Dual motor response to l-dopa and nociceptin/orphanin FQ receptor antagonists in 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) treated mice: Paradoxical inhibition is relieved by D2/D3 receptor blockade

Motor activity of mice acutely treated with the parkinsonian toxin 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) was monitored for 6 days using behavioral tests which provide complementary information on motor function: the bar, reaction time, drag, stair climbing, grip, rotarod and footprinting tests. These tests consistently disclosed a prolonged motor impairment characterized by akinesia, bradykinesia, speed reduction, loss of coordination and gait patterns. This impairment was associated with ∼ 60% loss of striatal dopamine terminals, as revealed by tyrosine hydroxylase immunohistochemistry, and was attenuated by dopaminergic drugs. Indeed, the dopamine precursor, l-dopa (1–10 mg/kg), and the D₃/D₂ receptor agonist pramipexole (0.0001–0.001 mg/kg) promoted stepping activity in the drag test (a test for akinesia/bradykinesia). The novel nociceptin/orphanin FQ receptor (NOP) antagonist 1-[1-(cyclooctylmethyl)-1,2,3,6-tetrahydro-5-(hydroxymethyl)-4-pyridinyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one (Trap-101, 0.001–0.1 mg/kg), an analogue of 1-[(3R,4R)-1-cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one (J-113397), also promoted stepping and synergistically or additively (depending on test) attenuated parkinsonism when combined to dopamine agonists. High doses of l-dopa (100 mg/kg), pramipexole (0.1 mg/kg), Trap-101 and J-113397 (1 mg/kg), however, failed to modulate stepping, worsening immobility time and/or rotarod performance. Low doses of amisulpride (0.1 mg/kg) reversed motor inhibition induced by l-dopa and J-113397, suggesting involvement of D₂/D₃ receptors. This study brings further evidence for a dopamine-dependent motor phenotype in MPTP-treated mice reinforcing the view that this model can be predictive of symptomatic antiparkinsonian activity provided the appropriate test is used. Moreover, it offers mechanistic interpretation to clinical reports of paradoxical worsening of parkinsonism following l-dopa. Finally, it confirms that NOP receptor antagonists may be proven effective in reversing parkinsonism when administered alone or in combination with dopamine agonists.     

Functional coupling of the nociceptin/orphanin FQ receptor in dog brain membranes

Nociceptin/orphanin FQ (N/OFQ) is the endogenous ligand for the N/OFQ receptor (NOP) which is yet to be functionally characterized in dog brain. Ligand binding data reports low NOP density (29 fmol mg(-1) protein) in dog. In this study using dog brain membranes, we have examined the effects of N/OFQ on [leucyl-(3)H]N/OFQ(1-17)OH ([leucyl-(3)H]N/OFQ) binding in the presence and absence of 120 mM NaCl and 100 microM GTPgammaS. Data from standard [(35)S]GTPgammaS binding and immunoprecipitation (G(alphai1-3)) assays are also presented, along with data from a limited number of control experiments with human NOP expressed in Chinese hamster ovary (CHO(hNOP)) cells. N/OFQ displaced [leucyl-(3)H]N/OFQ binding with pK(i) and slope values of 9.62+/-0.07 and 0.38+/-0.05, respectively. Addition of NaCl/GTPgammaS produced a steepening (slope 0.95+/-0.06, n=3) of the curve. N/OFQ stimulated [(35)S]GTPgammaS binding with pEC(50) and E(max) values of 8.21+/-0.17 and 1.17+/-0.01, respectively (in CHO(hNOP), pEC(50) and E(max) values were 8.47+/-0.01 and 7.01+/-0.63). N/OFQ stimulated [(35)S]GTPgammaS binding in dog and CHO(hNOP) cell membranes could be immunoprecipitated with an anti-G(alphai1-3) antibody, indicating coupling to a pertussis toxin (PTx)-sensitive G-protein. N/OFQ actions were competitively antagonized by the selective NOP antagonists, 100 nM J-113397, 1 microM [Nphe(1)]N/OFQ(1-13)NH(2) and 1 microM [Phe(1)Psi(CH(2)-NH)Gly(2)]N/OFQ(1-13)NH(2) (partial agonist) yielding pK(B) values of 8.58+/-0.21, 7.06+/-0.59 and 7.32+/-0.41, respectively (in CHO(hNOP), a pK(B) for J-113397 of 8.33+/-0.02 was obtained). Despite relatively low receptor density, we were able to detect functional activity of native dog NOP, with pharmacology consistent with reports for other species.     

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.     

Pharmacological profile of nociceptin/orphanin FQ receptors regulating 5-hydroxytryptamine release in the mouse neocortex

A synaptosomal preparation was employed to pharmacologically characterize the role of presynaptic nociceptin/orphanin FQ (N/OFQ) receptors (NOP receptors) in the regulation of 5-hydroxytryptamine release in the Swiss mouse neocortex. In the present study, the NOP receptor ligands N/OFQ, Ac-RYYRWK-NH(2) and [Phe(1)psi(CH(2)-NH)Gly(2)]N/OFQ(1-13)-NH(2) inhibited the K(+)-induced [(3)H]-5-HT overflow with similar maximal effects ( approximately -35%) but different potencies (pEC(50) of 8.56, 8.35 and 7.23, respectively). The novel agonist [Arg(14),Lys(15)]N/OFQ also inhibited [(3)H]-5-HT overflow, but the concentration-response curve was biphasic and the efficacy higher ( approximately -45%). Receptor selectivity of NOP receptor agonists was demonstrated by showing that synaptosomes from NOP receptor knockout mice were unresponsive to N/OFQ, [Arg(14),Lys(15)]N/OFQ and [Phe(1)psi(CH(2)-NH)Gly(2)]N/OFQ(1-13)-NH(2) but maintained full responsiveness to endomorphin-1. Moreover, the inhibitory effect of N/OFQ was prevented by peptide ([Nphe(1)]N/OFQ(1-13)-NH(2) and UFP-101) and nonpeptide (J-113397 and JTC-801) NOP receptor selective antagonists. Desensitization occurred under perfusion with high (3 and 10 microm) N/OFQ concentrations. This phenomenon was prevented by the protein kinase C inhibitor, bisindolylmaleimide. Moreover, N/OFQ-induced desensitization did not affect mu opioid receptor responsiveness. Finally, it was observed in a similar preparation of rat cerebrocortical synaptosomes, although it was induced by higher N/OFQ concentrations than that used in the mouse. Together, these findings indicate that presynaptic NOP receptors inhibit 5-hydroxytryptamine release in the mouse neocortex. Based on present and previous studies, we conclude that NOP receptors in the mouse are subtly different from the homologous receptor population in the rat, strengthening the view that there exist species differences in the pharmacology of central NOP receptors.     

Dopamine-nociceptin/orphanin FQ interactions in the substantia nigra reticulata of hemiparkinsonian rats: involvement of D2/D3 receptors and impact on nigro-thalamic neurons and motor activity

Nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor antagonists proved to be effective in alleviating experimental parkinsonism. Nonetheless, loss of effectiveness or even worsening of parkinsonian symptoms have been observed at high doses. With the aim of clarifying the circuitry underlying the dual action of NOP receptor antagonists and the role of endogenous dopamine, the NOP receptor antagonist 1-benzyl-N-[3-[spiroisobenzofuran-1(3H),4′-piperidin-1-yl]propyl]pyrrolidine-2-carboxamide (Compound 24) and the D₂/D₃ receptor antagonist raclopride were used in 6-hydroxydopamine hemilesioned rats. Systemically administered Compound 24 improved motor activity in the 0.1–10 mg/kg dose range being ineffective at 30 mg/kg. To confirm NOP selectivity, Compound 24 improved motor performance in wild-type mice at 1 and 10 mg/kg and inhibited it at 60 mg/kg, being ineffective in NOP receptor knockout mice. To prove that the bell-shaped profile was mediated by nigral NOP receptors, reverse dialysis of Compound 24 (0.03 μM) in substantia nigra reticulata ameliorated akinesia whereas Compound 24 (3 μM) was ineffective. To demonstrate that motor responses were mediated by tuning inhibitory and excitatory inputs to nigro-thalamic neurons, the low concentration elevated GABA and reduced glutamate in substantia nigra, simultaneously reducing GABA levels in ventro-medial thalamus. Conversely, the higher concentration reduced nigral and elevated thalamic GABA, without affecting nigral glutamate levels. Co-perfusion with raclopride (1 μM) abolished the antiakinetic action of Compound 24 (0.03 μM) and turned the ineffectiveness of Compound 24 (3 μM) into an antiakinetic effect. The low concentration reduced nigral but did not affect thalamic GABA whereas the higher concentration elevated nigral and reduced thalamic GABA. Neither concentration affected nigral glutamate. We conclude that dual motor effects of Compound 24 in hemiparkinsonian rats are accomplished through blockade of nigral NOP receptors resulting in opposite modulation of nigro-thalamic neurons. Endogenous dopamine contributes to these responses affecting the level of GABAergic inhibition of the nigral output via D₂/D₃ receptors.     

Orphanin FQ/nociceptin suppresses motor activity through an action along the mesoaccumbens axis in rats

OBJECTIVE: Intracerebroventricular administration of orphanin FQ/nociceptin (OFQ/N), the endogenous agonist ligand of the opioid receptor-like (ORL-1) receptor, decreases extracellular levels of dopamine and suppresses motor activity. The presence of the ORL-1 receptor on mesoaccumbal and nigrostriatal dopaminergic neurons raises the possibility that an action along these pathways may be one means by which OFQ/N produces motor suppression. Thus, the present study used local administration of OFQ/N into the ventral tegmental area (VTA), the substantia nigra, the nucleus accumbens and the striatum to determine the contribution of cell-body regions and terminal fields of the dopaminergic neurons to the motor-suppressant effect of OFQ/N. METHODS: Rats were implanted bilaterally with guide cannulae into one of the brain regions and tested 4 days later. First, the effect of a single dose of OFQ/N (30 microg/0.5 microL per side) on motor activity was determined after direct injection into the VTA, substantia nigra, nucleus accumbens or striatum. Rats were habituated to activity chambers for 1 hour and then injected with either artificial cerebrospinal fluid or OFQ/N into one of the brain regions, and motor activity was recorded for a further 1 hour. Next, the dose-response effect of intra-VTA or intranigral OFQ/N (3 microg or 30 microg/0.5 microL per side) on motor activity was examined. Finally, the effect of intra-VTA OFQ/N (3 microg or 30 microg/0.5 microL per side) on motor activity was determined in the presence of J-113397, an ORL-1 receptor antagonist. RESULTS: OFQ/N suppressed motor activity when injected into the VTA and to a lesser extent after direct injection into the nucleus accumbens. However, OFQ/N failed to attenuate motor activity significantly after injection into the substantia nigra or the striatum. Subsequent dose-response studies showed that OFQ/N suppressed motor activity even at a 10-fold-lower dose after intrategmental but not intranigral administration. The motor-suppressant action of intra-VTA OFQ/N was attenuated by J-113397 (1.5 microg/0.5 microL per side) administered into the VTA 10 minutes before administration of OFQ/N. CONCLUSION: Our results indicate that OFQ/N suppresses motor activity through activation of the ORL-1 receptor primarily through an action in the VTA.     

Activation of nociceptin/orphanin FQ peptide receptors disrupts visual but not auditory sensorimotor gating in BALB/cByJ mice: comparison to dopamine receptor agonists

Nociceptin/orphanin FQ (N/OFQ) peptide and its receptor (NOP receptor) have been implicated in a host of brain functions and diseases, but the contribution of this neuropeptide system to behavioral processes of relevance to psychosis has not been investigated. We examined the effect of the NOP receptor antagonists, Compound 24 and J-113397, and the synthetic agonist, Ro64-6198, on time function (2-2000?ms prepulse-pulse intervals) of acoustic (80?dB/10?ms prepulse) and visual (1000?Lux/20?ms prepulse) prepulse inhibition of startle reflex (PPI), a preattentive sensory filtering mechanism that is central to perceptual and mental integration. The effects of the dopamine D1-like receptor agonist, SKF-81297, the D2-like receptor agonist, quinelorane, and the mixed D1/D2 agonist, apomorphine, were studied for comparison. When acoustic stimulus was used as prepulse, BALB/cByJ mice displayed a monotonic time function of PPI, and consistent with previous studies, apomorphine and SKF-81279 induced PPI impairment, whereas quinelorane had no effect. None of the NOP receptor ligands was effective on acoustic PPI. When flash light was used as prepulse, BALB/cByJ mice displayed a bell-shaped time function of PPI and all dopamine agonists were active. Ro64-6198 was also effective in reducing visual PPI. NOP receptor antagonists showed no activity but blocked disruptive effect of Ro64-6198. Finally, coadministration of the typical antipsychotic, haloperidol, attenuated PPI impairment induced by Ro64-6198, revealing involvement of a dopaminergic component. These findings show that pharmacological stimulation of NOP or dopamine D2-like receptors is more potent in disrupting visual than acoustic PPI in mice, whereas D1-like receptor activation disrupts both. They further suggest that dysfunction of N/OFQ transmission may be implicated in the pathogenesis of psychotic manifestations.     

Effects of co-administration of anticonvulsant and putative anticonvulsive agents and sub-/suprathreshold doses of L-Dopa upon motor behaviour of MPTP-treated mice

The effects of co-administration of the dopamine precursor, L-Dopa, with anticonvulsant and putative anticonvulsive agents upon the motor activity of hypoactive MPTP-treated C57 BL/6 mice were measured in six experiments. In each case, MPTP (2 x 40 mg/kg, s.c., separated by a 24-hr interval) was administered four to six weeks prior to behavioural testing. Thus, the effects of these agents combined with either a single acute, subthreshold dose (5 mg/kg, s.c.) of L-Dopa, or, with chronically-administered, suprathreshold doses (20 mg/kg, s.c.) of L-Dopa were studied. In the former, lamotrigine, FCE 26743 and L-Deprenyl, injected 60 min before subthreshold L-Dopa (5 mg/kg), each induced an antiparkinsonian action in MPTP-treated mice that consisted of dose-specific, as opposed to dose-related, elevations of locomotion and rearing behaviour. In the latter, lamotrigine (all three measures of activity at 3 mg/kg), FCE 26743 (locomotion and total activity at 3; rearing at 1 and 3 mg/kg) and L-Deprenyl (locomotion and total activity at 1 and 3mg/kg), but not phenytoin (neither at 1 nor 3 mg/kg), reinstated the motor activity-stimulating effects of the threshold dose of L-Dopa (20 mg/kg) in L-Dopa-tolerant, MPTP-treated mice. Neurochemical analyses confirmed severe DA depletions in MPTP-treated mice. Since neither lamotrigine, FCE 26743 nor L-Deprenyl, nor subthreshold L-Dopa, by themselves increased the motor behaviour of MPTP-treated mice, a synergistic effect of the co-administration is concluded. Further, since the suprathreshold dose of L-Dopa by itself failed to stimulate motor activity in the MPTP mice following chronic (25 daily injections) administrations of the compound, it is suggested that a restorative effect, in combination with lamotrigine, FCE 26743 or L-Deprenyl was evidenced. The potential therapeutic benefits of anticonvulsant or putative anticonvulsive compounds for parkinsonian symptoms are discussed.     

NOP receptor antagonist, JTC-801, blocks cannabinoid-evoked hypothermia in rats

The present study used the endpoint of hypothermia to investigate cannabinoid and nociceptin/orphanin FQ (N/OFQ) interactions in conscious animals. Prior work has established that cannabinoids produce hypothermia by activating central cannabinoid CB(1) receptors. The administration of N/OFQ into the brain also causes significant hypothermia. Those data suggest a link between cannabinoid CB(1) receptors and N/OFQ peptide (NOP) receptors in the production of hypothermia. Therefore, we determined if NOP receptor activation is required for cannabinoid-evoked hypothermia and if cannabinoid CB(1) receptor activation is necessary for N/OFQ-induced hypothermia. In actual experiments, a cannabinoid agonist, WIN 55212-2 (2.5, 5, and 10 mg/kg, i.p.), caused significant hypothermia in male Sprague-Dawley rats (200-225 g). A NOP receptor antagonist, JTC-801 (1 mg/kg, i.p.), did not affect body temperature. For combined administration, JTC-801 (1 mg/kg, i.p.) blocked a significant proportion of the hypothermia caused by each dose of WIN 55212-2 (2.5, 5, and 10 mg/kg, i.p.). JTC-801 (1 mg/kg, i.p.) also blocked the hypothermia caused by another cannabinoid agonist, CP-55, 940 (1 mg/kg, i.p.). In separate experiments, the direct administration of N/OFQ (9 microg/rat, i.c.v.) into the brain produced significant hypothermia. The hypothermic effect of N/OFQ was blocked by JTC-801 (1 mg/kg, i.p.) but not by a selective cannabinoid CB(1) antagonist, SR 141716A (5 mg/kg, i.m.). The finding that a NOP receptor antagonist abolishes a significant percentage of cannabinoid-induced hypothermia suggests that NOP receptor activation is required for cannabinoids to produce hypothermia. This interaction, quantitated in the present study, is the first evidence that NOP receptors mediate a cannabinoid-induced effect in conscious animals.     

Endogenous nociceptin/orphanin‐fq in the dorsal hippocampus facilitates despair‐related behavior

Nociceptin/orphanin‐FQ (N/OFQ) peptide and its receptor (NOP: N/OFQ opioid peptide receptor) are highly expressed in the hippocampus, but their functional role remains poorly understood. We recently showed that hippocampal N/OFQ inhibits learning and memory abilities in mice. Here, we investigated whether the endogenous peptide also regulated emotional responses at the level of the hippocampus. Bilateral infusions of the selective NOP receptor antagonist, UFP‐101 (1–3 nmol/side), into the dorsal hippocampus produced antidepressant‐like effects in the mouse forced swim and tail suspension tests comparable with those obtained with the prototypical antidepressant, fluoxetine (10–30 mg/kg, intraperitoneal). In the light‐dark test, neither UFP‐101 (1–3 nmol/side) nor N/OFQ peptide (1–3 nmol/side) modified anxiety measures when injected at behaviorally active doses in the dorsal hippocampus. These findings show a clear dissociation in the involvement of hippocampal N/OFQ system in anxiety‐ and despair‐related behaviors. We conclude that the dorsal hippocampus is a brain region in which there is an important N/OFQ modulation of mnemonic processes and adaptive emotional responses associated to despair states. © 2010 Wiley‐Liss, Inc.     

Brain structures implicated in the four-plate test in naïve and experienced Swiss mice using injection of diazepam and the 5-HT2A agonist DOI

Four-plate test-retest (FPT-R) is a useful tool to study aversive memory and abolishment of benzodiazepine effects in experienced mice to four-plate test (FPT), namely one-trial tolerance. In the present study, we have used local injections paradigm, in order to localize structures implied in anxiolytic-like effects of two drugs in naïve and experienced mice: a benzodiazepine, diazepam that is only active in naïve mice; and a 5-HT_2A/2C agonist, DOI that exert its anxiolytic-like effect both in naïve and experienced mice. Periacqueductal grey substance, three sub-regions of hippocampus (CA1, CA2 and CA3) and two nuclei of amygdala (BLA and LA) have been studied. Local injections did not cause any modifications of ambulatory activity. DOI injections elicit anxiolytic-like effects only when injected into CA2, in naïve and experienced mice. Diazepam had an anxiolytic-like effect in naïve mice, only when injected into lateral nucleus of amygdala; and in experienced mice when injected into PAG. These results help us to better understand the way of action of these two compounds and the structures functionally involved in their effects and in one-trial tolerance (OTT).     

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     

Kainic Acid Down-regulates NOP Receptor Density and Gene Expression in Human Neuroblastoma SH-SY5Y Cells

Nociceptin (N/OFQ) is involved in neuronal excitability and in certain types of seizures. Kainate-induced seizures are associated with increased N/OFQ release in the rat thalamus and hippocampus, causing down-regulation of the N/OFQ receptor (NOP). In this study, we used the neuroblastoma SH-SY5Y cell line as a model to investigate the effects of kainate on NOP receptor density and gene expression. Exposure to kainate (10–50 μM) for 3 h did not affect NOP receptor density. In contrast, a NOP B _max down-regulation was detected in cells exposed to 10 μM kainate for both 6 and 24 h. Moreover, our data show that kainate causes a decrease in NOP mRNA levels after 3, 6, and 24 h, an effect blocked by the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). These findings show that kainate is able to affect the NOP system, both at biosynthesis and receptor density levels in SH-SY5Y cells, and that the kainate ionotropic receptor can contribute to the regulation of the NOP receptor. These data are in agreement with data obtained in vivo and provide new evidence concerning the existence of a cross-talk between NOP and kainate receptors, leading to an interplay between glutamate and N/OFQ circuits. Cannarsa and Landuzzi contributed equally to this work.     

Synergistic interactions between COMT-/MAO-inhibitors and L-Dopa in MPTP-treated mice

Summary Four experiments were performed to investigate the anti-akinesia effects of combining a sub-threshold dose (5mg/kg, s.c.) of L-Dopa with different doses and combinations of COMT and MAO inhibitors upon the hypokinesia observed in MPTP-treated mice. Ro 40-7592 (1 and 3 mg/kg, s.c.), a novel COMT inhibitor, 60 min before L-Dopa reinstated both locomotion and rearing during a 2-hr interval after L-Dopa in MPTP mice; control mice were unaffected. The combination of Ro 40-7592 (3 mg/kg, s.c.) and pargyline (5 mg/kg, s.c.), a MAO inhibitor, with L-Dopa produced increases in both the peak effect and duration of action indicating a distinct potentiation of the effects of Ro 40-7592 by pargyline. L-Deprenyl, a MAO_B inhibitor, together with L-Dopa, restored locomotion and rearing behaviour at all three doses applied (1, 3 and 10 mg/kg, s.c.); in control mice, motor activity was stimulated at the higher doses (3 and 10 mg/kg, s.c.), independent of L-Dopa administration. Combining L-Deprenyl (3 mg/kg, s.c.) with Ro 40-7592 (3 mg/kg, s.c.) one hr before L-Dopa to MPTP mice potentiated the restorative effects of each compound by itself, although no increase in peak effect was obtained. In the control mice, L-Deprenyl plus Ro 40-7592 or L-Deprenyl, by itself, stimulated motor activity following injection of L-Dopa. Marked dopamine (DA) depletions in the striatum of MPTP-treated mice were evident. The present results demonstrate that the effects of the COMT/MAO inhibitors in combination, and in conjunction with L-Dopa (at a dose that was without effect by itself), were well in excess of a summation of their individual effects. It was concluded therefore that a synergism of the restorative, anti-akinesic action of these compounds in MPTP-treated mice could offer a broader therapeutic spectrum in the treatment of Parkinson's disorder.     

Antiparkinsonian actions of CP-101,606, an antagonist of NR2B subunit-containing N-methyl-d-aspartate receptors

In the setting of nigrostriatal dopamine depletion, glutamatergic pathways to the striatum and basal ganglia output nuclei become overactive. Systemically administered glutamate receptor antagonists may have direct antiparkinsonian actions in rodents, but there is little evidence for this in primates. Glutamate antagonists may also potentiate conventional dopaminergic therapies; however, there is concern that broad spectrum, nonselective antagonists may have unwanted side-effects. Because subunit-selective antagonists may avoid these liabilities, we have examined the antiparkinsonian effects of a selective antagonist of the NR2B subunit of the NMDA receptor. In rats, CP-101,606 decreased haloperidol-induced catalepsy with an ED(50) of about 0.5 mg/kg. In MPTP-treated monkeys, CP-101,606 (1 mg/kg) reduced parkinsonian motor symptoms by 20%. At a dose of 0.05 mg/kg, CP-101,606 markedly potentiated the effect of a submaximal dose of levodopa, reducing motor symptoms by about 50% compared to vehicle and by about 30% compared to levodopa alone. No side-effects were apparent at any dose of CP-101,606. We conclude that CP-101,606 has direct antiparkinsonian actions in both rodents and monkeys and it synergistically potentiates levodopa in MPTP-treated monkeys. Clinical evaluation of selective NR2B antagonists may be warranted in Parkinson's disease.     

Effects of non-peptide nociceptin/orphanin FQ receptor ligands on methylphenidate-induced hyperactivity in mice: Implications for bipolar disorders

Bipolar disorder is a psychiatric pathology characterized by biphasic mood episodes of mania or hypomania and depression. The pharmacotherapy of bipolar disorder has significant adverse effects impairing treatment adherence and patient quality of life. The N/OFQ-NOP receptor system has been widely implicated with mood disorders. Clinical and preclinical findings suggest antidepressants actions for NOP antagonists. More recently, the administration of NOP agonists has shown to promote depressant states. The present study aimed to investigate the effects of non-peptide NOP ligands in methylphenidate-induced manic-like behavior in mice. The NOP agonist Ro 65–6570 (0.01–1 mg/kg, ip), at the higher dose, did not affect spontaneous locomotion per se, but prevented the methylphenidate (10 mg/kg, sc)-induced hyperlocomotion. The NOP partial agonist AT-090 (0.001–0.03 mg/kg, ip) and the NOP antagonist SB-612111 (1–10 mg/kg, ip) did not significantly affect the psychostimulant-induced hyperactivity. Experiments performed with mice lacking the NOP receptor (NOP(−/−)) demonstrated that the treatment with methylphenidate induced similar hyperlocomotion in NOP(−/−) and NOP(+/+) mice. In conclusion, these findings suggest a potential role for NOP agonists in the prevention of manic states, especially by counteracting the hyperactivity symptom of bipolar patients. However, more studies are necessary in order to evaluate these compounds in other features of bipolar disorder.     

Site-specific estrogen and progestin regulation of orphanin FQ/nociceptin and nociceptin opioid receptor mRNA expression in the female rat limbic hypothalamic system

The distributions of orphanin FQ (OFQ/N; also known as nociceptin) and its cognate receptor, opioid receptor-like receptor-1 (NOP), overlap steroid-responsive regions throughout reproductive circuits of the limbic system and hypothalamus. For example, in the ventromedial nucleus of the hypothalamus (VMH), OFQ/N facilitates lordosis in female rats through estrogen and progesterone regulation of nociceptin activity. We studied estrogen and progesterone regulation of OFQ/N and NOP mRNA expression in limbic-hypothalamic reproductive circuits. Ovariectomized rats were treated with 17beta-estradiol-benzoate (2 microg) and 26 hours later with oil or progesterone (500 microg) and were killed 30 hours after initial treatment. Alternate brain sections were processed for OFQ/N or NOP mRNA in situ hybridization. High levels of hybridization for NOP and OFQ/N and overlapping distributions were observed throughout the limbic hypothalamic reproductive circuits; however, in VMH, only NOP expression was observed. Estrogen treatment increased NOP mRNA expression in anteroventral periventricular nucleus (AVPV), median preoptic nucleus, and VMH. Subsequent progesterone treatment did not alter estrogen-induced expression of NOP mRNA in VMH or median preoptic nucleus but reduced expression in the AVPV. OFQ/N mRNA levels were also regulated by steroids. In the caudal part of the posterodorsal medial amygdala, estrogen increased OFQ/N mRNA levels, and progesterone did not alter this increase, whereas, in the medial part of the medial preoptic nucleus, estrogen and progesterone were needed to increase OFQ/N mRNA levels. Steroid regulation of OFQ/N and NOP in the medial preoptic nucleus and VMH is consistent with emerging data indicating that this opioid system regulates female reproduction.     

Effect of the alpha 2 adrenoreceptor antagonist, idazoxan, on motor disabilities in MPTP-treated monkey

1. The motor effect of the alpha 2 adrenoreceptor antagonist, idazoxan, was compared to that of L-dopa in MPTP-treated monkeys. 2. Idazoxan 2.0 mg/kg improved parkinsonian motor abnormalities which was comparable to the effects of a minimal effective dose of L-dopa. 3. At 2.0 and 5.0 mg/kg, the parkinsonian rigidity was the item most frequently alleviated by idazoxan (respectively 63.6% and 68.2%). 4. These findings provide support for the therapeutic utility of alpha 2 antagonists in the treatment of Parkinson's disease.     

Normal sensitivity to acute pain,but increased inflammatory hyperalgesia in mice lacking the nociceptin precursor polypeptide or the nociceptin receptor

Nociceptin/orphanin FQ (N/OFQ) is the endogenous agonist of the N/OFQ peptide receptor (NOP receptor). It is released from a larger precursor polypeptide, called prepro-nociceptin (ppN/OFQ) from which, in addition to N/OFQ, other biologically active neuropeptides may be derived. Increasing evidence indicates that exogenous application of N/OFQ to the central nervous system of mice and rats induces pro- and antinociceptive effects depending on the dose and site of administration. Much less is known about a potential contribution of endogenous N/OFQ to pain control. Here, we have used a genetic approach to address this topic. Mice deficient in either the NOP receptor (NOP-R-/- mice) or the N/OFQ precursor polypeptide (ppN/OFQ-/- mice) or both (double knockout mice) were compared with wild-type littermates in animal models of acute and tonic pain. Nociceptive responses to acute noxious heat of all three types of mutant mice were indistinguishable from those of wild-type mice. Accordingly, nociceptive behaviour was very similar in the early phase of the formalin test. However, NOP-R-/-, ppN/OFQ-/- and double knockout mice showed markedly stronger nociceptive responses during prolonged nociceptive stimulation in the second phase of the formalin test and significantly lower thermal pain thresholds in inflamed tissue after zymosan A injection. These results indicate that N/OFQ contributes significantly to endogenous pain control during prolonged nociceptive stimulation but does not affect acute pain sensitivity. Among the three types of mutant mice nociceptive behaviour was nearly identical, indicating that the lack of other potential ppN/OFQ products in the ppN/OFQ-/- mice was apparently without effect on the nociceptive phenotype.