Orexin-Corticotropin-Releasing Factor Receptor Heteromers in the Ventral Tegmental Area as Targets for Cocaine

Release of the neuropeptides corticotropin-releasing factor (CRF) and orexin-A in the ventral tegmental area (VTA) play an important role in stress-induced cocaine-seeking behavior. We provide evidence for pharmacologically significant interactions between CRF and orexin-A that depend on oligomerization of CRF1 receptor (CRF1R) and orexin OX1 receptors (OX1R). CRF1R–OX1R heteromers are the conduits of a negative crosstalk between orexin-A and CRF as demonstrated in transfected cells and rat VTA, in which they significantly modulate dendritic dopamine release. The cocaine target σ1 receptor (σ1R) also associates with the CRF1R–OX1R heteromer. Cocaine binding to the σ1R–CRF1R–OX1R complex promotes a long-term disruption of the orexin-A–CRF negative crosstalk. Through this mechanism, cocaine sensitizes VTA cells to the excitatory effects of both CRF and orexin-A, thus providing a mechanism by which stress induces cocaine seeking

Orexinergic Input to Dopaminergic Neurons of the Human Ventral Tegmental Area

The mesolimbic reward pathway arising from dopaminergic (DA) neurons of the ventral tegmental area (VTA) has been strongly implicated in reward processing and drug abuse. In rodents, behaviors associated with this projection are profoundly influenced by an orexinergic input from the lateral hypothalamus to the VTA. Because the existence and significance of an analogous orexigenic regulatory mechanism acting in the human VTA have been elusive, here we addressed the possibility that orexinergic neurons provide direct input to DA neurons of the human VTA. Dual-label immunohistochemistry was used and orexinergic projections to the VTA and to DA neurons of the neighboring substantia nigra (SN) were analyzed comparatively in adult male humans and rats. Orexin B-immunoreactive (IR) axons apposed to tyrosine hydroxylase (TH)-IR DA and to non-DA neurons were scarce in the VTA and SN of both species. In the VTA, 15.062.8% of TH-IR perikarya in humans and 3.260.3% in rats received orexin B-IR afferent contacts. On average, 0.2460.05 and 0.0560.005 orexinergic appositions per TH-IR perikaryon were detected in humans and rats, respectively. The majority(86–88%) of randomly encountered orexinergic contacts targeted the dendritic compartment of DA neurons. Finally, DA neurons of the SN also received orexinergic innervation in both species. Based on the observation of five times heavierorexinergic input to TH-IR neurons of the human, compared with the rat, VTA, we propose that orexinergic mechanism acting in the VTA may play just as important roles in reward processing and drug abuse in humans, as already established well in rodents

Acute High Fat Diet Consumption Activates the Mesolimbic Circuit and Requires Orexin Signaling in a Mouse Model

Overconsumption of palatable energy-dense foods has negative health implications and it is associated with obesity andseveral eating disorders. Currently, little is known about the neuronal circuitries activated by the acute ingestion of arewarding stimulus. Here, we used a combination of immunohistochemistry, pharmacology and neuronal tracing analysesto examine the role of the mesolimbic system in general, and the orexin neurons in particular, in a simple experimental testin which naı ̈ve mice are allowed to spontaneously eat a pellet of a high fat diet (HFD) for 2 h. We found that acute HFDactivates c-Fos expression in several reward-related brain areas, including the ventral tegmental area (VTA), nucleusaccumbens, central amygdala and lateral hypothalamic area. We also found that: i- HFD-mediated orosensory stimulationwas required for the mesolimbic pathway activation, ii- acute HFD differentially activates dopamine neurons of theparanigral, parabrachial pigmented and interfascicular sub-regions of the VTA, and iii- orexin neurons of the lateralhypothalamic area are responsive to acute HFD. Moreover, orexin signaling blockade, with the orexin 1 receptor antagonistSB-334867, reduces acute HFD consumption and c-Fos induction in the VTA but not in the other mesolimbic nuclei understudy. Finally, we found that most orexin neurons responsive to acute HFD innervate the VTA. Our results show that acuteHFD consumption recruits the mesolimbic system and that the full manifestation of this eating behavior requires theactivation of orexin signaling.Fil: Valdivia Torres, Lesly Spring. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; ArgentinaFil: Patrone, Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; ArgentinaFil: Reynaldo, Mirta Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; ArgentinaFil: Perello, Mario. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; Argentin

Hypocretin-1 receptors regulate the reinforcing and reward-enhancing effects of cocaine: pharmacological and behavioral genetics evidence.

Considerable evidence suggests that transmission at hypocretin-1 (orexin-1) receptors (Hcrt-R1) plays an important role in the reinstatement of extinguished cocaine-seeking behaviors in rodents. However, far less is known about the role for hypocretin transmission in regulating ongoing cocaine-taking behavior. Here, we investigated the effects of the selective Hcrt-R1 antagonist SB-334867 on cocaine intake, as measured by intravenous (IV) cocaine self-administration in rats. The stimulatory effects of cocaine on brain reward systems contribute to the establishment and maintenance of cocaine-taking behaviors. Therefore, we also assessed the effects of SB-334867 on the reward-enhancing properties of cocaine, as measured by cocaine-induced lowering of intracranial self-stimulation (ICSS) thresholds. Finally, to definitively establish a role for Hcrt-R1 in regulating cocaine intake, we assessed IV cocaine self-administration in Hcrt-R1 knockout mice. We found that SB-334867 (1-4 mg/kg) dose-dependently decreased cocaine (0.5 mg/kg/infusion) self-administration in rats but did not alter responding for food rewards under the same schedule of reinforcement. This suggests that SB-334867 decreased cocaine reinforcement without negatively impacting operant performance. SB-334867 (1-4 mg/kg) also dose-dependently attenuated the stimulatory effects of cocaine (10 mg/kg) on brain reward systems, as measured by reversal of cocaine-induced lowering of ICSS thresholds in rats. Finally, we found that Hcrt-R1 knockout mice self-administered far less cocaine than wildtype mice across the entire dose-response function. These data demonstrate that Hcrt-R1 play an important role in regulating the reinforcing and reward-enhancing properties of cocaine and suggest that hypocretin transmission is likely essential for establishing and maintaining the cocaine habit in human addicts

Single Gene Deletions of Orexin, Leptin, Neuropeptide Y, and Ghrelin Do Not Appreciably Alter Food Anticipatory Activity in Mice

Timing activity to match resource availability is a widely conserved ability in nature. Scheduled feeding of a limited amount of food induces increased activity prior to feeding time in animals as diverse as fish and rodents. Typically, food anticipatory activity (FAA) involves temporally restricting unlimited food access (RF) to several hours in the middle of the light cycle, which is a time of day when rodents are not normally active. We compared this model to calorie restriction (CR), giving the mice 60% of their normal daily calorie intake at the same time each day. Measurement of body temperature and home cage behaviors suggests that the RF and CR models are very similar but CR has the advantage of a clearly defined food intake and more stable mean body temperature. Using the CR model, we then attempted to verify the published result that orexin deletion diminishes food anticipatory activity (FAA) but observed little to no diminution in the response to CR and, surprisingly, that orexin KO mice are refractory to body weight loss on a CR diet. Next we tested the orexigenic neuropeptide Y (NPY) and ghrelin and the anorexigenic hormone, leptin, using mouse mutants. NPY deletion did not alter the behavior or physiological response to CR. Leptin deletion impaired FAA in terms of some activity measures, such as walking and rearing, but did not substantially diminish hanging behavior preceding feeding time, suggesting that leptin knockout mice do anticipate daily meal time but do not manifest the full spectrum of activities that typify FAA. Ghrelin knockout mice do not have impaired FAA on a CR diet. Collectively, these results suggest that the individual hormones and neuropepetides tested do not regulate FAA by acting individually but this does not rule out the possibility of their concerted action in mediating FAA

Selective loss of GABAB receptors in orexin/hypocretin-producing neurons results in disrupted sleep/wakefulness architecture

We generated mice with a selective loss of GABAB receptors in orexin neurons. Orexin neurons in these GABAB1<sup>-/-(orexin)</sup> mice showed reduced responsiveness to GABA<sub>A</sub> receptor agonists due to a compensatory increase in GABAA receptor-mediated inhibition. This increased GABA<sub>A</sub> receptor-mediated inhibition of orexin neurons is due to orexin-1 receptor-mediated activation of local GABAergic interneurons. Surprisingly, orexin neurons were also less responsive to glutamate, apparently because the augmented GABA<sub>A</sub> receptor-mediated inhibition increases the membrane conductance and shunts excitatory currents. These observations indicate that absence of GABA<sub>B</sub> receptors decreases the sensitivity of orexin neurons to both excitatory and inhibitory inputs. GABAB1<sup>-/-(orexin)</sup>mice exhibited severe fragmentation of sleep/wake states during both the light and dark periods without affecting total sleep time or inducing cataplexy, indicating that GABA<sub>B</sub> receptors are crucial regulators of orexin neurons and that "fine tuning" of orexin neurons by inhibitory and excitatory inputs is important for the stability of sleep/waking states

Animals-assisted therapy: A brief review

In rehabilitative setting, the presence of animals can be considered as an important stimulus for verbal and social communication, and for mood regulation. Interaction with an animal is beneficial for children's development and numerous psychological tests have revealed that growing up with pets has a beneficial effect on children's self-esteem and self-confidence, can improve empathy, a sense of responsibility and cognitive development, as well as social status within the peer group

DEC2 modulates orexin expression and regulates sleep.

Adequate sleep is essential for physical and mental health. We previously identified a missense mutation in the human DEC2 gene (BHLHE41) leading to the familial natural short sleep behavioral trait. DEC2 is a transcription factor regulating the circadian clock in mammals, although its role in sleep regulation has been unclear. Here we report that prepro-orexin, also known as hypocretin (Hcrt), gene expression is increased in the mouse model expressing the mutant hDEC2 transgene (hDEC2-P384R). Prepro-orexin encodes a precursor protein of a neuropeptide producing orexin A and B (hcrt1 and hcrt2), which is enriched in the hypothalamus and regulates maintenance of arousal. In cell culture, DEC2 suppressed prepro-orexin promoter-luc (ore-luc) expression through cis-acting E-box elements. The mutant DEC2 has less repressor activity than WT-DEC2, resulting in increased orexin expression. DEC2-binding affinity for the prepro-orexin gene promoter is decreased by the P384R mutation, likely due to weakened interaction with other transcription factors. In vivo, the decreased immobility time of the mutant transgenic mice is attenuated by an orexin receptor antagonist. Our results suggested that DEC2 regulates sleep/wake duration, at least in part, by modulating the neuropeptide hormone orexin

Genes and primary headaches: discovering new potential therapeutic targets

Genetic studies have clearly shown that primary headaches (migraine, tension-type headache and cluster headache) are multifactorial disorders characterized by a complex interaction between different genes and environmental factors. Genetic association studies have highlighted a potential role in the etiopathogenesis of these disorders for several genes related to vascular, neuronal and neuroendocrine functions. A potential role as a therapeutic target is now emerging for some of these genes. The main purpose of this review is to describe new advances in our knowledge regarding the role of MTHFR, KCNK18, TRPV1, TRPV3 and HCRTR genes in primary headache disorders. Involvement of these genes in primary headaches, as well as their potential role in the therapy of these disorders, will be discussed

ATP-Sensitive Potassium Channel-Mediated Lactate Effect on Orexin Neurons: Implications for Brain Energetics during Arousal

Active neurons have a high demand for energy substrate, which is thought to be mainly supplied as lactate by astrocytes. Heavy lactate dependence of neuronal activity suggests that there may be a mechanism that detects and controls lactate levels and/or gates brain activation accordingly. Here, we demonstrate that orexin neurons can behave as such lactate sensors. Using acute brain slice preparations and patch-clamp techniques, we show that the monocarboxylate transporter blocker α-cyano-4-hydroxycinnamate (4-CIN) inhibits the spontaneous activity of orexin neurons despite the presence of extracellular glucose. Furthermore, fluoroacetate, a glial toxin, inhibits orexin neurons in the presence of glucose but not lactate. Thus, orexin neurons specifically use astrocyte-derived lactate. The effect of lactate on firing activity is concentration dependent, an essential characteristic of lactate sensors. Furthermore, lactate disinhibits and sensitizes these neurons for subsequent excitation. 4-CIN has no effect on the activity of some arcuate neurons, indicating that lactate dependency is not universal. Orexin neurons show an indirect concentration-dependent sensitivity to glucose below 1mM, responding by hyperpolarization, which is mediated by ATP-sensitive potassium channels composed of Kir6.1 and SUR1 subunits. In conclusion, our study suggests that lactate is a critical energy substrate and a regulator of the orexin system. Together with the known effects of orexins in inducing arousal, food intake, and hepatic glucose production, as well as lactate release from astrocytes in response to neuronal activity, our study suggests that orexin neurons play an integral part in balancing brain activity and energy supply