Brain Interstitial Nociceptin/Orphanin FQ Levels are Elevated in Parkinson's Disease

Expression and release of nociceptin/orphanin FQ (N/OFQ) are elevated in the substantia nigra reticulata of 6‐hydroxydopamine‐hemilesioned rats, suggesting a pathogenic role for N/OFQ in Parkinson's disease. In this study, we investigated whether elevation of N/OFQ expression in 6‐hydroxydopamine‐hemilesioned rats selectively occurs in substantia nigra and whether hypomotility following acute haloperidol administration is accompanied by a rise in nigral N/OFQ levels. Moreover, to prove a link between N/OFQ and idiopathic Parkinson's disease in humans, we measured N/OFQ levels in the cerebrospinal fluid of parkinsonian patients undergoing surgery for deep brain stimulation. In situ hybridization demonstrated that dopamine depletion was associated with increase of N/OFQ expression in substantia nigra (compacta +160%, reticulata +105%) and subthalamic nucleus (+45%), as well as reduction in caudate putamen (?20%). No change was observed in globus pallidus, nucleus accumbens, thalamus, and motor cortex. Microdialysis coupled to the bar test allowed to demonstrate that acute administration of haloperidol (0.8 and 3 mg/kg) increased nigral N/OFQ levels (maximally of +47% and +53%, respectively) in parallel with akinesia. A correlation with preclinical studies was found by analyzing N/OFQ levels in humans. Indeed, N/OFQ levels were found to be ～3.5‐fold elevated in the cerebrospinal fluid of parkinsonian patients (148 fmol/ml) compared with nonparkinsonian neurologic controls (41 fmol/ml). These data represent the first clinical evidence linking N/OFQ to idiopathic Parkinson's disease in humans. They strengthen the pathogenic role of N/OFQ in the modulation of parkinsonism across species and provide a rationale for developing N/OFQ receptor antagonists as antiparkinsonian drugs. © 2010 Movement Disorder Society     

Pro-nociceptin/orphanin FQ and NOP receptor mRNA levels in the forebrain of food deprived rats

Forebrain injections of nociceptin/orphanin FQ (N/OFQ), the endogenous ligand of the NOP opioid receptor, previously referred to as ORL1 or OP4 receptor, stimulate feeding in freely feeding rats, while the NOP receptor antagonist [Nphe(1)]N/OFQ(1-13)NH(2) inhibits food deprivation-induced feeding. To further evaluate whether the N/OFQ-NOP receptor system plays a physiological role in feeding control, the present study evaluated forebrain mRNA levels for the N/OFQ precursor (pro-N/OFQ), as well as for the NOP receptor in food deprived rats. The results obtained show that food deprived rats have lower mRNA levels for the NOP receptor in several forebrain regions; a significant reduction was found in the paraventricular and lateral hypothalamic nuclei and in the central nucleus of the amygdala. Food deprived rats also exhibited lower pro-N/OFQ mRNA levels in the central amygdala. These results suggest that the N/OFQ-NOP receptor system may have a physiological role in feeding control. The observation that food deprivation reduces gene expression of the N/OFQ-NOP receptor system is apparently not consistent with a direct hyperphagic action for N/OFQ. Taking into account that N/OFQ exerts inhibitory actions at cellular level, the present results may be in keeping with the hypothesis that N/OFQ stimulates feeding by inhibiting neurons inhibitory for food intake; under conditions of food deprivation, these neurons may be silent and the N/OFQ-NOP receptor system, which controls them, may also be regulated at a lower level. Consistently, in the present study N/OFQ stimulated food intake in freely feeding rats, but did not further increase feeding in food deprived rats.     

Nociceptin/orphanin FQ-induced delay in gastric emptying: role of central corticotropin-releasing factor and glucocorticoid receptors

Abstract  When injected intracerebroventricularly (i.c.v.) in rats, nociceptin/orphanin FQ (N/OFQ) delays gastric emptying and increases plasma corticosterone levels. Our aim in this study was to investigate changes in gastric emptying of a phenol red meal, and the plasma corticosterone response to N/OFQ in adrenalectomized (ADX) rats, in ADX rats injected with corticosterone at 1, 24 and 72 h before the gastric emptying assay, and in intact rats i.c.v. pretreated with a glucocorticoid antagonist (RU486) and with a corticotropin-releasing factor receptor antagonist ( α -helical CRF_9–41). In adrenal intact rats, i.c.v. injection of N/OFQ (2.5 nmol rat⁻¹) significantly delayed gastric emptying (by 70%) and increased plasma corticosterone concentrations. Conversely, in ADX rats, N/OFQ left gastric emptying unchanged. In ADX rats, corticosterone injected at 1, 24 and 72 h before the gastric emptying assay almost restored the N/OFQ-induced delay in gastric emptying. Finally, pretreatment with RU486- and α -helical CRF_9–41 abolished the N/OFQ-induced inhibition of gastric emptying. These findings suggest that central N/OFQ inhibits gastric emptying through an integrated orphaninergic system–CRF interaction in which corticosterone plays a permissive role.     

Dissociation Between Behavioral and Hormonal Responses to the Forced Swim Stress in Lactating Rats

Retention of immobility in the Porsolt forced swim test is believed to be dependent upon glucocorticoid secretion in male rats. Because lactating females exhibit increased basal glucocorticoid secretion and blunted stress responses, we tested the hypothesis that lactation-induced changes in adrenal glucocorticoid and in circulating estrogen and progesterone levels would improve retention and/or acquisition of immobility. Immobility was recorded during 3 intervals of 5 min on day 1 (acquisition) and one 5 min interval 24 h later (retention). Blood samples were collected before the swim test and at various times after the onset of stress for plasma ACTH and corticosterone (B) determinations. Male rats (young = 200 g, old = 325 g) were compared to virgin females (V) and to lactating females in early (day 8-10, EL) and late (day 17–19, LL) lactation. Adrenalectomy (ADX) and ovariectomy (OVX) were performed 5 and 10 days prior to testing, respectively. All animals acquired immobility at the end of the 15 min swim on day 1, but only the young male group exhibited a significant retention of immobility on day 2. Total immobility was higher in males than females (V) although basal and stress-induced ACTH and B secretion were comparable on both testing days. Lactational status did not affect immobility in either the acquisition or retention phases. However, stress-induced ACTH secretion was greatly diminished in intact and ADX lactating females (EL and LL) compared to virgins (LL < EL < virgin), demonstrating a clear dissociation between behavioral and neuroendocrine responses. Following ADX, immobility in the retention phase was either decreased in males or increased in lactating females. Finally, OVX decreased immobility in both lactating (EL) and virgin females without significantly altering the magnitude of the ACTH and B responses to stress. In summary, our results demonstrated both sex-related and lactation-related differences in the behavioral and endocrine responses to the forced swim test of Porsolt. Although retention of the immobile response is thought to involve glucocorticoids and/or opioids secreted during the first testing session, we did not find evidence for a direct relationship between basal or stress-induced total corticosterone secretion, the magnitude of ACTH response to stress and behavioral scores in the retention period. However, experimental variables such as body weight, sex and water depth could significantly modify the outcome of behavioral testing and question the validity of glucocorticoid-mediated retention processes. Since the effect of ADX was reversed in lactating females compared to male rats, we hypothesize that glucocorticoid sensitivity of cognitive processes controlling behavioral reactivity is different from that controlling hypothalamic-adrenocortical function. Our results also demonstrated a clear dissociation between behavioral and neuroendocrine responses to the swim test, in particular during lactation. In early and late lactation, blunted responsiveness to stress was not caused by enhanced glucocorticoid feedback but might result from modifications in the inhibitiory and/or stimulatory inputs to hypothalamic neurons controlling adrenocortical activity.     

Orphanin FQ/nociceptin binds to functionally coupled ORL1 receptors on human immune cell lines and alters peripheral blood mononuclear cell proliferation

Orphanin FQ/nociceptin (OFQ/N) has been shown to modulate nociception, responses to stress and anxiety. We investigated OFQ/N function in human immune cells. We find that monocytic U937, T lymphocytic CEM, and MOLT-4 cell lines express OFQ/N binding sites at levels comparable to that of human SH-SY5Y neuroblastoma cells. We show that OFQ/N receptors are functionally coupled to G proteins in these cells. Finally OFQ/N decreases proliferation of phytohemagglutinin-stimulated peripheral blood mononuclear cells in vitro at doses ranging from 10(-13) to 10(-8) M. Thus, our data suggest that OFQ/N and OFQ/N receptor may act as an immunomodulatory system.     

Effects of Chronic Oestradiol,Progesterone and Medroxyprogesterone Acetate on Hippocampal Neurogenesis and Adrenal Mass in Adult Female Rats

Both natural oestrogens and progesterone influence synaptic plasticity and neurogenesis within the female hippocampus. However, less is known of the impact of synthetic hormones on hippocampal structure and function. There is some evidence that the administration of the synthetic progestin, medroxyprogesterone acetate (MPA) is not as beneficial as natural progesterone and can attenuate oestrogen‐induced neuroprotection. Although the effects of oestradiol have been well studied, little is known about the effects of natural and synthetic progestins alone and in combination with oestradiol on adult neurogenesis in females. In the present study, we investigated the effects of chronic oestradiol, progesterone, MPA and the co‐administration of each progestin with oestradiol on neurogenesis within the dentate gyrus of adult ovariectomised female rats. Twenty‐four hours after a bromodeoxyuridine (BrdU; 200 mg/kg) injection, female rats were repeatedly administered either progesterone (1 or 4 mg), MPA (1 or 4 mg), oestradiol benzoate (EB), progesterone or MPA in combination with EB (10 μg), or vehicle for 21 days. Rats were perfused on day 22 and brain tissue was analysed for the number of BrdU‐labelled and Ki67 (an endogenous marker of cell proliferation)‐expressing cells. EB alone and MPA + EB significantly decreased neurogenesis and the number of surviving BrdU‐labelled cells in the dorsal region of the dentate gyrus, independent of any effects on cell proliferation. Furthermore, MPA (1 and 4 mg) and MPA + EB treated animals had significantly lower adrenal/body mass ratios and reduced serum corticosterone (CORT) levels. By contrast, progesterone + EB treated animals had significantly higher adrenal/body mass ratios and 1 mg of progesterone, progesterone + EB, and EB significantly increased CORT levels. The results of the present study demonstrate that different progestins alone and in combination with oestradiol can differentially affect neurogenesis (via cell survival) and regulation of the hypothalamic‐pituitary‐adrenal axis. These findings have implications for women using hormone replacement therapies with MPA for both neuroprotection and stress‐related disorders.     

Evidence that Cholecystokinin Receptors are not Involved in the Hypothalamic-Pituitary-Adrenal Response to Intraperitoneal Administration of Interleukin-1β

The purpose of this study was to investigate a putative role for cholecystokinin (CCK) in the activation of the hypothalamic-pituitary-adrenal (HPA) axis following intraperitoneal (i.p.) administration of interleukin-1-beta (IL-1beta). Previous studies predict that CCKA receptors on vagal sensory afferents may be involved in the initiation of the stress response following an acute i.p. injection of IL-1beta. Adult male rats were given an i.p. injection of a specific CCKA (devazepide, 1 mg/kg) or CCKB (CI-988, 1 mg/kg) receptor antagonist, 30 min prior to an i.p. injection of rat recombinant IL-1beta (rrIL-1beta), 0.5 microg/kg in 0.9% sterile saline/0.01% rat serum albumin. Blood samples were obtained via an indwelling jugular vein catheter, and the plasma levels of the stress hormones ACTH (adrenocorticotropin hormone) and corticosterone analysed over time as an indicator of HPA axis activation. This dose of rrIL-1beta resulted in a significant release of ACTH and corticosterone, peaking at 30-60 min, and returning to basal levels by 2 h. Pretreatment with either devazepide or CI-988 had no effect on the rrIL-1beta induced ACTH or corticosterone release. In contrast, the same dose of devazepide completely inhibited the ACTH and corticosterone response to i.p. CCK (octapeptide, sulphated form, CCK-8S), 5 microg/kg. It is concluded that CCK receptors are not involved in the hormonal stress response to a submaximal i.p. dose of rrIL-1beta.     

The Effect of Long-Term Castration on the Neuronal and Physiological Responses to Acute or Repeated Restraint Stress: Interactions with Opioids and Prostaglandins

Sixty min supine restraint stress induced a marked, but transient, hypothermic response in intact male rats, which tended to recover towards pre-stress levels or slightly overshoot during the later stages of restraint. Castration reduced the initial hypothermia but increased overshoots. Baseline (pre-stress) core temperature was also higher in castrated than intact rats, but the reduction in stress-induced hypothermia was still present even when this difference had been taken into account. The hypothermic response was not altered during the course of 10 sessions of daily repeated restraint in either intact or castrated rats. Castration did not alter cardiac responses to restraint. Both intact and castrated rats showed marked tachycardia during the initial 12 min of restraint, followed by a gradual fall towards baseline values. Repeated restraint accentuated the second phase of the cardiac response, without modifying the initial tachycardia, in both intact and castrated animals. The response of blood corticosterone levels to the first period of restraint was unaltered by castration but the attenuation observed after 10 sessions of stress was more complete in castrated rats. The neuronal c-fos response 60 min after the last of the series of repeated restraints was less in the hypothalamic paraventricular nucleus, medial amygdala, and locus coeruleus compared with that following the first session, but not in the lateral septum or the bed nucleus of the stria terminalis. Castration did not change the c-fos profile following the same number of restraint sessions. Castration depleted completely the vasopressinergic innervation in the lateral septum, diagonal band of Broca and medial amygdala. Five mg/kg naloxone i.p. prior to the onset of stress resulted in persistent hypothermic responses throughout the stress period in intact males. This was reduced in castrated rats. Blockade of prostaglandins synthesis by 12 mg/kg indornethacin i.p. 3 h prior to stress had little effect on hypothermic stress responses. The results of these experiments suggest that an androgen-dependent process, possibly AVP-containing systems in the forebrain, may regulate both baseline and stress-induced levels of core temperature and have selective effects on components of the response to stress, including restraint-induced hypothermia and the adaptation of corticoid responses to repeated restraint. Steroid-dependent opioid systems may also be implicated in the hypotherrnic response to restraint stress.     

UFP‐112 a Potent and Long‐Lasting Agonist Selective for the Nociceptin/Orphanin FQ Receptor

Nociceptin/orphanin FQ (N/OFQ) controls several biological functions via selective activation of the N/OFQ peptide receptor (NOP). [(pF)Phe⁴Aib⁷Arg¹⁴Lys¹⁵]N/OFQ‐NH₂ (UFP‐112) is an NOP receptor ligand designed using a combination of several chemical modifications in the same peptide sequence that increase NOP receptor affinity/potency and/or reduce susceptibility to enzymatic degradation. In the present review article, we summarize data from the literature and present original findings on the in vitro and in vivo pharmacological features of UFP‐112. Moreover, important biological actions and possible therapeutic indications of NOP receptor agonists are discussed based on the results obtained with UFP‐112 and compared with other peptide and nonpeptide NOP receptor ligands.     

Paradoxical Effects of Chronic Morphine Treatment on the Temperature and Pituitary-Adrenal Responses to Acute Restraint Stress: A Chronic Stress Paradigm

Body temperature and pituitary-adrenal responses to restraint (15 min or 4 h) stress were evaluated in nondependent and morphine-dependent rats. Male Sprague-Dawley rats were treated twice daily with increasing doses of morphine (10-100 mg/kg, s.c.) for 16 days. Transmitters were implanted in the peritoneal cavity to monitor body temperature and blood was collected for hormone assays. Acute withdrawal from chronic morphine treatment was associated with reduced body weight, increased adrenal weight and decreased thymus weight. Sixteen days after termination of chronic morphine treatment, rats had recovered normal adrenal size, but still displayed marked thymus involution and reduced body weight. Restraint-induced hyperthermia was attenuated in morphine-dependent rats that had undergone 12-h withdrawal. Sixteen days after withdrawal, rats still had not fully recovered the hyperthermic response to restraint. Chronic morphine treatment resulted in a marked elevation of basal corticosterone concentrations. Despite the negative-feedback effects of elevated basal corticosterone concentrations, morphine-dependent rats that had undergone 12-h withdrawal displayed a potentiated and prolonged corticosterone response to restraint stress. In contrast, rats that had undergone 8-day and 16-day morphine withdrawal had recovered normal basal pituitary-adrenal activity, but displayed significantly reduced and shorter adrenocorticotropic hormone and corticosterone responses to restraint. These results suggest that chronic morphine dependence is a chronic stressor, resulting in profound and long-lasting changes in the temperature and pituitary-adrenal responses to acute restraint stress in a time-dependent manner. This morphine-dependence model may be useful in understanding the role that hormonal stress responses play in the maintenance and relapse to opioid use in humans.     

Acute and Long-Term Treatments with the Selective Serotonin Reuptake Inhibitor Citalopram Modulate the HPA Axis Activity at Different Levels in Male Rats

It is well established that the maximal therapeutic effect of selective serotonin reuptake inhibitors (SSRI) are achieved in depressive patients after several weeks of treatment, but the adaptive processes leading to the therapeutic effects are unclear. It has been shown that hyperactivity in the hypothalamic-pituitary-adrenal (HPA) axis in depressive patients is affected by long-term antidepressant treatment. These changes occur in association with the mood normalising effect, suggesting that antidepressants affect the HPA axis and this effect is associated with the therapeutic effect. Male Wistar rats were treated with the SSRI, citalopram, to investigate time-related changes in components that may be involved in the desensitization of the HPA axis. A single injection of citalopram (10 mg/kg, s.c.), increased the plasma levels of ACTH and corticosterone in a dose-dependent manner and increased the number of c-Fos containing cells in the hypothalamic paraventricular nucleus. A daily treatment with the same compound (10 mg/kg, s.c.) for 14 days decreased the expression of POMC mRNA ( approximately 40%). In addition, a blunted response to citalopram was observed in animals long-term treated with citalopram. Also CRF-stimulated cAMP accumulation in the pituitary was altered. In conclusion, acute citalopram activated the HPA-axis at the hypothalamic level and long-term citalopram treatment desensitized the HPA-axis at the pituitary level. These results support the hypothesis that the therapeutic effects of long-term antidepressant treatments reduce HPA axis responsiveness.     

Vasopressin and the Output of the Hypothalamic Biological Clock

The physiological effects of vasopressin as a peripheral hormone were first reported more than 100 years ago. However, it was not until the first immunocytochemical studies were carried out in the early 1970s, using vasopressin antibodies, and the discovery of an extensive distribution of vasopressin‐containing fibres outside the hypothalamus, that a neurotransmitter role for vasopressin could be hypothesised. These studies revealed four additional vasopressin systems next to the classical magnocellular vasopressin system in the paraventricular and supraoptic nuclei: a sexually dimorphic system originating from the bed nucleus of the stria terminalis and the medial amygdala, an autonomic and endocrine system originating from the medial part of the paraventricular nucleus, and the circadian system originating from the hypothalamic suprachiasmatic nuclei (SCN). At about the same time as the discovery of the neurotransmitter function of vasopressin, it also became clear that the SCN contain the main component of the mammalian biological clock system (i.e. the endogenous pacemaker). This review will concentrate on the significance of the vasopressin neurones in the SCN for the functional output of the biological clock that is contained within it. The vasopressin‐containing subpopulation is a characteristic feature of the SCN in many species, including humans. The activity of the vasopressin neurones in the SCN shows a pronounced daily variation in its activity that has also been demonstrated in human post‐mortem brains. Animal experiments show an important role for SCN‐derived vasopressin in the control of neuroendocrine day/night rhythms such as that of the hypothalamic‐pituitary‐adrenal and hypothalamic‐pituitary‐gonadal axes. The remarkable correlation between a diminished presence of vasopressin in the SCN and a deterioration of sleep‐wake rhythms during ageing and depression make it likely that, also in humans, the vasopressin neurones contribute considerably to the rhythmic output of the SCN.     

Increased Expression of Type 1 Angiotensin II Receptors in the Hypothalamic Paraventricular Nucleus following Stress and Glucocorticoid Administration

Double staining in situ hybridization studies have shown that angiotensin II (All) type 1 receptors (AT1) in the hypothalamic paraventricular nucleus (PVN) are located primarily in corticotropin releasing hormone (CRH) neurons of the parvicellular subdivision. The purpose of these studies was to investigate the role of All regulating the hypothalamic-pituitary adrenal (HPA) axis, by correlating AT₁ receptor expression levels in the PVN with the known changes in activity of the HPA axis under different stress paradigms, and manipulation of circulating glucocorticoids. AT₁ receptor mRNA was measured by in situ hybridization using ³⁵S-labelled cRNA probes and All binding by autoradiography using ¹²⁵I[Sar¹,lle⁸]All in slide mounted hypothalamic sections. AT₁ receptor mRNA levels and All binding in the PVN were reduced by about 20% 18 h after adrenalectomy remaining at these levels up to 6 days after. This effect was prevented by corticosterone administration in the drinking water, or dexamethasone injection (100 mg, s.c., daily). Conversely, dexamethasone injection in intact rats caused a 20% increase in AT₁ receptor mRNA in the PVN. AT₁ receptor mRNA and binding in the PVN increased 4 h after exposure to stress paradigms associated with activation of the HPA axis (immobilization for 1 h, or i.p. injection of 1.5 M NaCl), and remained elevated after repeated daily stress for 14 days. Unexpectedly, two osmotic stress models associated with inhibition of the HPA axis (60 h water deprivation or 12 days of 2% saline intake) also resulted in increased AT₁ receptor mRNA levels and All binding in the parvicellular PVN. In intact rats, the stimulatory effect of acute stress on AT₁ receptor mRNA in the PVN was significantly enhanced by dexamethasone administration (100 μg, s.c., 14 h and 1 h prior to stress), while in adrenalectomized rats, with or without glucocorticoid replacement, stress reduced rather than increased, AT₁ receptor mRNA. Dexamethasone, 100 μg, injected sc within 1 min the beginning of immobilization in adrenalectomized rats, increased AT₁ receptor mRNA in the PVN to levels significantly higher than those after dexamethasone alone, indicating that the stress induced glucocorticoid surge is required for the stimulatory effect of stress on AT₁ receptor mRNA. The data suggest that AT₁ receptor expression in the PVN is under dual control during stress: stress-activated inhibitory pathways and the stimulatory effect of glucocorticoids. The lack of specificity of the changes in AT₁ receptor expression in the PVN following stressors with opposite effects on ACTH secretion (osmotic and physical-psychological stress) does not support a role for All as a major determinant of the response of the HPA axis during stress.     

Prenatal Stress Increases the Hypothalamo-Pituitary-Adrenal Axis Response in Young and Adult Rats

Prenatal stress is considered as an early epigenetic factor able to induce long-lasting alterations in brain structures and functions. It is still unclear whether prenatal stress can induce long-lasting modifications in the hypothalamo-pituitary-adrenal axis. To test this possibility the effects of restraint stress in pregnant rats during the third week of gestation were investigated in the functional properties of the hypothalamo-pituitary-adrenal axis and hippocampal type I and type II corticosteroid receptors in the male offspring at 3, 21 and 90 days of age. Plasma corticosterone was significantly elevated in prenatally-stressed rats at 3 and 21 days after exposure to novelty. At 90 days of age, prenatally-stressed rats showed a longer duration of corticosterone secretion after exposure to novelty. No change was observed for type I and type II receptor densities 3 days after birth, but both receptor subtypes were decreased in the hippocampus of prenatally-stressed offspring at 21 and 90 days of life. These findings suggest that prenatal stress produces long term changes in the hypothalamo-pituitary-adrenal axis in the offspring.