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This biographical note pays a small tribute to Robert Naquet, one of the pillars of French epileptology together with Henri Gastaut, his mentor. It can be said that the Marseilles school of epilepsy left an indelible mark in the neurological sciences and that there is no epileptologist, in France and abroad, who was not influenced by the teachings of these two gentlemen. It is practically impossible to separate the scientific achievements of Robert Naquet from his multiple facets as a humanist, artist, and teacher. His warm personality and legendary modesty may serve as an example for future generations of neuroscientists.

The development of treatments for acute neurodegenerative diseases (stroke and brain trauma) has focused on (i) re‐establishing blood flow to ischemic areas as quickly as possible (i.e. mainly antithrombotics or thrombolytics for stroke therapy) and (ii) on protecting neurons from cytotoxic events (i.e. neuroprotective therapies such as anti‐excitotoxic or anti‐inflammatory agents for stroke and neurotrauma therapies). This paper reviews the preclinical data for enoxaparin in in vivo models of ischemia and brain trauma in rats. Following a photothrombotic lesion in the rat, enoxaparin significantly reduced edema at 24 h after lesion when the treatment was started up to 18 h after insult. Enoxaparin was also tested after an ischemic insult using the transient middle cerebral artery occlusion (tMCAO) model in the rat. Enoxaparin, 2 × 1.5 mg/kg i.v., significantly reduced the lesion size and improved the neuroscore when the treatment was started up to 5 h after ischemia. Enoxaparin, administered at 5h after insult, reduced cortical lesion size in a dose‐dependent manner. In permanent MCAO, enoxaparin (5 and 24 h after insult) significantly reduced lesion size and improved neuroscore. A slight and reversible elevation of activated partial thromboplastin time (APTT) suggests that enoxaparin is neuroprotective at a non‐hemorrhagic dose. Traumatic brain injury (TBI) is often accompanied by secondary ischemia due in part to edema‐induced compression of blood vessels. When enoxaparin, at 0.5 mg/kg i.v. + 4 × 1 mg/kg s.c., was administered later than 30h after TBI, it significantly reduced edema in hippocampus and parietal cortex. At one week after TBI the lesion size was significantly reduced and the neurological deficit significantly improved in enoxaparin treated animals. Finally, the cognitive impairment was significantly improved by enoxaparin at 48 h to 2 weeks after TBI. The anticoagulant properties of unfractionated heparin and specifically enoxaparin can explain their anti‐ischemic effects in experimental models. Furthermore, unfractionated heparin and specifically enoxaparin, have, in addition to anticoagulant, many other pharmacological effects (i.e. reduction of intracellular Ca(2+) release; antioxidant effect; anti‐inflammatory or neurotrophic effects) that could act in synergy to explain the neuroprotective activity of enoxaparin in acute neurodegenerative diseases. Finally, we demonstrated, that in different in vivo models of acute neurodegenerative diseases, enoxaparin reduces brain edema and lesion size and improves motor and cognitive functional recovery with a large therapeutic window of opportunity (compatible with a clinical application). Taking into account these experimental data in models of ischemia and brain trauma, the clinical use of enoxaparin in acute neurodegenerative diseases warrants serious consideration.

Taking advantage of a progressive nonhuman primate model mimicking Parkinson's disease (PD) evolution, we monitored transcriptional fluctuations in the substantia nigra using Affymetrix microarrays in control (normal), saline-treated (normal), 6 days-treated (asymptomatic with 20% cell loss), 12 days-treated (asymptomatic with 40% cell loss) and 25 days-treated animals (fully parkinsonian with 85% cell loss). Two statistical methods were used to ascertain the regulation and real-time quantitative PCR was used to confirm their regulation. Surprisingly, the number of deregulated transcripts is limited at all time points and five clusters exhibiting different profiles were defined using a hierarchical clustering algorithm. Such profiles are likely to represent activation/deactivation of mechanisms of different nature. We briefly speculate about (i) the existence of yet unknown compensatory mechanisms is unraveled, (ii) the putative triggering of a developmental program in the mature brain in reaction to progressing degeneration and finally, (iii) the activation of mechanisms leading eventually to death in final stage. These data should help development of new therapeutic approaches either aimed at enhancing existing compensatory mechanisms or at protecting dopamine neurons.

Excessive release of glutamate, a potent excitatory neurotransmitter, is thought to play an important role in a variety of acute and chronic neurological disorders, suggesting that excitatory amino acid antagonists may have broad therapeutic potential in neurology. Here, we describe the synthesis, pharmacological properties and neuroprotective activity of 9-carboxymethyl-imidazo-[1-2a]indeno[1-2e]pyrazin-4-one-2-carboxylic acid (RPR117824), an original selective AMPA antagonist. RPR117824 can be obtained through a six-step synthesis starting from (1-oxo-indan-4-yl) acetic acid, which has been validated on a gram-scale with an overall yield of 25%. Monosodium or disodium salts of the compound exhibit excellent solubility in saline (> or = 10 g/L), enabling intravenous administration. RPR117824 displays nanomolar affinity (IC(50)=18 nM) for AMPA receptors and competitive inhibition of electrophysiological responses mediated by AMPA receptors heterologously expressed in Xenopus oocytes (K(B)=5 nM) and native receptors in rat brain slices (IC(50)=0.36 microM). In in vivo testing, RPR117824 behaves as a powerful blocker of convulsions induced in mice or rats by supramaximal electroshock or chemoconvulsive agents such as pentylenetetrazole, bicuculline, isoniazide, strychnine, 4-aminopyridine and harmaline with half maximal effective doses ranging from 1.5 to 10 mg/kg following subcutaneous or intraperitoneal administration. In disease models in rats and gerbils, RPR117824 possesses significant neuroprotective activity in global and focal cerebral ischemia, and brain and spinal cord trauma.

The purpose of this study was to set-up a battery of behavioral tests to assess sensorimotor and cognitive deficits following a moderate traumatic brain injury (TBI) in rats. Coordinated walking ability was evaluated in an accelerated rotarod test. Vestibulomotor function and fine motor coordination were assessed by using a beam-walking task. Rotarod and beam-walking performances were both altered in injured rats compared to sham-operated and control rats. A more pronounced and longer-lasting deficit was measured in the beam-walking test. Cognitive function was studied by using the Lashley maze paradigm. A spatial localization deficit was significant for 4 weeks posttrauma in TBI rats. The beam-walking task and the Lashley maze are robust and sensitive methods in detecting sensorimotor and cognitive impairment after TBI in rats, respectively. These tests are proposed for evaluating the ability of new pharmacological agents to improve the functional recovery after a TBI in rats.

An abnormal increase in the activity of neurons of the subthalamic nucleus is a key pathophysiological feature of Parkinson's disease. We sought to determine whether riluzole, a sodium channel inhibitor that interferes with glutamatergic neurotransmission, affects neuronal activity in this brain region. Intravenous administration of riluzole reduced the discharge rate of subthalamic neurons in rats with 6-OHDA-induced lesions of the midbrain. By contrast, no effect was observed in nonlesioned control animals. This property may contribute to the neuroprotective effects of riluzole in animal models of PD through the modulation of the glutamatergic inputs these neurons feedback to nigral dopaminergic neurons.

The overstimulation of excitatory amino acid receptors such as the glutamate AMPA receptor has been implicated in the physiopathogenesis of epilepsy as well as in acute and chronic neurodegenerative disorders. An original series of readily water soluble 4-oxo-10-substituted-imidazo[1,2- a ]indeno[1,2- e ]pyrazin-2-carboxylic acid derivatives was synthesized. The most potent derivative 6a exhibited nanomolar binding affinity (IC 50 =35 nM) and antagonist activity (IC 50 =6 nM) at ionotropic AMPA receptor. This compound also demonstrated potent anticonvulsant properties in MES in mice and rats with long durations of action with ED 50 values in the 1–3 mg/kg dose range following ip and iv administration.

A novel series of 2- and 9-disubstituted heterocyclic-fused 4-oxo-indeno[1,2- e ]pyrazin derivatives was synthesized. One of them, the 9-(1 H -tetrazol-5-ylmethyl)-4-oxo-5,10-dihydroimidazo[1,2- a ]indeno[1,2- e ]pyrazin-2-yl phosphonic acid 4i exhibited a strong and a selective binding affinity for the AMPA receptor (IC 50 =13 nM) and demonstrated potent antagonist activity (IC 50 =6 nM) at the ionotropic AMPA receptor. This compound also displayed good anticonvulsant properties against electrically-induced convulsions after ip and iv administration with ED 50 values between 0.8 and 1 mg/kg. Furthermore, a strong increase in potency was observed when given iv 3 h before test (ED 50 =3.5 instead of 25.6 mg/kg for the corresponding 9-carboxymethyl-2-carboxylic acid analogue). These data confirmed that there is an advantage in replacing the classical carboxy substituents by their bioisosteres such as tetrazole or phosphonic acid groups.

The over-stimulation of excitatory amino acid receptors such as the glutamate AMPA receptor has been suggested to be associated with neurodegenerative disorders. Here we describe an original series of readily water soluble 4-oxo-imidazo[1,2-a] indeno[1,2-e]pyrazin-8- and -9-carboxylic (acetic) acid derivatives. One of these compounds, 4f, exhibited nanomolar binding affinity, potent competitive antagonism at the ionotropic AMPA receptor and a long duration of anticonvulsant activity after administration by parenteral route in vivo.

Water soluble 8-methylureido-10-amino-10-methyl-imidazo[1,2-a]indeno[1,2-e]pyraz ine-4-one 4 represents a novel class of highly potent and selective AMPA receptors antagonists with in vivo activity. The dextrorotatory isomer (+)-4 was found to display the highest affinity with an IC50 of 10 nM. It also exhibited very good anticonvulsant effects after i.p., s.c. and i.v. administration in mice subjected to electrical convulsions (MES) and i.p. in audiogenic seizure-e in DBA/2 mice (ED50's < or = 10 mg/kg).

Indeno[1,2-b]pyrazin-2,3-diones have been identified as a novel series of potent ligands on the glycine site of the NMDA receptor. To improve their in vivo activities, an acetic acid-type side chain was introduced to the 5-position, giving water-soluble compounds when formulated as the sodium salt (>10 mg/mL). Introduction of a chlorine atom in the 8-position led to a dramatic improvement of anticonvulsant activity and this was surprising since this change did not improve binding affinity. A plausible explanation is a reduced recognition by a Na + ,K + -ATPase active transport system responsible for the excretion of these compounds from the brain and kidney. This promising new chemical series led to the optically active isomer (-)-10i (RPR 118723), a glycine/NMDA antagonist with nanomolar binding affinity and in vivo activity in animal model of convulsions and electrophysiology at doses in the range of 2-3 mg/kg following iv administration.

A novel series of 2-substituted-4,5-dihydro-4-oxo-4H-imidazo[1,2-a]indeno[1,2-e]pyrazine derivatives was synthesised. One of them, Figure 1 , Scheme 1 Download : Download high-res image (100KB) Download : Download full-size image Figure 1 . Download : Download high-res image (175KB) Download : Download full-size image Scheme 1 . Synthesis of compounds Figure 1 , Scheme 1 . Experimental conditions: (a) 7a: 6a, NaH, DMF, 15 °C, 45 min then 5a , CH2Cl2, rt, 48 h, flash chromatography on silica gel (CH2Cl2), 42%; 7b: 6a, 5b , K2CO3, acetone, reflux, 2 h, flash chromatography on silica gel (cyclohexane:EtOAc 30:70), 47%; (b) 6b , NaH, DMF, 15 °C, 45 min then 5a , CH2Cl2, rt, 48 h, flash chromatography on silica gel (cyclohexane:EtOAc 50:50), 49%; (c) Figure 1 , Scheme 1 : 7a, AcOH, NH4Ac, reflux, 2 h, 38%; Figure 1 , Scheme 1 : 7b, AcOH, NH4Ac, reflux, 2h, 16%; (d) 8a : 7a, MeOH, NH3 (gas), 30 min then rt, 48h, 70%; 8b : 7c, MeOH, NH3 (gas), 30 min then rt, 20 h, 80%; (e) Figure 1 , Scheme 1 , concd H2SO4, −5 °C to 0 °C, 30 min then potassium nitrate, 0 °C (30 min) to rt (12 h), 96% (f) H2 (pressure of hydrogen: 28 psi), cat.Pd/C (10%), DMF, rt, 24 h, 58%; (g) 6N HCl, reflux, 24 h, 59% (h) MeNCO, DMF, rt, 12 h, 46%; (i) 1N NaOH, dioxane, rt, 12 h then 1N HCl until pH=1 and precipitation, 67%; (j) AcOH, reflux, 7 h, 83%; (k) 6N HCl, reflux, 16 h, 39.5% (l) AcOH, reflux, 7h, 54%. —a highly water soluble compound—exhibited a nanomolar affinity and demonstrated competitive antagonist properties at the ionotropic AMPA receptors. This compound also displayed potent anticonvulsant properties against electrically or sound-induced convulsions in mice after systemic administration, thus suggesting adequate brain penetration.

The effect of riluzole administration, an antiglutamatergic compound, on the electrophysiological activity of the pallidal complex of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys is compared with those induced by two dosages of levodopa (L-DOPA), the first affording the best clinical alleviation, the second sufficient to induce dyskinesias. Both dosages of L-DOPA reduced sharply the firing frequency of globus pallidus pars internalis (GPi) neurons (respectively, 43.8+/-23.0 and 27.4+/-20.2 vs. 111. 2+/-31.4 Hz), decreased the percentage of bursting cells (respectively, 60.7 and 50.0 vs. 80.3%) and augmented the number of regular cells (respectively, 6.5 and 33.0 vs. 4.8%). Riluzole restored the firing frequency (75.0+/-26.9 Hz) and the firing pattern of the GPi (39.7% bursting, 9.5% regular and 50.8% irregular). These results suggest that the emergence of dyskinesia may well be due to a modification of the neuronal messages transmitted from the GPi to the motor nuclei of the thalamus. Riluzole would represent an interesting alternative to dopamine therapy in Parkinson's disease since it regularizes firing but does not cause dyskinesia.

The excessive release of glutamate, a potent excitatory neurotransmitter, is thought to play an important role in a variety of acute and chronic neurological disorders. Consequently, excitatory amino acid antagonists may have an important therapeutic potential in the treatment of these diseases. Glutamate interacts with at least three types of receptor: 1) NMDA (N-methyl-D-aspartic acid) receptors; 2) AMPA [2-amino-3-(3-hydroxy-5-methylisoxazol-4-yl)propionic acid]/kainic acid (KA) receptors; and 3) metabotropic receptors. Blockade of ionotropic AMPA/KA receptors has been shown to prevent cerebral ischemia insult in experimental models. This article describes the synthesis, pharmacological activity, and neuroprotective properties of 5H,10H-imidazo[1,2-a]indeno[1,2-e]pyrazine-4-one (1), a novel AMPA/KA antagonist which showed micromolar affinity at AMPA/KA receptors and competitively inhibited functional responses mediated by these receptors. In mice, 1 had significant anticonvulsive properties and conferred protection against hypobaric hypoxia and KCN intoxication. In rats and gerbils, 1 possesses significant activity in models of global or focal cerebral ischemia, as well as in a model of neurotrauma. Compound 1 was prepared from 2-bromo-indanone using two synthetic pathways in two or three steps with moderate (30%) or good (70%) yields, respectively. Drug Dev. Res. 48:121–129, 1999. © 1999 Wiley-Liss, Inc.

Original spiro-imidazo[1,2-a]indeno[1,2-e]pyrazine-4-one derivatives were synthesised and led to the identification of 3e which showed good affinities for both the AMPA and the NMDA glycine-site receptors, and displayed good anticonvulsant effects after i.p. and i.v. administrations in the electroshock-induced convulsion assay in mice. The corresponding dextrorotatory isomer (+)-3e was notably more potent than the levorotatory isomer (-)-3e in in vitro and in vivo assays.

1. The aim of this study was to investigate the effect of N-(3-(aminomethyl)benzyl)acetamidine (1400W), a selective inhibitor of inducible calcium-independent nitric oxide synthase (iNOS), on the functional and histopathological outcomes of experimental transient focal cerebral ischaemia in rats. 2. Transient ischaemia was produced by the occlusion for 2 h of both the left middle cerebral artery and common carotid artery. Treatments with 1400W (20 mg kg(-1)) or vehicle were started 18 h after occlusion of the arteries and consisted in seven subcutaneous injections at 8 h interval. Ischaemic outcomes and NOS activities (constitutive and calcium-independent NOS) were evaluated 3 days after ischaemia. 3. 1400W significantly reduced ischaemic lesion volume by 31%, and attenuated weight loss and neurological dysfunction. 4. 1400W attenuated the calcium-independent NOS activity in the infarct by 36% without affecting the constitutive NOS activity. 5. These findings suggest that iNOS activation contributes to tissue damage and that selective inhibitors of this isoform may be of interest for the treatment of stroke.

The aim of our study was to assess polymorphonuclear neutrophil infiltration into the injured parenchyma after a traumatic brain injury (TBI). Myeloperoxidase (MPO) activity was assayed on the hippocampus, temporal and parietal cortex 6, 24, 48, 72, and 120 h post-trauma. MPO activity occurred in these structures from 6 h post-trauma and was maximum at 24-48 h. It was resolved by 72 h in the hippocampus and the parietal cortex, but persisted in the temporal cortex until 120 h after trauma. This suggests that neutrophil infiltration is a delayed phenomenon in the physiopathology of TBI. Considering that a large therapeutic window may be crucial in the management of TBI, inhibition of neutrophil infiltration needs to be further investigated following cerebral trauma.

The neuroprotective effects of Riluzole, a compound with several mechanisms of action including the inhibition of sodium channel activity and glutamate release, were evaluated in a rat model of parasagittal fluid-percussion (FP) brain injury. Male Sprague-Dawley rats (350-400 g, n = 17) were anesthetized with sodium pentobarbital (60 mg/kg i.p.) and subjected to parasagittal FP brain injury of moderate severity (2.3-2.5 atm). Fifteen min following injury, animals randomly received an i.v. bolus of either Riluzole (8 mg/kg, n = 8) or vehicle (n = 9), followed by subcutaneous injections (identical dose) at 6 hr and 24 hr. Two weeks after injury and drug treatment, animals were sacrificed and a series of brain sections, stained with Hematoxylin and Eosin (H&E) or cresyl violet, were evaluated for quantitative cortical lesion volume and cell counts of hippocampal CA3 neurons, respectively, using a computerized image analysis system. Administration of Riluzole significantly reduced FP-induced tissue loss in the temporal/occipital cortices ipsilateral to the site of impact by 46%, compared to vehicle-treated, brain-injured animals (P = 0.01). In contrast, the selective neuronal loss observed in the CA3 region of the ipsilateral hippocampus was unaffected by Riluzole treatment. The present study demonstrates that Riluzole can attenuate cortical lesion size following brain trauma. These neuroprotective effects may be related to the synergy of the different mechanisms of action of Riluzole.

This study examined high affinity Na+-dependent uptake of glutamate in synaptosomal preparations from spinal cord in mice that express a dominant mutation of human copper/zinc superoxide dismutase (SOD1) and represent an animal model of amyotrophic lateral sclerosis (ALS). Their muscle strength was also monitored by a grip traction test throughout their lifespan. The high affinity Na+-dependent uptake of [3H]glutamate was decreased between 120 and 150 days of age. A marked and significant decrease in Vmax (-40.2%; p < 0.001) on whole spinal cord synaptosomes was observed at 150 days, with no change in Km. This significant decrease was reached a week before the animals died (157.2 +/- 2.2 days) and corresponded to a considerable fall in muscle strength (25% loss between 120 and 140 days, p < 0.001). The FALS mouse model therefore reproduces the decrease in glutamate uptake reported in humans suffering from sporadic or familial ALS. These results are discussed in terms of a possible tardive involvement of glutamate uptake deficiency in human ALS.

This study investigates the effect of enoxaparin (Lovenox, Klexane), a low-molecular-weight heparin, on edema following a photothrombotic lesion using rose bengal dye in the rat. An area of cerebral ischemia was provoked in the right hemisphere of rats. Edema developed over 24 h after the lesion, as seen comparing water content of a core sample from the right hemisphere to that of a similar sample from the left hemisphere of each rat. Enoxaparin at 0.5 mg/kg i.v. plus 2 mg/kg s.c. reduced edema 24 h after lesion induction by 32% (p < 0.01) when the treatment was started 2 h after photothrombotic insult, with maintenance doses of 2 mg/kg s.c. enoxaparin at 6 and 18 h. When the same initial treatment with enoxaparin was started 18 h after insult, there was still a significant reduction of 20% (p < 0.01) in cerebral edema. Administration of enoxaparin 18 h after insult reduced cerebral edema in a dose-dependent manner. There was no evidence of intracranial hemorrhages in any of the animal groups and when the hemoglobin content of the brain samples was assayed by the method of Drabkin, no increase in hemoglobin content was seen compared to sham-operated animals.

Brain trauma is the main cause of morbidity and mortality in young adults. One delayed events that occurs after a head trauma and compromises the survival of patients is cerebral edema. The present study examined first the occurrence of cerebral edema after a traumatic brain injury (TBI) induced by moderate fluid percussion in rats. Brain water content was measured from 1 h to 7 days posttrauma, in the hippocampus and cortex, on both ipsi- and contralateral hemispheres. Second, the effects of mannitol, an osmotic agent frequently used in the clinic, and riluzole, a neuroprotective compound, were investigated on regional edema formation. After TBI, the ipsilateral edema began early at 1-6 h, was maximal at 48 h and was resorbed by 5-7 days. No edema was observed in the contralateral hemisphere. Mannitol at 1 g/kg or vehicle was administered iv 15 min, 2 h and 4 h postinjury. At this dose, mannitol significantly attenuated the ipsilateral injured cortex edema measured at 6 h (p < 0.05). Riluzole at 4 and 8 mg/kg or vehicle was administered 15 min (IV) and 6 h, 24 h, and 30 h (SC) post-TBI. Riluzole at 4 x 4 mg/kg significantly reduced edema measured at 48 h, in the ipsilateral hippocampus (p < 0.05), whereas at 4 x 8 mg/kg, the reduction was observed in the hippocampus (p < 0.01) and the injured cortex (p < 0.05). Our results demonstrate that (1) cerebral edema begins early after the injury and is resorbed over 1 week; (2) mannitol could attenuate cerebral edema; and (iii) riluzole in addition to its neuroprotective effects reduces the brain edema. Thus, riluzole could be useful in human TBI treatment.

The aim of this study was to investigate the effects of riluzole on the lesion induced by a permanent middle cerebral artery occlusion (MCAO) in rats. Riluzole at 4 or 8 mg/kg i.v. significantly reduced the cortical ischemic brain damage. With the most effective dose of 8 mg/kg, the time evolution of the lesion was assessed by T2-weighted magnetic resonance imaging (MRI) repeated on the same animals after MCAO. MRI obtained at 24, 48, and 72 hours after MCAO showed a progressive increase of the ischemic lesion, except in the cortex of the riluzole-treated rats (8 mg/kg i.v.). Furthermore, there was no difference between lesion volumes as measured by MRI or by histology. This study indicates that MRI may be a valuable method to quantify in vivo the neuroprotective profile of a drug.

Riluzole has been shown recently to increase life expectancy in patients with amyotrophic lateral sclerosis. A number of experimental studies also suggest that this compound may be a neuroprotectant. We have investigated in baboons whether riluzole would protect striatal neurons from a prolonged 3-nitropropionic acid (3NP) treatment and ameliorate the associated motor symptoms. In animals receiving 3NP and the solvent of riluzole, 12 weeks of high-dose 3NP treatment resulted in the appearance of persistent leg dystonia and significant increases in the incidence of three categories of abnormal movements and in the dyskinesia index in the apomorphine test (0.5 mg/kg i.m.). Quantitative assessment of these behavioral deficits using a video movement analysis system demonstrated a significant decrease in locomotor activity and peak tangential velocity in 3NP-treated animals compared to controls. Histological analysis showed the presence of severe, bilateral, striatal lesions, localized in both caudate and putamen. Cotreatment with riluzole (4 mg/kg i.p., twice daily) significantly reduced the dyskinesia index (-35%, P0.02) in the apomorphine test. In the quantitative behavioral analysis, riluzole significantly ameliorated the decrease in peak tangential velocity (P0.02) but not the decrease in locomotor activity observed after 3NP. Comparative histological analysis of the two groups of treated animals did not demonstrate a clear neuroprotective effect of riluzole. The present study suggests that one potential therapeutic interest for riluzole in neurodegenerative disorders may reside in the reduction of motor symptoms associated with striatal lesions.

In the present study we have assessed whether riluzole stimulates the production of trophic activities for motoneurons by spinal astrocyte cultures. Astrocyte monolayers prepared from new-born rats were exposed to vehicle or riluzole (1-10 microM) for 30-36 h, then washed and further incubated without riluzole for 24 h in L15 medium to obtain the astrocyte conditioned media (ACM). Motoneuron-enriched cultures were used to test the ability of the ACM to support motoneuron viability. Astrocyte monolayers exposed to 1 microM riluzole did not show changes in morphology or in DNA or protein synthesis. However, the conditioned medium obtained from astrocyte monolayers after this treatment increased motoneuron survival compared to that from vehicle-treated cultures. A similar effect was found when astrocytes were exposed to a higher riluzole concentration (10 microM) but with greater dilutions of the conditioned medium. This trophic activity was abolished by boiling or after treatment with trypsin. These findings strongly suggest the existence of a new trophic mechanism, through which riluzole may exert motoneuron protection.

This study examined the influence of zopiclone, a third generation hypnotic, on the transition from slow wave sleep to paradoxical sleep (PS) which is increased at the expense of PS by barbiturates and benzodiazepines. The compound decreased sleep latency and increased the latency of the intermediate stage (IS) and PS at 2.5, 5 and 7.5 mg/kg IP. The amount of the IS was decreased because of the decrease in phase number up to 6 h at all doses. PS amount was decreased during 2 h at 2.5 mg/kg and during 4 h at 5 and 7.5 mg/kg also because of the decrease in phase number. The IS never substituted for PS. The IS spindle characteristics were not modified and the theta rhythm frequency slightly decreased at 5 mg/kg (IS) and 7.5 mg/kg (PS).

Several potential mechanisms are involved in mediating the pathophysiology of traumatic brain injury (TBI), including inflammatory processes and excitotoxicity. In the present study, we evaluated the ability of the use-dependent sodium channel inhibitor Riluzole to attenuate cognitive and neurologic motor deficits and reduce regional cerebral edema and histologic cell damage following lateral fluid-percussion (FP) brain injury in rats (n = 109). In study 1, 58 anesthetized male Sprague-Dawley rats (350-400 g) were subjected to FP brain injury of moderate severity (2.3-2.5 atm). Fifteen minutes following brain injury, animals randomly received an i.v. bolus of either Riluzole (4 mg/kg, n = 11), Riluzole (8 mg/kg, n = 11), or glycol vehicle (n = 20), followed by 6 h and 24 h s.c. injections (identical dose). Surgically prepared but uninjured animals received vehicle (n = 16) and served as controls. Animals were evaluated for cognitive deficits at 48 h postinjury and killed for assessment of regional brain edema. Administration of vehicle or Riluzole (4 mg/kg x 3) had no significant effect on memory or edema, whereas Riluzole (8 mg/kg x 3) significantly attenuated post-traumatic cognitive dysfunction (p0.05). In study 2, a second group of animals (n = 25) was injured, treated with Riluzole (8 mg/kg x 3 doses, n = 13) or vehicle (n = 12), and evaluated for neurologic motor function over 2 weeks. Animals treated with Riluzole demonstrated significantly improved motor scores beginning 1 week postinjury (p0.05). In study 3, brain-injured animals were treated with Riluzole (8 mg/kg x 3 doses, n = 10) or vehicle (n = 10), and posttraumatic lesion volume was assessed at 48 h postinjury using 2,3,5-triphenyltetrazolium chloride (TTC) staining. Treatment with Riluzole had no significant effect on posttraumatic lesion volume. The present study demonstrates that use-dependent sodium channel inhibitors, such as Riluzole, can attenuate both cognitive and neuromotor dysfunction associated with brain trauma.

THIS study evaluated treatment with riluzole, a neuroprotective agent, following thoracic spinal cord compression in the rat. Animals received riluzole (2 mg kg -1 ) or vehicle twice daily for 10 days following the trauma. Motor deficits, somatosensory evoked potentials (SEP) and lesion histology were evaluated. Although paralysis was seen following trauma, seven of 10 animals receiving riluzole recovered motor function and nearly normal behaviour, unlike animals receiving vehicle. Trauma dramatically disturbed SEPs with falls in amplitude and increases in latency. After riluzole SEP returned towards pre-injury levels, while untreated animals showed no recovery. Morphological studies revealed significant (53%) reduction in the degree of spinal cord infarcted after riluzole treatment.

1. 1. The authors describe here the effects of intravenous administration of RP 60180, a novel kappa agonist, on conscious baboons of the species Papio papio , which spontaneously present photically induced epileptic responses. 2. 2. Animals (n = 2) were chronically implanted with epidural recording electrodes and tested whilst seated in a primate chair. The electrocorticogram (ECoG) and electrocardiogram (ECG) were recorded during a control period of at least 30 minutes before the injection of RP 60180 (1 to 4.5 mg/kg i.v.) and immediately afterwards. 3. 3. Qualitatively, up to the dose of 4.5 mg/kg i.v., RP 60180 did not modify ECoG backgroundin term of paroxysmal activity in comparison with that observed during the control period. It did not cause any manifest focal or generalized seizure discharges, nor did it consistently enhance or reduce photically induced myoclonic responses. 4. 4. From the dose of 1 mg/kg i.v., RP 60180 slowed ECG frequency. This effect, which lasted for about 30 minutes post-injection, was most often seen at the higher doses. 5. 5. In another set of experiments, one baboon received the kappa agonist U-50488 (a benzacetamide derivative of spiradoline) at 1 and 3 mg/kg i.v. U-50488, at 3 and to a lesser degree at 1 mg/kg i.v., induced paroxysmal bursts of slow wave ECoG activity and a slowing of the ECG. These effects lasted about 1 hour post-drug administration. During this period, we observed spontaneous vocalization, as if the animal were complaining, as well as shaking.

Excitatory amino acid-mediated neurotoxicity was investigated in motoneuron-enriched cultures from fetal rats at 12-14 days of gestation. The cultures were mainly composed of differentiated motoneurons identified by choline acetyl transferase and calcitonin gene-related peptide (CGRP) immunoreactivity. Addition of glutamate (600 microM) to the conditioned medium induced no acute neuronal swelling. However, it was followed by a widespread neuronal degeneration over the next 24 h, accounting for 77% of the total cell number. Glutamate toxicity was dose dependent, with an EC50 around 300 microM. Treatment for 24 h with the agonists, N-methyl-D-aspartate (NMDA, 100 microM), kainate (500 microM) or RS-alpha-amino-3-hydroxy-5-methyl-4-isoxalopropionate (AMPA, 10 microM), also induced a significant cell loss. Riluzole (2 amino 6-trifluoromethoxybenzothiazole), a compound known to interfere with glutamatergic transmission pre- and postsynaptically, significantly reduced glutamate and NMDA neurotoxicity in a dose-dependent manner. These results suggest that a prolonged activation of one or more subtypes of ionotropic excitatory amino acid receptors can lead to motoneuron degeneration in vitro, and provide direct experimental evidence supporting the neuroprotective effect of riluzole in cultured motoneurons.

Optical isomers of the riluzole derivative, sulfoxide RP 64406, were prepared in pure forms. No significant difference was found between the (+) and (−) isomers in their powerful antiglutamate activity.

Novel strategies are needed to treat epilepsy, in order to ensure efficiency, security and prevention. The search for innovating anti-epileptics is based on finding appropriate target molecules, among which the most pertinent appear to be chlore and potassium channels. Transcriptomics and proteomics are also prone to detect genes or proteins implicated in the disease, in particular when biopsies from healthy and epileptic brains are compared. Animal genetic models provide information about epilepsies with a unique origin. Finally some targets are identified through fortuitous findings from research in other fields, notably that of pro-inflammatory cytokines.

Nitric oxide production in the cerebellum and induction of long-term potentiation (LTP) in the hippocampus have some characteristics in common: both phenomena are induced by activation of N-methyl-d-aspartate receptors and both are highly dependent on calcium-mediated processes. Here we provide evidence that endogenous nitric oxide production is necessary for synaptic plasticity in the CA1 hippocampus of the rat. LTP recorded in slices was blocked in a concentration-dependent manner by the nitric oxide synthase inhibitors l-NG-nitroarginine and l-NG-nitroarginine methyl ester, but l-NG-monomethylarginine was only marginally active. Bathing the slices with haemoglobin, a protein that scavenges nitric oxide, also resulted in a concentration-dependent blockade of LTP. Nitric oxide released locally from hydroxylamine produced a stable potentiation of synaptic transmission that was not additive with LTP induced by high-frequency stimulation. These results are fully consistent with the presumed retrograde messenger role of nitric oxide in LTP.

The excitatory neurotransmitter glutamate interacts with ionotropic and metabotropic receptors that mediate a variety of normal signalling processes in the brain. However, excessive stimulation of these receptors appears to be involved in neurodegenerative processes, at least in animal models. Ionotropic glutamate receptors can be divided into NMDA and non-NMDA (AMPA and KA) subtypes on the basis of t heir preferential affinities for the synthetic excitatory aminoacids N-methyl-D-aspartic acid (NMDA) or 2-amino-3- (3-hydroxy-5-methylisoxazol-4-yl) propionic acid (AMPA), respectively. Although most of the early evidence favoured a role for NMDA receptors in the excitotoxic processes, it has been recognised that AMPA receptors may also be significantly involved in neuronal death. As a consequence, the synthesis of specific AMPA antagonists has raised much interest as source of potential drugs for epilepsy and acute and chronic neurodegenerative diseases. The discovery of RPR117824, a potent and selective AMPA receptors antagonist endowed with anticonvulsant and neuroprotective properties, induced growing interest on dihydro-4-oxo-4Himidazo[ 1,2-a]indeno[1,2-e]pyrazine series. This review covers the main chemical course leading to the most promising compounds as well as the pharmacological properties of this original class of AMPA receptor antagonists.

Traumatic brain injury (TBI) is often accompanied by secondary ischemia due, in part, to edema-induced blood vessel compression. Enoxaparin, a low-molecular weight heparin, which is efficacious in models of myocardial and brain ischemia was studied in lateral fluid percussion-induced TBI in rats. Enoxaparin was administered 2 h post-TBI at 0.5 mg/kg i.v. followed by 4 x 0.5, 4 x 1, or 4 x 2 mg/kg s.c. over 30 h. Brain edema was measured in the hippocampus, temporal cortex and parietal cortex. Edema was reduced by enoxaparin (0.5 + 4 x 0.5 mg/kg) in the hippocampus (-53%, p = 0.07) and the parietal cortex (-39%, ns). At 0.5 + 4 x 1 mg/kg edema was reduced in the hippocampus (-63%, p < 0.05) and the parietal cortex (-47%, p = 0.06). At 0.5 + 4 x 2 mg/kg, the reduction was more important in the hippocampus (-69%, p < 0.01) and in the parietal cortex (-50%, p < 0.05). No reduction was seen in the temporal cortex. The lesion size was reduced by enoxaparin at 0.5 + 4 x 1 mg/kg (-50%, p < 0.05), and at 0.5 + 4 x 2 mg/kg (-35%, ns). The neurological deficit evaluated with a 9-point scale was also improved with enoxaparin at 0.5 + 4 x 1 mg/kg 1 week post-TBI (p < 0.05). The cognitive impairment evaluated with a Lashley maze task was improved with enoxaparin (0.5 + 4 x 1 mg/kg) from 48 h (p < 0.05) to 2 weeks post-TBI (p < 0.01). Our results demonstrate for the first time that enoxaparin significantly reduces the brain contusion and edema, and improves the functional outcomes induced by a TBI. Therefore, enoxaparin could be a candidate drug to treat acute brain-injured patients.

The neuroprotective drug riluzole (Rilutek®) is a sodium channel blocker and anti-excitotoxic drug which is marketed for the treatment of amyotrophic lateral sclerosis (ALS). Previous studies have shown that riluzole prolongs survival of transgenic mice harboring the mutated form of Cu,Zn-superoxide dismutase found in familial forms of the human disease. In this study we have examined the effect of treatment with riluzole in mice suffering from progressive motor neuronopathy (pmn), a hereditary autosomal recessive wasting disease which shares some symptoms of ALS. These mutants display hind limb weakness starting during the 3rd week of life and leading to paralysis and death during the 7th week of life. Daily treatment with 8 mg/kg of riluzole by oral route significantly retarded the appearance of paralysis, increased life span and improved motor performance on grip test and electromyographic results in the early stage of the disease. There was no effect of riluzole on weight gain. These data demonstrate that riluzole significantly prolongs life span, retards the onset of paralysis and slows the evolution of functional parameters connected with muscle strength in the pmn mouse model of motor neuron disease.

A novel series of readily water soluble 8-methylureido-4,5-dihydro-4-oxo-10 H -imidazo[1,2- a ]indeno[1,2- e ]pyrazines were synthesized. The -10-yl acetic acid ((+)- 3 Download high-res image (146KB) Download full-size image Scheme 3 . Synthesis of (±) -3, (+) -3, (−) -3 and 4 . Reagents and conditions: (a) HCOCO 2 H, NaH, DMF, rt then 1 N HCl, rt, 64%; (b) ZnCl 2 , Ac 2 O, reflux, 23%; (c) KNO 3 , concd H 2 SO 4 , rt, 92%; (d) MeOH, concd HCl, Fe, 65 °C, 83.5%; (e) MeNCO, K 2 CO 3 , DMF, 6 h, rt, 84.5%; (f) 1 N NaOH, 35 °C then 1 N HCl, 46%; (g) preparative HPLC (see text), (+) -14 : 26%, (−) -14 : 27%; (h) 8 N HCl, dioxane, 40 °C, (+) -3 : 67%, (−) -3 : 63.5%, (i) SnCl 2 , concd HCl, 40 °C, 94%; (j) MeNCO, K 2 CO 3 , DMF:dioxane 1:1, rt, 35%. ) and -10-carboxylidene ( 4 ) derivatives exhibit potent affinities (IC 50 =4 and 19 nM, respectively) and antagonist properties (IC 50 =2 and 3 nM, respectively) at the ionotropic AMPA receptor. These compounds also display anticonvulsant properties against both electrically and sound-induced convulsions in mice after ip, sc and iv administration with ED 50 values between 0.9 and 11 mg/kg, thus suggesting adequate brain penetration.

The effects of systemic treatment with the CCK-B receptor antagonist L-365,260, its 3S-(-) enantiomer and the CCK-A receptor antagonist devazepide were assessed in the plus-maze procedure in mice. L-365,260 (1-1000 micrograms.kg-1 i.p.) produced a dose-dependent increase in the percentage of entries into, and the percentage of time spent in the open arms. Total arm entry was not consistently modified. Neither the 3S-(-) enantiomer of L-365,260 nor devazepide (both administered at 100-10,000 micrograms.kg-1) enhanced the exploratory behavior of mice. These results suggest that CCK-B, rather than CCK-A antagonists may possess 'anxiolytic' properties in mice.

A cryogenic lesion was produced under halothane anaesthesia in the mouse by placing a cotton swab soaked in liquid nitrogen onto the surface of the cranium. This provoked an oedematous lesion which developed within the hour after the insult and evolved over the following week. Treatment with mannitol at 3 g/kg i.v. caused a significant 22% reduction in oedema 1 h later, when administered immediately after lesion, but not when administered 23-h post lesion. Likewise riluzole (16 mg/kg, i.v.) significantly reduced oedema by 17% when administered immediately after lesion, or 13% (P < 0.05) when administered 23 h after lesion. Repeated doses (2 x 16 mg/kg, i.p.) of riluzole were also able to reduce oedema significantly (24%, P < 0.05) at 24 h post lesion. Riluzole, in four repeated doses of 8 mg/kg i.p. was also able to reduce lesion surface size by 16% (P < 0.05) 48 h after lesion.

Two series of analogues of riluzole, a blocker of excitatory amino acid mediated neurotransmission, have been synthesized: monosubstituted 2-benzothiazolamines and 3-substituted derivatives. Of all the compounds prepared in the first series, only 2-benzothiazolamines bearing alkyl, polyfluoroalkyl, or polyfluoroalkoxy substituents in the 6-position showed potent anticonvulsant activity against administration of glutamic acid in rats. The most active compounds displaying in vivo "antiglutamate" activity were the 6-OCF(3) (riluzole), 6-OCF(2)CF(3), 6-CF(3), and 6-CF(2)CF(3) substituted derivatives with ED(50) values between 2.5 and 3.2 mg/kg i.p. Among the second series of variously substituted benzothiazolines, compounds as active as riluzole or up to 3 times more potent were identified in two series: benzothiazolines bearing a beta-dialkylaminoethyl moiety and compounds with an alkylthioalkyl chain and their corresponding sulfoxides and sulfones. The most potent derivatives were 2-imino-3-(2-methylthio)- and 2-imino-3-(2-methylsulfinyl)-ethyl-6-trifluoromethoxybenzothiazolines (61 and 64, ED(50) = 1.0 and 1.1 mg/kg i.p., respectively). In addition, intraperitoneal administration of some of the best benzothiazolines protected mice from mortality produced by hypobaric hypoxia.

Physical injury to the central nervous system (CNS) remains one of the main causes of mortality and disability in young adults. Numerous therapies have been successfully evaluated in experimental traumatic brain or spinal cord injuries (TBI, SCI) and, although some of them are currently under clinical trials for these indications, no drug therapy is at present available. Thus, an interesting approach to reduce the CNS injury-induced damage could be the blockade of Na+-channels by drugs such as riluzole which is neuroprotective in models of TBI or SCI as summarized in this review. Repeated doses ranging from 2 to 8mg/kg were administered between 24h to 10 days post-injury, with a first administration given either at 15 min or up to 6h post-injury. In these models riluzole was found to reduce both the size of spinal cord and brain lesions as well as brain edema, and to restore the neurological, motor and cognitive impairments consequent of these injuries. The largest therapeutic time window obtained was 1 to 6h in TBI. Thus such a compound should be considered as an interesting candidate for the treatment or SCI or TBI.

Preliminary studies have shown that riluzole, a Na + channel blocker with antiglutamatergic activity, has neuroprotective efficacy in several models of acute dopaminergic neurodegeneration. A chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) monkey model which comes closer to the slow evolution of Parkinson's disease has recently been developed in order to allow dynamic studies. The present results show that riluzole delayed the appearance of parkinsonian motor abnormalities in this dynamic model, using from 10.2±1.6 daily injections for the MPTP-treated monkeys ( n =4) to 16.5±2.0 daily injections for the MPTP+riluzole-treated monkeys ( n =4). These results strongly suggest that riluzole may be beneficial to slow down the rate of progression of Parkinson's disease.

The putative neuroprotective effect of riluzole was investigated in a rat model of progressive striatal neurodegeneration induced by prolonged treatment (three weeks, intraperitoneal) with 3-nitropropionic acid, an irreversible inhibitor of succinate dehydrogenase. Quantitative analysis of motor behaviour indicated a significant protective effect (60%) of riluzole (8 mg/kg/day) on 3-nitropropionic acid-induced motor deficits as assessed using two independent motor tests. Furthermore, quantitative analysis of 3-nitropropionic acid-induced lesions indicated a significant 84% decrease in the volume of striatal damage produced by 3-nitropropionic acid, the neuroprotective effect apparently being more pronounced in the posterior striatum and pallidum. In addition, it was checked that this neuroprotective effect of riluzole against systemic 3-nitropropionic acid did not result from a decreased bioavailability of the neurotoxin or a direct action of riluzole on 3-nitropropionic acid-induced inhibition of succinate dehydrogenase. We found that riluzole significantly decreased by 48% the size of striatal lesions produced by stereotaxic intrastriatal injection of malonate, a reversible succinate dehydrogenase inhibitor. Furthermore, the inhibition of cortical and striatal succinate dehydrogenase activity induced by systemic 3-nitropropionic acid was left unchanged by riluzole administration. The present results, consistent with a beneficial effect of riluzole in amyotrophic lateral sclerosis, suggest that this compound may be useful in the treatment of chronic neurodegenerative diseases.

Riluzole (2-amino 6-trifluoromethoxy-benzothiazole) was studied in a rat model of traumatic brain injury (TBI) induced by a fluid percussion applied laterally to the right parietal cortex. Study I: vehicle or riluzole (4 or 8 mg/kg) was administered 15 min (i.v.), 6 h and 24 h (s.c.), after TBI. Brain lesions were quantified 1 week after insult. Riluzole significantly reduced the size of TBI-induced lesions by approximately 44% with either dose regime (P < 0.05). Study II: vehicle or riluzole (8 mg/kg) was administered 15 min (i.v.), 6 h (i.p.) and then twice daily (i.p.) for 6 days, after injury. One, 2 and 3 weeks after TBI, a neurological examination was performed. Control injured rats had a significant neurological deficit at 1, 2 and 3 weeks (P < 0.001). Riluzole treatment did not modify the neurological status evaluated for the first 2 weeks after TBI. However at 3 weeks, riluzole significant improved the neurological function of injured rats (P < 0.05). These results suggest that riluzole may be beneficial in the clinical treatment of TBI. The protective action of riluzole may result from (i) stabilization of the inactivated state of voltage-dependent sodium channels, (ii) indirect action on the glutamatergic pathway, and/or (iii) indirect neurotrophic effect.