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Orexin/hypocretin neurons are located in and around the perifornical hypothalamus. Disinhibition of this area in the anaesthetized preparation evokes cardiorespiratory changes that can be reduced to nearly half or more by systemic Almorexant, a dual receptor antagonist of the two known orexin receptors, Ox1R and Ox2R. It is not clear if these reductions result from the blockade of one receptor or both. To determine the contribution of the two receptors, we compared the effects of Almorexant to those of the selective Ox1R antagonist ACT335827 and the selective Ox2R antagonists EMPA and TCS- OX2-29. Bicuculline (20 pmol) was injected in the perifornical hypothalamus of urethane-anaesthetized rats before and after administration of the drugs (all 15 mg/kg, intravenously). The pressor, tachycardic and tachypneic responses to bicuculline were attenuated/reduced by ACT335827 (by 19%, ns; 10%, ns and 24%, P < 0.01, respectively), EMPA (by 35% P < 0.01; 6%, ns; and 26% P < 0.05) and TCS- OX2-29 (by 13%, ns; 10%, ns and 42%, P < 0.001). These reductions represented only a fraction of the reduction after Almorexant (by 43%, P < 0.001; 42%, P < 0.001 and 65% P < 0.001). However, when the selective Ox1R and Ox2R antagonists were given in combination, the reductions were greater and closer to those of Almorexant ( ACT335827 + EMPA, by 26%, P < 0.05; 24%, P < 0.05 and 47%, P < 0.001; ACT335827 + TCS- OX2-29, by 40%, P < 0.01; 26%, P < 0.001 and 59%, P < 0.0001). This was particularly clear with the tachypneic response. These results suggest that both orexin receptors contribute to the cardiorespiratory response evoked from the hypothalamus under anaesthesia. They are consistent with our previous study in the conscious animal. [ABSTRACT FROM AUTHOR]

New Findings What is the central question of this study? Blockade of orexin receptors reduces blood pressure in spontaneously hypertensive rats (SHRs) but not in normotensive Wistar-Kyoto (WKY) rats, suggesting that upregulation of orexin signalling underlies the hypertensive phenotype of the SHR. However, it is not known what causes this upregulation., What is the main finding and its importance? Using orexin immunolabelling, we show that SHRs have 20% more orexin neurons than normotensive WKY and Wistar rats in the medial hypothalamus, which is a good match to their blood pressure phenotype. In contrast, there is no such match for the orexin neurons of the lateral hypothalamus. Essential hypertension may be linked to an increase in orexin neurons in the medial hypothalamus., The neuropeptide orexin contributes to the regulation of blood pressure as part of its role in the control of arousal during wakefulness and motivated behaviour (including responses to psychological stress). Recent work shows that pharmacological blockade of orexin receptors reduces blood pressure in spontaneously hypertensive rats (SHRs) but not in normotensive Wistar-Kyoto (WKY) rats. It is not clear why orexin signalling is upregulated in the SHR, but one possibility is that these animals have more orexin neurons than their normotensive WKY and Wistar relatives. To test this possibility, SHRs, WKY and Wistar male rats (6-16 weeks old) were killed, perfused and their brains sectioned and immunolabelled for orexin A. Labelled neurons were plotted and counted in the six best labelled hemisections (120 μm apart) of each brain. There were significantly more orexin neurons (+20%) in the medial hypothalamus (medial to fornix) of SHRs compared with WKY and Wistar rats (126 ± 4 versus 106 ± 5 and 104 ± 5 per hemisection, respectively, P < 0.05), which matches well the blood pressure phenotypes. In contrast, counts in the lateral hypothalamus did not match the blood pressure phenotypes (69 ± 2 versus 50 ± 3 and 76 ± 4, respectively). The results support the idea that orexin signalling is upregulated in the SHR and suggest that this is due, at least in part, to a greater number of orexin neurons in the medial hypothalamus. These medial orexin neurons, which are also involved in hyperarousal and stress responses, may contribute to the development of essential hypertension. [ABSTRACT FROM AUTHOR]

Hypocretin/orexin has a well-established role in wakefulness and in the maintenance of arousal. Because stress is associated with arousal, it has been proposed that hypocretin is also involved in stress. However, it is not clear if this is true for all forms of stress. To clarify this issue, we compared four conditions combining high arousal with no or low stress (wakefulness and exploration) or high stress (contextual fear and restraint) in the rat. We looked at Fos expression in hypocretin neurons, hypocretin-1 levels in cerebrospinal fluid and cardiovascular and behavioural changes after pharmacological blockade with the dual hypocretin receptor antagonist, almorexant. Fos expression in hypocretin neurons was highest with wakefulness and exploration, also high with fear but not significant with restraint. Hypocretin-1 levels were consistent with this pattern, although the differences were not as marked. Hypocretin receptor blockade with almorexant reduced the pressor, tachycardic and locomotor responses of wakefulness and exploration as well as the pressor and sympathetic component of the tachycardic response of fear. In contrast, almorexant did not reduce the pressor and tachycardic responses of restraint and nor did it reduce the pressor, tachycardic and locomotor responses of another stressor, i.e. cold exposure. Thus, hypocretin is not involved in all forms of stress. Comparison of the different conditions suggests that, regardless of stress, hypocretin involvement occurs when the arousal associated with the response includes increased attention to environmental cues. When it does, hypocretin will at least contribute to the cardiovascular response. The findings are of clinical relevance to some forms of psychological stress. [ABSTRACT FROM AUTHOR]

Conditioned fear to context, a pure form of psychological stress, is associated with sympathetically mediated changes including a marked hypertension. To identify the possible premotor sympathetic neurons mediating these changes, we conducted double-immunolabelling experiments combining fear-induced Fos with retrograde tracing from the thoracic cord (T2-L1). Presympathetic groups showing the greatest increase in the proportion of spinally projecting cells double-labelled with Fos compared with resting controls were the perifornical area (PeF; 22.7% vs. 0.4%) and paraventricular nucleus (Pa; 10.5% vs. 0.2%) in the hypothalamus, and the A5 noradrenergic group (33.6% vs. 0.2%) in the pons. In contrast, there was only a small increase in the presympathetic groups of the rostral ventral medulla, including the lateral paragigantocellular group (LPGi; 4.3% vs. 0.5%), raphe magnus and pallidus (1.1% vs. 0.6% and 1.8% vs. 0.5%), and the vasopressor group of the rostral ventrolateral medulla (RVLM; 1.9% vs. 0.8%). PeF, Pa, A5 and LPGi accounted for 21, 15, 16 and 6% of all the double-labelled cells, respectively, and RVLM for only 1%. Double-immunolabelling of Fos and tyrosine hydroxylase confirmed that many A5 neurons were activated (19%) and that practically no C1 neurons in RVLM were (1.3%). The results suggest that the main premotor sympathetic drive of the fear response comes from hypothalamic (PeF and Pa) and A5 neurons that project directly to the thoracic cord and bypass medullary presympathetic groups, and that the vasopressor premotor sympathetic neurons of the RVLM are unlikely to mediate the hypertensive pressure response of contextual fear. [ABSTRACT FROM AUTHOR]

1. The present study investigates the contribution of the sympathetic and vagal parasympathetic systems to the tachycardic response of long-lasting (40 min) conditioned fear responses to context. 2. The conditioned fear response evoked by re-exposure to a footshock chamber was tested 10 min after intravenous injection of the β-adrenoceptor antagonist propranolol (2 mg/kg) or the muscarinic antagonist atropine methyl nitrate (2 mg/kg) in rats implanted with radiotelemetric probes. 3. Compared with saline controls, the drugs did not change the behavioural component of the response (freezing, 22 kHz ultrasonic vocalizations) or its pressor component (+28 mmHg). 4. Propranolol abolished the tachycardic response of fear, whereas atropine more than doubled it (from +75 to +175 b.p.m. above resting baseline). 5. The results demonstrate that both sympathetic and vagal parasympathetic outflows to the heart are strongly activated during conditioned fear. The vagal activation may act to hold back cardiac acceleration while the animal waits for the aversive stimulus to come. [ABSTRACT FROM AUTHOR]

Infrared thermography was used to image changes in cutaneous temperature during a conditioned fear response to context. Changes in heart rate, arterial pressure, activity and body (i.p.) temperature were recorded at the same time by radio-telemetry, in addition to freezing immobility. A marked drop in tail and paws temperature (−5.3 and−7.5 °C, respectively, down to room temperature), which lasted for the entire duration of the response (30 min), was observed in fear-conditioned rats. In sham-conditioned rats, the drop was on average half the magnitude and duration. In contrast, temperature of the eye, head and back increased (between + 0.8 and + 1.5 °C), with no difference between the two groups of rats. There was a similar increase in body temperature although it was slightly higher and delayed in the fear-conditioned animals. Finally, ending of the fear response was associated with a gradual decrease in body temperature and a rebound increase in the temperature of the tail (+ 3.3 °C above baseline). This study shows that fear, and to some extent arousal, evokes a strong cutaneous vasoconstriction that is restricted to the tail and paws. This regionally specific reduction in blood flow may be part of a preparatory response to a possible fight and flight to reduce blood loss in the most exposed parts of the rat's body in case of injury. The data also show that the tail is the main part of the body used for dissipating internal heat accumulated during fear once the animal has returned to a safe environment. [ABSTRACT FROM AUTHOR]