT thermogenesis over a long time course. Combining viral retrograde tracing from BAT, or CTb tracing from rRPa, with orexin immunohistochemistry revealed synaptic connections to BAT from orexin neurons in PeF-LH and from rRPa neurons with closely apposed, varicose orexin fibers, as well as a direct, orexinergic projection from PeF-LH to rRPa. These results indicate a potent modulation of BAT thermogenesis by orexin released from the terminals of orexin neurons in PeFLH directly into the rRPa and provide a potential mechanism contributing to the disrupted regulation of body temperature and energy metabolism in the absence of orexin.

Sleep architecture is often disturbed after a stressful event; nevertheless, little is known about the brain circuitry responsible for the sleep perturbations induced by stress. We exposed rats to a psychological stressor (cage exchange) that initially causes an acute stress response, but several hours later generates a pattern of sleep disturbances similar to that observed in stress-induced insomnia in humans: increased sleep latency, decreased non-REM (nREM) and REM sleep, increased fragmentation, and high-frequency EEG activity during nREM sleep. We examined the pattern of Fos expression to identify the brain circuitry activated, and found increased Fos in the cerebral cortex, limbic system, and parts of the arousal and autonomic systems. Surprisingly, there was simultaneous activation of the sleep-promoting areas, most likely driven by ongoing circadian and homeostatic pressure. The activity in the cerebral cortex and arousal system while sleeping generates a novel intermediate state characterized by EEG high-frequency activity, distinctive of waking, during nREM sleep. Inactivation of discrete limbic and arousal regions allowed the recovery of specific sleep components and altered the Fos pattern, suggesting a hierarchical organization of limbic areas that in turn activate the arousal system and subsequently the cerebral cortex, generating the high-frequency activity. This high-frequency activity during nREM was eliminated in the stressed rats after inactivating parts of the arousal system. These results suggest that shutting down the residual activity of the limbic-arousal system might be a better approach to treat stress-induced insomnia, rather than potentiation of the sleep system, which remains fully active.

Sympathetic premotor neurons in the rostral medullary raphe (RMR) regulate heat conservation by tail artery vasoconstriction and brown adipose tissue thermogenesis. These neurons are a critical relay in the pathway that increases body temperature. However, the origins of the inputs that activate the RMR during cold exposure have not been definitively identified. We investigated the afferents to the RMR that were activated during cold by examining Fos expression in retrogradely labeled neurons after injection of cholera toxin B subunit (CTb) in the RMR. These experiments identified a cluster of Fos-positive neurons in the dorsomedial hypothalamic nucleus and dorsal hypothalamic area (DMH/DHA) with projections to the RMR that may mediate cold-induced elevation of body temperature. Also, neurons in the median preoptic nucleus (MnPO) and dorsolateral preoptic area (DLPO) and in the A7 noradrenergic cell group were retrogradely labeled but lacked Fos expression, suggesting that they may inhibit the RMR. To investigate whether individual or common preoptic neurons project to the RMR and DMH/DHA, we injected CTb into the RMR and Fluorogold into the DMH/DHA. We found that projections from the DLPO and MnPO to the RMR and DMH/DHA emerge from largely separate neuronal populations, indicating they may be differentially regulated. Combined cell-specific lesions of MnPO and DLPO, but not lesions of either one alone, caused baseline hyperthermia. Our data suggest that the MnPO and DLPO provide parallel inhibitory pathways that tonically inhibit the DMH/DHA and the RMR at baseline, and that hyperthermia requires the release of this inhibition from both nuclei.