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Objective Mutations in the CDKL5 gene cause an early-onset epileptic encephalopathy. To date, little is known about effective antiepileptic treatment in this disorder. Method Accordingly, the aim of this retrospective study was to explore the role of different antiepileptic drugs (AEDs) and the ketogenic diet (KD) in the treatment of this rare genetic disorder. We evaluated the efficacy in 39 patients with CDKL5 mutations at 3, 6 and 12 months after the introduction of each treatment. One patient was lost to follow-up after 6 and 12 months. Results The responder rate (>50% reduction in seizure frequency) to at least one AED or KD was 69% (27/39) after 3 months, 45% (17/38) after 6 months and 24% (9/38) after 12 months. The highest rate of seizure reduction after 3 months was reported for FBM (3/3), VGB (8/25), CLB (4/17), VPA (7/34), steroids (5/26), LTG (5/23) and ZNS (2/11). Twelve patients (31%) experienced a seizure aggravation to at least one AED. Most patients showed some but only initial response to various AEDs with different modes of actions. Significance Considering both age-related and spontaneous fluctuation in seizure frequency and the unknown impact of many AEDs or KD on cognition, our data may help defining realistic treatment goals and avoiding overtreatment in patients with CDKL5 mutations. There is a strong need to develop new treatment strategies for patients with this rare mutation. [ABSTRACT FROM AUTHOR]

Abstract: Rasmussen encephalitis (RE) is a chronic inflammatory disease leading to unilateral hemispheric atrophy, associated with progressive neurological dysfunction and intractable seizures. The best approach to RE is hemispherectomy. However long-term immunotherapy seems to prevent or slow down hemispheric tissue loss and the associated functional decline. We describe a girl with epilepsia partialis continua (EPC) and progressive neurological dysfunction compatible with RE. The brain MRI showed a lesion that was initially interpreted as focal cortical dysplasia. Combined antiepileptic and immunomodulation were administered for two years with initial beneficial effects. The follow-up MRI, 4year later showed. atrophic change in right parietal region. The association of antiepileptic and immunomodulation therapies may inhibit pathogenetic mechanisms responsible for neuronal loss in RE, slowing down the progression of the disease. [Copyright &y& Elsevier]

Abstract: Nociceptin/orphanin FQ (N/OFQ), the endogenous NOP receptor ligand, centrally modulates gastric motor and secretory functions and prevents ethanol-induced gastric lesions in rats. A recently synthesized N/OFQ analog, [(pF)Phe4Aib7Arg14Lys15]N/OFQ-NH2 (UFP-112), acts as a highly potent and selective peptide agonist for NOP receptors and produces longer-lasting in vitro and in vivo effects in mice than the natural ligand N/OFQ. In this study, we evaluated the effects of centrally (intracerebroventricularly/icv) and peripherally (intraperitoneally/ip) injected UFP-112 on gastric emptying and gastric acid secretion, and on the development of gastric mucosal lesions induced by 50% ethanol in the rat. When injected icv, it dose-dependently delayed gastric emptying of a phenol red meal (by up to 70%), decreased gastric secretion in water-loaded rats after 90 pylorus ligature, and reduced ethanol-induced gastric lesions (by up to 87%). In all three assays, UFP-112 was more effective than N/OFQ. The highly selective NOP receptor antagonist, UFP-101, decreased the efficacy of UFP-112, thus confirming that central NOP receptors mediate inhibitory control on these functional and pathological conditions in rats. Ip injected N/OFQ and UFP-112 induced non-dose-related gastric hypersecretory and antiulcer effects, which UFP-101 partially abolished. Ip N/OFQ appeared equiactive but about 30–100 times less potent than ip UFP-112 in stimulating gastric acid secretion and preventing lesion formation. When ip injected, both UFP-112 and N/OFQ left gastric emptying in rats unchanged, suggesting that peripheral NOP receptors have a role in mediating gastric hypersecretory and antiulcer effects but are not involved in regulating gastric motility. In addition, the inhibitory effects induced by this novel NOP receptor agonist lasted longer than those induced by N/OFQ. In conclusion, UFP-112 is a promising new pharmacological tool for studying the functional roles of the central and peripheral N/OFQ receptor system. [Copyright &y& Elsevier]

Intracerebral microdialysis was employed in awake freely moving rats to investigate the effects of nociceptin/orphanin FQ receptor ligands on glutamate extracellular levels in the substantia nigra pars reticulata. Nociceptin/orphanin FQ, ineffective at 0.1 μM, induced a prolonged stimulation of nigral glutamate levels at 1 and 10 μM (mean effect of 137±9 and 167±13%, respectively, of basal values). These effects were prevented by the novel nociceptin/orphanin FQ receptor antagonist [Nphe1]nociceptin/orphanin FQ(1-13)NH2 (100 and 300 μM, respectively) but not by the non-selective opioid receptor antagonist naloxone (10 μM). [Nphe1]nociceptin/orphanin FQ(1-13)NH2 (100 μM) inhibited by itself glutamate outflow (maximal reduction to 71±4%) while naloxone was ineffective. The nociceptin/orphanin FQ receptor ligand [Phe1ψ(CH2-NH)Gly2]nociceptin/orphanin FQ(1-13)NH2 also facilitated glutamate outflow at 10 μM (mean effect of 145±10%). Intranigral perfusion with tetrodotoxin (1 μM) or with the dopamine D2 receptor antagonist raclopride (1 μM), failed to affect basal glutamate output and prevented the facilitatory effect of nociceptin/orphanin FQ (10 μM). However, perfusion with the GABAA receptor antagonist bicuculline (10 μM) increased local glutamate extracellular levels by itself and attenuated the effect of the peptide.Our data suggest that nociceptin/orphanin FQ increases glutamate extracellular levels in the substantia nigra pars reticulata via activation of nociceptin/orphanin FQ receptors located on non-glutamatergic, possibly dopaminergic and GABAergic, neuronal elements. [Copyright &y& Elsevier]

Background: While producing good-quality analgesia, µ-opioid (MOP) receptor activation produces a number of side-effects including tolerance. Simultaneous blockade of δ-opioid (DOP) receptors has been shown to reduce tolerance to morphine. Here, we characterize a prototype bifunctional opioid H-Dmt-Tic-Gly-NH-Bzl (UFP-505).Methods: We measured receptor binding affinity in Chinese hamster ovary (CHO) cells expressing recombinant human MOP, DOP, k-opioid (KOP), nociceptin/orphanin (NOP) receptors. For activation, we measured the binding of GTPγ(35)S to membranes from CHO(hMOP), CHO(hDOP), rat cerebrocortex, and rat spinal cord. In addition, we assessed 'end organ' responses in the guinea pig ileum and mouse vas deferens.Results: UFP-505 bound to CHO(hMOP) and CHO(hDOP) with (binding affinity) pK(i) values of 7.79 and 9.82, respectively. There was a weak interaction at KOP and NOP (pK(i) 6.29 and 5.86). At CHO(hMOP), UFP-505 stimulated GTPγ(35)S binding with potency (pEC(50)) of 6.37 and in CHO(hDOP) reversed the effects of a DOP agonist with affinity (pK(b)) of 9.81 (in agreement with pK(i) at DOP). UFP-505 also stimulated GTPγ(35)S binding in rat cerebrocortex and spinal cord with pEC(50) values of 6.11-6.53. In the guinea pig ileum (MOP-rich preparation), UFP-505 inhibited contractility with pEC(50) of 7.50 and in the vas deferens (DOP-rich preparation) reversed the effects of a DOP agonist with an affinity (pA(2)) of 9.15.Conclusions: We have shown in a range of preparations and assays that UFP-505 behaves as a potent MOP agonist and DOP antagonist; a MOP/DOP bifunctional opioid. Further studies in dual expression systems and whole animals with this prototype are warranted. [ABSTRACT FROM AUTHOR]

Abstract: Nociceptin/orphanin FQ (N/OFQ) administered into the lateral left cerebral ventricle of rats has been reported to inhibit in vivo gut motor and secretory functions. Recently, a novel N/OFQ analog, [Arg14, Lys15] N/OFQ, was synthesized and demonstrated to behave as a highly potent agonist at the human recombinant N/OFQ peptide (NOP) receptors and to produce long-lasting effects in vivo in mice compared with the natural ligand N/OFQ. In the present study, the pharmacological profile of [Arg14, Lys 15] N/OFQ was further evaluated and compared with that of N/OFQ in vitro on guinea pig exocrine pancreas and in vivo on gastric emptying, colonic propulsion and gastric acid secretion in rats. [Arg14, Lys15] N/OFQ and N/OFQ significantly decreased the KCl-evoked amylase secretion from isolated pancreatic lobules of the guinea pig. In in vivo experiments, [Arg14, Lys15] N/OFQ mimicked the effects of N/OFQ, inducing, after intracerebroventricular injection, a delay (up to 70%) in the gastric emptying of a phenol red meal, an increase (about 40 times) of the mean bead colonic expulsion time and a decrease (up to 90%) of gastric acid secretion in water loaded rats after 90min pylorus ligature. In all these assays, [Arg14, Lys15] N/OFQ was more effective than N/OFQ, and its effective doses were at least 10-fold lower than N/OFQ effective doses. The highly selective NOP receptor antagonist, UFP-101, decreased the efficacy of [Arg14, Lys15] N/OFQ in in vitro and in vivo assays above reported. These findings: (a) show that pancreatic NOP receptors mediate an in vitro inhibitory effect on stimulated guinea pig amylase secretion; (b) confirm that the stimulation of central NOP receptors exerts an inhibitory control on gastric emptying, colonic motility and gastric secretion in rats and (c) put in evidence that [Arg14, Lys15] N/OFQ, being more potent and effective than the natural ligand N/OFQ, represents a new pharmacological tool for the study of the physiological and pharmacological roles mediated by the N/OFQ–NOP receptor system. [Copyright &y& Elsevier]

Myxoid glioneuronal tumor (MGT) is a benign glioneuronal neoplasm recently introduced in the World Health Organization (WHO) classification of the central nervous system (CNS) tumors. MGTs are typically located in the septum pellucidum, foramen of Monro or periventricular white matter of the lateral ventricle. They were previously diagnosed as dysembryoplastic neuroepithelial tumors (DNT), showing histological features almost indistinguishable from classical cortical DNT. Despite that, MGTs have been associated with a specific dinucleotide substitution at codon 385 in the platelet-derived growth factor receptor alpha (PDGFRA) gene, replacing a lysine residue with either leucine or isoleucine (p. LysK385Leu/Iso). This genetic variation has never been described in any other CNS tumor. Thirty-one consecutive tumors, previously diagnosed as DNTs at the Meyer Children's Hospital IRCCS between January 2010 and June 2021 were collected for a comprehensive study of their clinical, imaging, pathological features, and molecular profile. In six out of the thirty-one tumors we had previously diagnosed as DNTs, we identified the recurrent dinucleotide mutation in the PDGFRA. All six tumors were typically located within the periventricular white matter of the lateral ventricle and in the septum pellucidum. We then renamed these lesions as MGT, according to the latest WHO CNS classification. In all patients we observed an indolent clinical course, without recurrence. MGT represent a rare but distinct group of neoplasm with a typical molecular profiling, a characteristic localization, and a relative indolent clinical course. [ABSTRACT FROM AUTHOR]

Background and purpose Nociceptin/orphanin FQ (N/OFQ) receptor (NOP) agonists produce anxiolytic-like effects in rodents while antagonists promote antidepressant-like effects. The aim of this study was to investigate the effect on anxiety and depression of NOP receptor partial agonists such as the peptides [F/G]N/OFQ(1-13)NH 2 and UFP-113 and the non-peptide AT-090. Experimental approach In vitro AT-090, UFP-113, and [F/G]N/OFQ(1-13)NH 2 were tested for their ability to promote NOP/G-protein and NOP/β-arrestin 2 interaction, using a bioluminescence resonance energy transfer assay. In vivo , they were tested in mice in the elevated plus maze (EPM) and in the forced swim (FST) tests. NOP partial agonists effects were systematically compared to those of full agonists (N/OFQ and Ro 65-6570) and antagonists (UFP-101 and SB-612111). Key results In vitro , AT-090, UFP-113, and [F/G]N/OFQ(1-13)NH 2 promoted NOP/G protein interaction, with maximal effects lower than those evoked by N/OFQ and Ro 65-6570. AT-090 behaved as a NOP partial agonist also in inducing β-arrestin 2 recruitment, while UFP-113 and [F/G]N/OFQ(1-13)NH 2 were inactive in this assay. In vivo , AT-090 induced anxiolytic-like effects in the EPM but was inactive in the FST. Opposite results were obtained with UFP-113 and [F/G]N/OFQ(1-13)NH 2 . Conclusions and implications NOP ligands producing similar effects on NOP/G protein interaction (partial agonism) but showing different effects on β-arrestin 2 recruitment (partial agonism vs antagonism) elicited different actions on anxiety and mood. These results suggest that the action of a NOP ligand on emotional states is better predicted based on its β-arrestin 2 rather than G-protein efficacy. [ABSTRACT FROM AUTHOR]

Background. Opioid tolerance is a limiting factor in chronic pain. Delta opioid peptide (DOP)(d) receptor antagonism has been shown to reduce tolerance. Here, the common clinical mu opioid peptide (MOP)(m) receptor agonist fentanyl has been linked to the DOP antagonist Dmt-Tic (2',6'-dimethyl-L-tyrosyl-1,2,3,4-tetrahydrisoquinoline-3-carboxylic acid) to create new bivalent compounds. Methods. Binding affinities of bivalents(#9, #10, #11, #12 and #13) were measured in Chinese hamster ovary (CHO) cells expressing recombinant human MOP, DOP, Kappa opioid peptide (KOP)(μ) and nociceptin/orphanin FQ opioid peptide (NOP) receptors. Functional studies, measuring GTPγ[35S] or β-arrestin recruitment, were performed in membranes or whole cells respectively expressing MOP and DOP. Results. The new bivalents bound to MOP(pKi :#9:7.31; #10:7.58; #11:7.91; #12:7.94; #13:8.03)and DOP (#9:8.03; #10:8.16; #11:8.17; #12:9.67; #13:9.71). In GTPγ[35S] functional assays, compounds #9(pEC50:6.74; intrinsic activity:0.05) #10(7.13;0.34) and #11(7.52;0.27) showed weak partial agonist activity at MOP. Compounds #12 and #13, with longer linkers, showed no functional activity at MOP. In antagonist assays at MOP, compounds #9 (pKb:6.87), #10(7.55) #11(7.81) #12(6.91) and #13(7.05) all reversed the effects of fentanyl. At DOP, all compounds showed antagonist affinity (#9:6.85; #10:8.06; #11:8.11; #12:9.42; #13:9.00), reversing the effects of DPDPE ([D-Pen2,5]enkephalin). In β-arrestin assays, compared with fentanyl (with response at maximum concentration (RMC):13.62), all compounds showed reduced ability to activate barrestin (#9 RMC:1.58; #10:2.72; #11:2.40; #12:1.29; #13:1.58). Compared with fentanyl, the intrinsic activity was: #9:0.12; #10:0.20; #11:0.18; #12:0.09 and #13:0.12. Conclusions. The addition of a linker between fentanyl and Dmt-Tic did not alter the ability to bind to MOP and DOP, how ever a substantial loss in MOP functional activity was apparent. This highlights the difficulty in multifunctional opioid development. [ABSTRACT FROM AUTHOR]

Highlights: [•] The in vitro and in vivo pharmacological profile of novel NPSR ligands was assessed. [•] In vitro QA1 and PI1 behaved as pure and potent NPSR antagonists. [•] In vitro QA1 was more potent than PI1. [•] In vivo in mice QA1 and PI1 were only poorly active. [ABSTRACT FROM AUTHOR]

Abstract: Neuropeptide S (NPS) produces several biological actions by activating a formerly orphan GPCR, now named NPS receptor (NPSR). It has been previously demonstrated that NPS stimulates murine leukocyte chemotaxis in vitro. In the present study we investigated the ability of NPS, in comparison with the proinflammatory peptide formyl-Met-Leu-Phe (fMLP), to stimulate human monocyte chemotaxis. At a concentration of 10−8 M fMLP significantly stimulated chemotaxis. NPS produced a concentration dependent chemotactic action over the concentration range 10−12 to 10−5 M. The NPSR antagonists [d-Cys( t Bu)5]NPS, [ t Bu-d-Gly5]NPS and SHA 68 were used to pharmacologically characterize NPS action. Monocyte chemoattractant effect of NPS, but not fMLP, was completely blocked by either peptide antagonists or SHA with the nonpeptide molecule being more potent. None of the NPSR antagonists modified per se random cell migration. Thus, the present study demonstrated that NPS is able to stimulate human monocyte chemotaxis and that this effect is entirely due to selective NPSR activation. [Copyright &y& Elsevier]

Abstract: Neuropeptide S (NPS) regulates various biological functions by selectively activating the NPS receptor (NPSR). Recently, the NPSR ligand [ t Bu-d-Gly5]NPS was generated and in vitro characterized as a pure antagonist at the mouse NPSR. In the present study the pharmacological profile of [ t Bu-d-Gly5]NPS has been investigated. [ t Bu-d-Gly5]NPS activity was evaluated in vitro in the calcium mobilization assay at the rat NPSR and in vivo in the locomotor activity and righting reflex tests in mice and in the elevated plus maze and defensive burying assays in rats. In vitro, [ t Bu-d-Gly5]NPS was inactive per se while it inhibited the calcium mobilization induced by 30nM NPS (pKB 7.42). In Schild analysis experiments [ t Bu-d-Gly5]NPS (0.1–10μM) produced a concentration-dependent rightward shift of the concentration–response curve to NPS, showing a pA2 value of 7.17. In mouse locomotor activity experiments, supraspinal injection of [ t Bu-d-Gly5]NPS (1–10nmol) dose dependently counteracted NPS (0.1nmol) stimulant effects. In the mouse righting reflex assay [ t Bu-d-Gly5]NPS (0.1–10nmol) fully prevented the arousal-promoting action of the natural peptide (0.1nmol). Finally, [ t Bu-d-Gly5]NPS (3–30nmol) was able to completely block NPS (1nmol) anxiolytic-like actions in rat elevated plus maze and defensive burying assays. Collectively, the present results demonstrated that [ t Bu-d-Gly5]NPS behaves both in vitro and in vivo as a pure and potent NPSR antagonist. This compound represents a novel and useful tool for investigating the pharmacology and neurobiology of the NPS/NPSR system. [Copyright &y& Elsevier]

Abstract: Neuropeptide S (NPS) is the endogenous ligand of a previously orphan receptor now named NPSR. In the brain NPS regulates several biological functions including anxiety, arousal, locomotion, food intake, learning and memory, pain and drug abuse. Mice lacking the NPSR gene (NPSR(−/−)) represent an useful tool to investigate the neurobiology of the NPS/NPSR system. NPSR(−/−) mice have been generated in a 129S6/SvEv genetic background. In the present study we generated CD-1 congenic NPSR(+/+) and NPSR(−/−) mice and investigated their phenotype and sensitivity to NPS in various behavioural assays. The phenotype analysis revealed no locomotor differences between NPSR(+/+) and NPSR(−/−) mice. The behaviour of NPSR(+/+) and NPSR(−/−) mice in the righting reflex test was superimposable. No differences were recorded between the two genotypes in the elevated plus maze, open field and stress-induced hyperthermia tests, with the exception of rearing behaviour that was reduced in knockout animals. Moreover the behaviour of NPSR(+/+) and NPSR(−/−) mice in the forced swimming, novel object recognition and formalin assays was similar. The stimulatory effects of NPS in the locomotor activity test and its anxiolytic-like actions in the elevated plus maze and open field assays were evident in NPSR(+/+) but not NPSR(−/−) animals. In conclusion, the present study indicates that the NPS/NPSR system does not tonically control locomotion, sensitivity to diazepam, anxiety, depressive-like behaviours, memory and pain transmission in mice. Furthermore our results clearly show that the product of the NPSR gene represents the mandatory protein for all the NPS biological effects so far described. [Copyright &y& Elsevier]

Abstract: Carbamoyl Phosphate Synthetase 1 deficiency (CPS1D) is a rare autosomal recessive urea cycle disorder, potentially leading to lethal hyperammonemia. Based on the age of onset, there are two distinct phenotypes: neonatal and late form. The CPS1 enzyme, located in the mitochondrial matrix of hepatocytes and epithelial cells of intestinal mucosa, is encoded by the CPS1 gene. At present more than 220 clear-cut genetic lesions leading to CPS1D have been reported. As most of them are private mutations diagnosis is complicated. Here we report an overview of the main clinical findings and biochemical and molecular data of 13 CPS1D Italian patients. In two of them, one with the neonatal form and one with the late form, cadaveric auxiliary liver transplant was performed. Mutation analysis in these patients identified 17 genetic lesions, 9 of which were new confirming their “private” nature. Seven of the newly identified mutations were missense/nonsense changes. In order to study their protein level effects, we performed an in silico analysis whose results indicate that the amino acid substitutions occur at evolutionary conserved positions and affect residues necessary for enzyme stability or function. [Copyright &y& Elsevier]

Abstract: Fabry Disease (FD) is an X-linked multisystemic lysosomal disorder caused by mutations of α-galactosidase (GLA) gene. Only a few of the 450 genetic lesions identified so far have been characterised by in vitro expression studies. Thus the significance of newly identified GLA nucleotide variants in FD patients which lead to α-galactosidase (GAL-A) amino acid substitutions or intronic changes can be uncertain. We identified three GLA mutations, c.155G>A (p.C52Y), c.548G>C (p.G183A), c.647A>G (p.Y216C) in as many individuals (two male; one female) and performed in vitro expression studies and Western blot analysis in order to clarify their functional effects. Reduced GAL-A activity and normal or partially reduced mutant proteins were present in all overexpressed mutant systems in which three-dimensional structural analysis showed that the active site was not directly involved. We hypothesize that the three new mutations affect the GAL-A protein, leading to conformational FD. When mutant proteins overexpressed in COS-1 cells and in patients'' lymphocytes were tested in the presence of the 1-deoxygalactonojirimicin (DGJ) chaperone, the p.G183A and p.Y216C systems showed increased GAL-A enzyme activities and protein stabilisation while p.C52Y was not responsive. We underline that genetic, biochemical and functional studies are helpful in clarifying the consequences of the missense genetic lesions detected in FD. ERT is the elective therapy for Fabry patients, but it is not always possible to issue the enzyme''s active form in all involved organs. Our study endorses the hypothesis that an active site-specific chemical chaperone, which could be administered orally, might be effective in treating GAL-A conformational defects. [Copyright &y& Elsevier]

Abstract: Neuropeptide S (NPS) and its receptor NPSR comprise a recently deorphaned G-protein-coupled receptor system. There is a body of evidence suggesting the involvement of NPS in wakefulness, anxiety, locomotor activity and oxidative stress damage. Considering that mood stabilizers block the stimulatory effect of psychostimulants in rodents, the present study aimed to investigate the effects of the pretreatment with lithium and valproate on the hyperlocomotion evoked by NPS. Another relevant action induced by lithium and valproate is the neuroprotection against oxidative stress. Thus, aiming to get further information about the mechanisms of action of NPS, herein we evaluated the effects of NPS, lithium and valproate, and the combination of them on oxidative stress damage. Behavioral studies revealed that the pretreatment with lithium (100mg/kg, i.p.) and valproate (200mg/kg, i.p.) prevented hyperlocomotion evoked by NPS 0.1nmol. Importantly, the dose of valproate used in this study reduced mouse locomotion, although it did not reach the statistical significance. Biochemical analyses showed that lithium attenuated thiobarbituric reactive species (TBARS) formation in the striatum, cerebellum and hippocampus. NPS per se reduced TBARS levels only in the hippocampus. Valproate did not significantly affect TBARS levels in the brain. However, the combination of mood stabilizers and NPS blocked, instead of potentiate, the neuroprotective effects of each one. No relevant alterations were observed in carbonylated proteins after all treatments. Altogether, the present findings suggested that mainly the mood stabilizer lithium evoked antagonistic effects on the mediation of hyperlocomotion and protection against lipid peroxidation induced by NPS. [Copyright &y& Elsevier]

Abstract: Neuropeptide S (NPS) is a recently discovered peptide which induces hyperlocomotion, anxiolysis and wakefulness. This study aimed to compare behavioral and biochemical effects of NPS with amphetamine (AMPH), and diazepam (DZP). To this aim, the effects of NPS (0.01, 0.1 and 1 nmol, ICV), AMPH (2 mg/kg, IP) and DZP (1 mg/kg, IP) on locomotion and oxidative stress parameters were assessed in mouse brain structures. The administration of NPS and AMPH, but not DZP, increased locomotion compared to control. Biochemical analyses revealed that AMPH increased carbonylated proteins in striatum, but did not alter lipid peroxidation. DZP increased lipid peroxidation in the cortex and cerebellum, and increased protein carbonyl formation in the striatum. In contrast, NPS reduced carbonylated protein in the cerebellum and striatum, and also lipid peroxidation in the cortex. Additionally, the treatment with AMPH increased superoxide dismutase (SOD) activity in the striatum, while it did not affect catalase (CAT) activity. DZP did not alter SOD and CAT activity. NPS inhibited the increase of SOD activity in the cortex and cerebellum, but little influenced CAT activity. Altogether, this is the first evidence of a putative role of NPS in oxidative stress and brain injury. [Copyright &y& Elsevier]

Abstract: Objective: To examine the reorganisation of the somatosensory system after early brain lesions. Methods: We studied 12 young patients with congenital hemiplegia. Causative lesions were brain malformations, periventricular injuries and cortico-subcortical lesions. We explored the somatosensory system using evoked potentials, fMRI during sensory stimulation and clinical assessment of sensory function. To correlate sensory and motor function, we also performed transcranial magnetic stimulation, fMRI of hand movement and assessment of motor function by means of Melbourne test. Results: Eleven patients showed a perilesional reorganisation of primary somatosensory function, as expressed by short latency potentials following stimulation of the paretic hand; in a remaining patient, delayed latency responses (N27.1) were only elicited over the ipsilateral undamaged hemisphere. Five of the eleven patients with perilesional somatosensory representation of the affected hand showed contralesional shifting of motor function, thus exhibiting sensory-motor dissociation. Significant correlation was found between sensory deficit and fMRI activation during sensory stimulation. Conclusions: In subjects with early brain lesions, somato-sensory function is generally reorganised within the affected hemisphere. A contralesional shifting is uncommon and poorly efficient in function restoration. Significance: This study confirms and further explores the difference in reorganisation capabilities of the motor and sensory system following early brain injury of different etiologies and timing. [Copyright &y& Elsevier]

Abstract: Nociceptin/orphanin FQ (N/OFQ) produces several biological actions by activating the N/OFQ peptide receptor (NOP). It has been previously shown that N/OFQ stimulates leukocyte chemotaxis both in vitro and in vivo. In the present study we investigated the ability of N/OFQ, in comparison with the proinflammatory peptide formyl-Met-Leu-Phe (fMLP), to stimulate human neutrophil and monocyte chemotaxis and the release of lysozyme and superoxide anion (O2 −) production from neutrophils. fMLP stimulated all the leukocyte functions examined. N/OFQ stimulated monocyte (pEC50 12.15) but not neutrophil chemotaxis. The production of O2 − from neutrophils was not affected by N/OFQ while the release of lysozyme was increased in a concentration dependent manner (pEC50 11.00) although the maximal effects evoked by N/OFQ were about half of those of fMLP. The NOP ligands [Arg14, Lys15]N/OFQ, N/OFQ(1-13)NH2, Ro 64-6198, UFP-101 and the opioid antagonist naloxone were used for pharmacologically characterizing the receptor involved in the monocyte chemoattractant action of N/OFQ. [Arg14, Lys15]N/OFQ, N/OFQ(1-13)NH2, and Ro 64-6198 mimicked the action of N/OFQ showing similar maximal effects and the following order of potency: [Arg14, Lys15]N/OFQ (pEC50 13.22)>Ro 64-6198 (pEC50 12.96)>N/OFQ(1-13)NH2 (pEC50 12.67)>N/OFQ (pEC50 12.15). Moreover, the monocyte chemoattractant action of N/OFQ was not modified by naloxone 1μM while antagonized by UFP-101 10μM (pA2 7.00). Thus, the order of potency of agonists and the antagonist selectivity demonstrated that N/OFQ stimulates human monocyte chemotaxis via NOP receptor activation. [Copyright &y& Elsevier]

The neuropeptide nociceptin/orphanin FQ (N/OFQ) is the endogenous ligand for the nociceptin receptor (NOP). In an attempt to identify high potency NOP agonists for use in the brain we have compared the activity of a novel N/OFQ analogue [Phe1Ψ(CH2-O)Gly2]N/OFQ(1-13)NH2 ([F/G-O]) with the existing [Phe1Ψ(CH2-NH)Gly2]N/OFQ(1-13)NH2 ([F/G]). Both peptides are modified between the first two N-terminal amino acids and are further compared with the agonist template N/OFQ(1-13)NH2 in [3H]N/OFQ binding, GTPγ[35S] binding and cAMP inhibition studies using Chinese hamster ovary cells expressing the recombinant human NOP. All peptides displaced [3H]N/OFQ, stimulated GTPγ[35S] binding and inhibited cAMP formation. In [3H]N/OFQ binding and GTPγ[35S] binding the rank order affinity and potency was N/OFQ(1-13)NH2>[F/G-O]>[F/G]. In GTPγ[35S] binding [F/G] was a clear partial agonist with intrinsic activity (Emax stimulation factor, mean±SEM, n=4) of 7.75±1.02 compared with N/OFQ(1-13)NH2 of 11.13±1.76. The efficacy of [F/G-O] (10.17±1.88) approached that of the full agonist N/OFQ(1-13)NH2. Downstream, at the level of cAMP formation, all peptides were full agonists with the following rank order potency: N/OFQ(1-13)NH2>[F/G-O]=[F/G]. The enhanced potency and intrinsic activity of the novel [F/G-O] modification makes this an interesting peptide for further in vivo analysis. [Copyright &y& Elsevier]

Recently, the cloning of a novel preprotachykinin gene (PPT-C) has been reported. This gene codes for a novel peptide named hemokinin 1 (HK-1). In contrast with the known tachykinins, which are exclusively expressed in neuronal tissues, PPT-C mRNA was detected primarily in hematopoietic cells. In this study, we pharmacologically characterised the effects of HK-1 using three tachykinin monoreceptor systems, namely the rabbit jugular vein (rbJV) for NK1, the rabbit pulmonary artery (rbPA) for NK2, and rat portal vein (rPV) for NK3 receptors. In all these preparations substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) elicited concentration dependent contractions showing similar maximal effects and the following rank order of potency: SP > NKA = NKB in the rbJV, NKA > NKB ≫ SP in the rbPA, and NKB > NKA > SP in the rPV. In those vessels HK-1 behaved as a full agonist displaying potencies similar (rbPA and rPV) or slightly higher (rbJV) than those of SP. In the rbJV, SR 140333, a selective NK1 receptor antagonist, antagonised the effects of HK-1 and SP with similar high potencies (pKB 9.3 and 9.5, respectively). Similar results were obtained with the pseudopeptide NK1 antagonist, MEN 11467 (pKB 8.8 and 8.6, respectively). Taken together, these data indicate that HK-1 behaves as a NK1 preferring receptor agonist. [Copyright &y& Elsevier]

Abstract: We describe a case of hemiconvulsion–hemiplegia–epilepsy (HHE) syndrome documented by longitudinal magnetic resonance imaging (MRI). A two-year and nine-month-old boy had a prolonged hemiconvulsion during fever followed by right hemiparesis. Seven days later the imaging abnormality on T2 and diffusion-weighted images (DWI) was limited to the white matter of the left hemisphere. One month later severe gliosis and unilateral brain atrophy were already evident. MRI is useful in the early stages of prolonged seizures and T2 and DWI abnormalities appear to be well correlated with parenchymal damage that results from sustained ictal activity. The neuroradiological findings in our case and in the few HHE patients reported in the literature seem to be very characteristic and, if confirmed in larger series, could permit an early diagnosis. [Copyright &y& Elsevier]

Morphine is a gold standard analgesic acting at μ-opioid receptors (MOP) with analgesia accompanied by side-effects, for example, tolerance. If δ-opioid receptors (DOP) are blocked at the same time that MOP is activated, then analgesia with reduced tolerance results. As part of an ongoing programme to design mixed MOP agonist/DOP antagonist drugs (bifunctional) we have characterized the prototype pseudopeptide H-Dmt-Tic-Gly-NH-CH2-Ph (UFP505). We have measured ability to interact with MOP and DOP receptors with differential agonism/antagonism and the effects of 1 h pretreatment on cell surface MOP receptor numbers. We have used Chinese Hamster Ovary cells expressing human recombinant MOP, DOP KOP (κ), and NOP (nociceptin/orphanin FQ) receptors (CHOhMOP/hDOP/hKOP/ hNOP). Receptor binding was measured using the radioligand [3]diprenorphine ([3]DPN) in saturation format to determine receptor density (Bmax) and affinity (pKD) or displacement format to determine the affinity (pKi) of UFP505. Functional activity at MOP and DOP was assessed by measuring agonist stimulated GTPγ[35S] binding. Loss of cell surface receptors was measured using [3]DPN in saturation format after 1 h treatment of CHOhMOP with 10 µM UFP505. UFP505 displaced [3]DPN binding to MOP and DOP receptor with pKi 7.79 (0.18) and 9.82 (0.06) [mean (SEM), n=5], respectively. Affinity at KOP and NOP was negligible. At MOP in a GTPγ[35S] binding assay, UFP505 behaved as a full agonist (compared with MOP agonist endomorphin-1) with a pEC50 (potency) and Emax (efficacy:stimulation factor) of 6.23 (0.15) and 2.24 (0.15) (n=3), respectively. At DOP, UFP505 (10 µM) reversed the effects of the DOP agonist DPDPE in the GTPγ[35S] assay with potency (pKB) of 9.81 (0.18) (n=3), in agreement with pKi determined in [3]DPN binding assays. Pretreatment of CHOhMOP cells with 10 µM UFP505 for 1 h produced substantial loss of cell surface receptors with no change in DPN pKD. Bmax and pKD values for control and 1 h UFP505 treated cells were 332 (53) fmol mg-1 protein and 9.20 (0.11) and 155 (12) fmol mg-1 protein and 9.15 (0.10), respectively (n=5). Here we show that UFP505 is a bifunctional MOP agonist/ DOP antagonist capable of producing tolerance like actions in a single MOP expression system. The next stage is to examine the effects of this ligand in a double (MOP and DOP) expression system. [ABSTRACT FROM AUTHOR]