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The removal of lead from drinking water was investigated to develop a point-of-use water filter that could meet the regulation imposed by the new European Directive 98-83 lowering lead concentration in drinking water below 10 μg L −1 . The objective of this research was to assess the potential of different adsorbents (zeolites, resins, activated carbon, manganese oxides, cellulose powder) to remove lead from tap water with a very short contact time. To begin, the repartition of the lead species in a tap water and a mineral water was computed with the computer model CHESS. It showed that in bicarbonated waters lead is mainly under lead carbonate form, either in the aqueous or in the mineral phase. Batch experiments were then conducted to measure the equilibrium adsorption isotherms of the adsorbents. Then, for five of them, dynamic experiments in micro-columns were carried out to assess the outlet lead concentration level. Three adsorbents gave rise to a leakage concentration lower than 10 μg L −1 and were then selected for prototypes experiments: chabasite, an activated carbon coated with a synthetic zeolite and a natural manganese oxide. The proposed method clearly showed that the measurement of equilibrium isotherms is not sufficient to predict the effectiveness of an adsorbent, and must be coupled with dynamic experiments.

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 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).

Although several adaptive mechanisms have been identified that mask the existence of Parkinson's disease and delay the onset and aggravation of motor symptoms, the timescale and implications of this compensatory process remain an enigma. In order to examine: (i) the nature of the dopaminergic adaptive mechanisms that come into action; (ii) their sequential activation in relation to the severity of degeneration; and (iii) their efficacy with regard to the maintenance of a normal level of basal ganglia activity, we analysed the brains of mice treated daily with 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP, 4 mg/kg, i.p.) and killed at 5-day intervals from day 0 (D0) to D20. Our results demonstrate the sequential activation of two compensatory mechanisms: (i) an increase in striatal tyrosine hydroxylase (TH) protein content attested by the persistence of TH immunolabelling up to D15, contrasting with the decrease observed in both the number of nigral TH-immunoreactive neurons (-70.2%) and striatal dopamine content (-38.4%); (ii) a downregulation of DA uptake in surviving terminals at D20 (73.4% of nigral degeneration). At this point, the failure of adaptive mechanisms to maintain striatal dopaminergic homeostasis is also illustrated by an increase in the cytochrome oxidase activity of substantia nigra pars reticulata, a marker of neuronal function. It has been postulated that an increase in dopamine release per pulse could constitute an adaptive mechanism. The data we present from our MPTP mice model infirm this hypothesis. This study explores the link between the degree of nigral degeneration and the sequential activation of dopaminergic compensatory mechanisms in the nigrostriatal pathway and, in so doing, proposes a rethink of the paradigm applied to these mechanisms.

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.

Riluzole, has previously been shown to be protective in animal models of Parkinson's disease in vivo. In the present study the effects of riluzole on the intrastriatal formation and accumulation of MPP(+), after i.p. injection of MPTP were tested in mice, using two different experimental protocols. In the first protocol, mice were treated with a single dose (15 mg/kg i.p.) of MPTP and MPP(+) accumulation was measured 30 min, 1 h and 3 h after the injection of the toxin. Riluzole (10 mg/kg p.o.), administered 30 min before MPTP, did not modify the accumulation kinetic of MPP(+). Contrarily to riluzole, a single dose of 50 mg/kg p.o. of 7-nitroindazole (7-NI), a non-selective non hypertensive inhibitor of nitric oxide synthase (NOS), significantly decreased MPP(+) levels. In the second protocol, consisting of 3 injections of MPTP (15 mg/kg i.p.), riluzole, administered 4 times at the dose of 5 mg/kg p.o., had no effect on MPP(+) levels. The protective effect of repeated treatments of riluzole and 7-NI against MPTP-induced depletion of dopamine (DA) is also reported. Our data obtained with 7-NI (in agreement with previous studies reported by others) suggest that a part of the protection observed with this NOS inhibitor is probably due to in vivo inhibition of monoamine oxidase-B (MAO-B). That riluzole does not modify MPP(+) accumulation demonstrates that its protective effect against MPTP toxicity was not due to an in vivo interference with MPTP metabolism.

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 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.

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.

The effects of thalidomide, a sedative, anti-inflammatory and immunosuppressive agent were studied in the MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) murine model of Parkinson's disease. The striatal levels of dopamine (DA) and of its main metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were measured both in the MPTP control group (3 x 15 mg/kg intraperitoneally) and in the thalidomide groups (repeated treatments at 25 mg/kg or 50 mg/kg postoperatively). For mice treated with thalidomide, a dose-dependent protection was observed against the MPTP-induced decrease in DA. The decrease in HVA levels was totally antagonized by thalidomide at both doses. That thalidomide has activity in this model suggests that an inflammatory process may be involved in the induction of lesions by MPTP in DAergic neurons.

Metapramine, a pharmacological compound with antidepressant activity in humans, was tested for possible antiglutamatergic activity, in vitro. We investigated the effects of metapramine on the N-methyl-d-aspartic acid (NMDA) receptor complex, by determining whether this compound would interfere with the binding of [3H]N-[1-(2-thienyl)cyclohexyl]-3,4-piperidine ([3H]TCP) to rat cortical membranes in the presence of either glycine NMDA, or both. Metapramine in the micromolar range inhibited the binding of [3H]TCP in the presence of both NMDA and glycine (ic50 = 1.4 ± 0.2 μM). That very similar affinities were observed when either NMDA or glycine was present suggests that metapramine exerted a direct action at the PCP site. The affinity of metapramine for this site was about 25 and 350 times lower than that of PCP and MK-801, respectively. Metapramine inhibited the NMDA-evoked increase in guanosine 3′,5′-cyclic monophosphate (cGMP) levels of neonatal rat cerebellar slices (ic50 = 13 μM). These results suggest that metapramine is a low-affinity antagonist of the NMDA receptor complex channel. This paper discusses the potential application of metapramine to the treatment of diseases linked to excessive stimulation of glutamatergic NMDA receptors. © 1997 Elsevier Science Ltd. All rights reserved.

The aim of the present study was to analyse whether riluzole, a compound that interacts with the voltage-dependent sodium channel and impairs glutamatergic transmission, would exhibit a neuroprotective activity in a model of Parkinson's disease in the rat. Impaired skilled forelimb use, circling behavior, and altered dopaminergic metabolism of the mesotelencephalic system were evaluated in unilaterally 6-hydroxydopamine-lesioned rats. Riluzole was administered twice 15 min before, and 24 h after, the lesion. Riluzole reduced both the contralateral rotations induced by apomorphine and the ipsilateral ones elicited by amphetamine. Moreover, the decreased dopaminergic metabolism seen after 6-hydroxydopamine injection was attenuated in the riluzole-treated animals, at both the striatal and nigral levels. These biochemical and behavioral results demonstrate the ability of riluzole partially to protect the degeneration of the nigrostriatal dopaminergic neurons induced by the toxin 6-hydroxydopamine. Perhaps, the most striking evidence for the protective effect of riluzole was that this compound improved the skilled paw use, a complex sensorimotor behavior which is not easily ameliorated by palliative therapies such as dopaminergic grafts. These results extend previous data showing that riluzole counteracts the toxicity induced by 1-methyl-4-1,2,3,6-tetrahydropyridine and 1-methyl-4-phenylpyridinium in rodent dopaminergic neurons. The use of riluzole may be considered of potential interest for the neuroprotective therapy of Parkinson's disease.

The NMDA antagonist and neuroprotective effects of RPR 104632 (2H-1,2,4-benzothiadiazine-1-dioxide-3-carboxylic acid), a new benzothiadiazine derivative, with affinity for the glycine site of the NMDA receptor-channel complex are described. RPR 104632 antagonized the binding of [3H]5,7-dichlorokynurenic acid to the rat cerebral cortex, with a Ki of 4.9 nM. This effect was stereospecific, since the (-)-isomer was 500-fold more potent than the (+)-isomer. The potent affinity of RPR 104632 for the glycine site was confirmed by the observation that RPR 104632 inhibited [3H]N-[1-(2-thienyl)cyclohexyl]-3,4-piperidine ([3H]TCP) binding in the presence of N-methyl-D-aspartate (NMDA) (IC50 = 55 nM), whereas it had no effect on the competitive NMDA site or on the dissociative anaesthetic site. RPR 104632 inhibited the NMDA-evoked increase in guanosine 3',5'-cyclic monophosphate (cGMP) levels of neonatal rat cerebellar slices (IC50 = 890 nM) in a non-competitive manner and markedly reduced NMDA-induced neurotoxicity in rat hippocampal slices and in cortical primary cell cultures. These results suggest that RPR 104632 is a high-affinity specific antagonist of the glycine site coupled to the NMDA receptor channel with potent neuroprotective properties in vitro.

This study compares certain behavioural consequences of partial and complete unilateral lesions of the dopaminergic mesotelencephalic system. We investigated skilled forelimb use, rotations induced by apomorphine and amphetamine, and dopaminergic metabolism of the nigrostriatal system of rats that had received a unilateral injection of 6-hydroxydopamine into the medial forebrain bundle. The rats classified Apo(+), that rotated after the administrations of apomorphine, had a complete lesion of the nigrostriatal system, whereas those classified Apo(−), that did not rotate after the administration of apomorphine, had a partial lesion of the nigrostriatal system. In the Apo(+) rats, 99.8% of the dopamine in the striatum was depleted, as was 85% of that in the substantia nigra. For the Apo(−) rats, 72% of the dopamine in the striatum was depleted as was 56% of that in the substantia nigra. When investigated with the staircase test, the animals with the most severe dopamine depletions were those most impaired in the paw reaching task. Complete and partial unilateral depletions of the dopaminergic mesotelencephalic system impaired the heirarhic phases of paw reaching differently. A complete dopamine depletion, but not a partial one, decreased the number of attempts made with the contralateral paw, and induced a bias towards the ipsilateral paw. A partial dopamine lesion impaired the sensorimotor co-ordination of both paws, whereas the complete dopamine lesion had a greater effect on the contralateral paw than on the ipsilateral paw. The mild paw reaching impairments observed in animals with moderate depletions of dopamine are proposed as a model of the early symptoms of Parkinson's disease that may be useful for the development of protective or restorative therapies.

The effects of riluzole and lamotrigine, two agents which interfere with the release of glutamate (GLU), and MK-801, a blocker of N-methyl-D-aspartate (NMDA) receptors, were compared in the model of methamphetamine-induced depletion of dopamine (DA) levels in mice. Repeated injections with methamphetamine (4 x 5 mg/kg i.p.) markedly decreased levels of DA, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) levels. When mice were treated with riluzole (2 x 10 mg/kg p.o.), no protection was observed against the decrease in DA and the two metabolites. Lamotrigine (2 x 10 mg/kg p.o.) was also inactive. Treatment with MK-801 (2 x 2.5 mg/kg i.p.) antagonized the decrease in DA, DOPAC and HVA levels induced by the neurotoxin. Thus, unlike an NMDA blocker, drugs that interfere with GLU release did not antagonize the methamphetamine-induced DA neurotoxicity in mice. The consequences of this inactivity are discussed in terms of the reliability of this model to test new drugs with putative efficacy in the treatment of Parkinson's disease.

Superfusion of the rat striatum with 100 microM of 1-methyl-4-phenylpyridinium (MPP+) induced a 70-fold increase in dopamine (DA) release and a decrease in the extracellular levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). Pretreatment with riluzole (8 mg kg-1, i.p.), a compound that interferes with glutamatergic transmission, partially antagonized the effect of MPP+ on the release of DA, but did not change the effects of this toxin on the efflux of DOPAC and HVA. Riluzole did not affect the increase in DA release induced by MPP+ in vitro. The in vivo efficacy of riluzole on MPP(+)-induced DA release could be due to its central interference with glutamatergic transmission. Our data point to a protective role of riluzole with regard to DA release, a marker of the neuronal impairment induced by MPP+, a pro-parkinsonian neurotoxin.

RP 66055, a riluzole derivative, has been characterized as a potent anticonvulsant and in vivo neuroprotective agent.

We have studied the effects of neurotensin (NT) on the release of [3H]dopamine ([3H]DA) evoked by terminal depolarization with either K+, veratridine or 4-aminopyridine (4-AP). NT (1-1000 nM) induced a net potentiation (up to 170%) of the K+ (25 mM)-evoked release of [3H]DA. The capacity of NT to potentiate the effect of K+ ions decreased as the K+ concentration rose from 25 to 50 mM and totally disappeared at this high K+ concentration. NT (100 nM; 1,000 nM) had no significant effect on the veratridine (1.5; 5 microM) or 4-AP (20 microM) -evoked release of [3H]DA. The relevance of these experimental models of DA release to physiological transmitter release remains to be established. Those data highlight the complexity of the modulation of evoked neurotransmitter release by pharmacological agents.

We studied the effects of KATP channel blockers on [3H]GABA release in the absence of glucose in rat hippocampal slices. The omission of glucose induced a marked increase in the efflux of [3H]GABA, which was antagonized by TTX (1 microM), but not by MK 801 (1 microM) or DNQX (100 microM). Glibenclamide (10-100 microM) increased dose-dependently the release of [3H]GABA evoked in the absence of glucose. An increase in [3H]GABA release was also observed with gliquidone (100-300 microM), another sulfonylurea. The potentiation of [3H]GABA release induced by glibenclamide (100 microM) was antagonized by DNQX but not by MK 801. Thus, in the absence of glucose, KATP channel blockers enhance the release of GABA from rat hippocampal slices; this effect seems to be mediated by an overstimulation of non-NMDA glutamate receptors. On the basis of results reported in the present paper, we suggest that KATP channels may play a role in the regulation of GABAergic activity during hypoglycemia.

The effects of different potassium channel blockers on tritiated dopamine ([3H]DA) release were investigated in rat striatal slices in the presence of pargyline and nomifensine (10 μM each). 4-Aminopyridine (4-AP; 10 and 30 μM) and 3,4-diaminopyridine (3,4-DAP; 30 μM) markedly increased the basal tritium outflow, whereas tetraethylammonium (TEA; 100–1000 μM) was without effect. The facilitating effect of 4-AP (10 μM) on spontaneous release was Ca2+- and K+-dependent. Moreover, the 4-AP-induced increase in spontaneous release was abolished in the presence of tetrodotoxin, indicating that voltage-dependent Na+ channels were involved in the release mechanism. 4-AP (10 and 30 μM) induced a dose-dependent decrease in K+-evoked [3H]DA release. This effect was confirmed with 3,4-DAP (30 μM). When striatal slices were depolarized with veratridine (5 μM), these two aminopyridines increased the evoked release of [3H]DA. TEA increased both K+- and veratridine-evoked [3H]DA release. These biochemical results are consistent with electrophysiological differences between the mechanism of action of aminopyridines and that of TEA.

γ-Butyrolactone is a drag known to inhibit the flow of electrical impulses in dopaminergic neurones and to prevent dopamine release in the striatum. In this study, we investigated the effects of this compound on the retrograde transport of neurotensin in the nigrostriatal dopanunergic pathway. The amount of radioactivity measured in the substantia nigra after injection of iodinated neurotensin into the striatum and the time course of its accumulation were not found to be modified in γ-butyrolactone-treated rats. We therefore conclude that the retrograde axonal transport of neurotensin in the nigrostriatal pathu-γ is not affected by reduced dopaminergic firing or by inhibition of the striatal release of dopamine.

RPR 118723 ((8-chloro-5-methyl-2,3-dioxo-1,4-dihydro-5H-indeno[1, 2-b]pyrazin-5-yl) acetic acid) was previously reported to exhibit potent affinity for the glycine site of the N-methyl-D-aspartate (NMDA) receptor-channel complex in the nanomolar range (K(i)=3.1+/-0. 8 nM). We now report on the effects of RPR 118723 in two functional tests reflecting the interaction between the glycine site and the NMDA receptor. First, RPR 118723 potently inhibited [3H]N-[1-(2-thienyl)cyclohexyl]-3,4-piperidine ([3H]TCP) binding in the presence of NMDA (IC(50)=3.5+/-0.4 nM). Second, RPR 118723 antagonized the NMDA-induced increase in [3H]dopamine release in mouse striatal slices (IC(50)=8.0+/-1.1 nM). In both experimental models, an excess of glycine reversed the effect of RPR 118723. These results show that RPR 118723 interferes functionally in the nanomolar range with the glycine site coupled to the NMDA receptor in vitro. The blockade of the glycine site with RPR 118723 may be useful for the therapy of the disorders linked to excessive NMDA stimulation.

We studied the effects of ebselen (a seleno-organic anti-oxidant), on the release of the apoptogenic factor, cytochrome c, in two different experimental situations damaging mitochondria: (1) Fe2+/citrate, known to induce lipid peroxidation consecutively to an oxidative stress; and (2) atractyloside, a ligand of the adenine nucleotide translocator. The effects of ebselen were compared to those of butylated hydroxytoluene (BHT, an inhibitor of lipid peroxidation), and cyclosporine A (CsA, a classical pore antagonist). Ebselen, like BHT, inhibited Fe2+/citrate-induced release of cytochrome c, whereas CsA was inactive. On the contrary, neither ebselen nor BHT inhibited atractyloside-induced release of cytochrome c, whereas CsA was potently active. The antioxidant properties of ebselen may protect mitochondria from the consequences of the release of cytochrome c. Thus, it is suggested that the neuroprotective effect of ebselen previously demonstrated in humans and in animals may be due, at least in part, to a mitochondrial protection.

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.

Riluzole is an anti-excitotoxic substance with sodium channel blocking properties that has been shown to possess neuroprotective activities in a wide variety of animal models of neurodegeneration including models of amyotrophic lateral sclerosis (ALS), Huntington’s disease, ischaemia and trauma. Riluzole is currently approved for the treatment of patients suffering from ALS. Studies in rodents have shown that riluzole is able to counteract toxin-induced degeneration of nigro-striatal neurons. For example, dosage of mice brain for dopamine (DA) content by HPLC indicate that riluzole prevents the diminution of striatal DA levels induced by co-treatment with N-methyl-4-phenyl-l,2,3,6-tetrahydropyridine (MPTP). Similarly, riluzole attenuates the turning behavior produced by administration of apomorphine or amphetamine to rats that have received an unilateral injection of 6-hydroxydopamine. Behavioral and histological experiments were conducted in marmosets to investigate the antiparkinsonian profile of riluzole in primates. Systemic administration of a low doses regimen of MPTP (2 mg/kg subcutaneously twice, 1 week apart) produced a slowly evolving parkinsonian syndrome as evidenced by quantification of locomotor activity via computerized image analysis and clinical examination using a neurological evaluation scale. Administration of riluzole (10 mg/kg orally twice daily) significantly attenuated the locomotor alterations produced by the toxin. This effect was protracted over the 18 day observation period, at the end of which the brain of the animals was fixed for histobogical examination. Analysis of tyrosine hydroxylase immunofluorescence and cresylviolet staining revealed a significant preservation of the density of dopaminergic cells in the substantia nigra pars compacta (SNpc).

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.

We have examined the effects of riluzole, a neuroprotective drug which stabilizes voltage-dependent sodium channels in their inactivated state and inhibits the release of glutamate in-vivo and in-vitro, on the release of newly taken up [3H]dopamine induced by ouabain, a potent and selective inhibitor of Na+/K+-ATPase in mouse striatal slices in-vitro. Riluzole potently (IC50 (concentration resulting in 50% inhibition) = 0.9±0.3 μM) and dose-dependently antagonized ouabain-stimulated [3H]dopamine release, the effect being observed at low concentrations. Tetrodotoxin (1 μM) and nomifensine (10 μM) also abolished ouabain-induced [3H]dopamine release. Blockade of glutamate receptors with dizocilpine (1 μM) and 6-(1H-imidazol-1-yl)-7-nitro-2,3(1H,4H)-quinoxalinedione (YM-90K; 10 μM), alone or in combination, was without effect. Incubation of striatal slices with 50 μM La3+, which blocks voltage-dependent calcium channels, did not inhibit [3H]dopamine release induced by ouabain. The potent effects of riluzole observed in this model are probably related to its ability to block voltage-dependent sodium channels. The consequences of this activity are critically discussed in relation to the protective action of riluzole previously reported in various models of Parkinson’s disease and other neurodegenerative disorders.

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.

3−Nitropropionic acid (3−NPA) is a metabolic poison that produces lesions of striatal intrinsic neurones such as γ-aminobutyric acid (GABA) neurones. This study was carried out to determine whether 3−NPA would impair the ability of striatal GABAergic neurones to withstand hypoglycaemic stress. 3−NPA (500 μM) did not affect [3H]GABA release from striatal slices under normal (11 mM) glucose concentrations. When the glucose concentration was lowered to 0ṁ3 mM, however, 3−NPA greatly potentiated the leakage of [3H]GABA from the slices. Blockade of N-methyl-d-aspartate (NMDA) or α-amino−3−hydroxy−5−methyl−4−isoxazolepropionic acid (AMPA) receptors with 1 μM 5−methyl−10,11−dihydro−5H-dibenzo[a,d]cyclohepten−5,10−imine (MK−801) or 10 μM 2,3−dihydroxy−6−nitro−7−sulpha-moylbenzo[F]quinoxaline (NBQX), respectively, or a combination of both, had no effect. However, blockade of voltage-dependent sodium channels with tetrodotoxin totally antagonized the [3H]GABA overflow induced by the combination of 3−NPA and hypoglycaemia. Riluzole (10 to 100 μM), a neuroprotective agent that stabilizes the inactivated state of the voltage-dependent sodium channel, also dose-dependently antagonized the increase in [3H]GABA release induced by the combination of the two stresses.

Previous studies have shown that riluzole (2-amino-6-trifluoromethoxy-benzothiazole), a drug which interferes with glutamate neurotransmission, has a neuroprotective action in rodent models of global and focal cerebral ischemia. In this pilot study, the protective and palliative effects of riluzole have been examined using an animal model of Parkinson's disease. Two monkeys were rendered hemiparkinsonian by one intracarotid injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and motor signs were evaluated using clinical examination and electromyographic recordings. When riluzole (4 mg/kg) was administered before the injection of MPTP, parkinsonian motor symptoms, in particular bradykinesia and rigidity, were absent. When injected daily in one monkey which presented stable motor symptoms, bradykinesia and rigidity were significantly reduce d. Riluzole pretreatment induced a persistent increase in dopamine turnover when compared to MPTP alone. Thus, a possible neuroprotection and a facilitation of dopamine release may explain the behavioural effects reported with riluzole treatment. These preliminary results suggest that riluzole could possess neuroprotective and palliative effects in a primate model of Parkinson's disease.

There are only a few ligands available for labelling brain receptors simply because in vivo binding requires more severe experimental conditions than in vitro binding. We now describe the in vivo binding properties of [3H]RP 62203, a new potent and selective 5-HT2 antagonist. After intravenous injection into rats, [3H]RP 62203 accumulated predominantly in brain regions containing 5-HT2 receptors, with a frontal cortex/cerebellum ratio of 6 to 7. A good correlation was obtained between the regional distribution of [3H]RP 62203 in the brain and the density of 5-HT2 receptors measured in vitro. In vivo binding of [3H] RP 62203 was saturable in the frontal cortex but not in the cerebellum. The Bmax in the frontal cortex was equal to 42.5 fmol/mg, thus in the same range as was found in vitro. The 5-HT2 selectivity was ascertained by displacement (prevention) experiments; 5-HT2 antagonists or the agonist 2,5-dimethoxy- 4-iodophenylisopropylamine could prevent specific labelling of [3H]RP 62203 only in brain regions containing 5-HT2 receptors. Interestingly, the radioactivity remaining in various brain regions after displacement with pipamperone corresponded exactly to that measured in the cerebellum, with or without pipamperone. In conclusion, [3H]RP 62203 possesses striking properties of in vivo binding which make it a suitable candidate for examining 5-HT2 receptors in human brain by positron emission tomography scanning.

The effects of veratridine-induced depolarization on [3H]dopamine ([3H]DA) release in the presence of a physiological (5 Mm) or a depolarizing (25 Mm) concentration of K+ were studied in-vitro in rat superfused striatal ribbons. A combination of the two depolarizing agents induced a marked potentiation in the overflow of [3H]DA, giving an overall 3- to 5-fold increase in veratridine activity. This potentiation was completely antagonized by tetrodotoxin (100 Nm). These studies indicated that K+-induced depolarization can increase the potency of veratridine in releasing dopamine from terminals.

Two hours after the injection of (3-[125I]iodotyrosyl3) neurotensin into the striatum, a labeling was observed in the ipsilateral substantia nigra. In the present study, we demonstrated by HPLC that this radioactivity corresponded to intact neurotensin and to degradation products of this peptide. This finding provides the first clearcut evidence that a neuropeptide can be internalized and retrogradely transported in brain neurons. Therefore, the fact that intact neurotensin can be seen to exist over a long period of time in the cell body suggests that the retrograde transport process could perhaps be involved in the long-term effects of neuropeptides.

Hydrogen bromide gas phase corrosion of welded 316L electropolished stainless steel tubings to be used for the distribution of ultra clean gases has been investigated by using two techniques: one is leaching of the corroded tubings and subsequent chemical analysis of the dissolved metallic elements, the other one is bromine penetration depth at the stainless steel surface by Auger spectroscopy. These test procedures were applied to optimize the back shielding gas used for orbital welding of such tubings. A correlation is found between the oxygen concentration in the back shielding gas and the gas phase corrosion. Purified argon as back shielding gas insures best corrosion resistance due to the minimum thickness of superficial iron oxide, and maximal chromium concentration underlying the top iron/manganese oxide layer. Tubings made of stainless steel containing very small concentration of manganese exhibit lower level of corrosion. A corrosion enhancement mechanism by manganese is proposed.

The effects of a single injection of the palmitic ester of pipotiazine on dopamine synthesis were examined in slices of the striatum, the olfactory tubercule + nucleus accumbens and the frontal and anterior cingulate parts of the cerebral cortex in the rat. In the two first structures, dopamine synthesis was determined by measuring the rate of formation of 3H−H2O during the conversion of l-3,5-3H-tyrosine into 3H-Dopa. Newly synthesized 3H-dopamine and 3H-noradrenaline were measured in the cerebral cortex. In some experiments, the specific activity of tyrosine in the tissues was determined in order to calculate the conversion index of tyrosine into dopamine.

The use of [ 3 H]thioproperazine (a neuroleptic of the phenothiazine family) as a ligand for neuroleptic receptors has been studied in vitro in various brain regions of the rat. Specific thioproperazine binding has been observed in two regions rich in dopamine terminals: striatum and nucleus accumbens. No specific binding was observed in frontal cortex and hippocampus. Thioproperazine binds with high affinity ( K d = 0.30 nM) to rat microsomal striatal fraction. In this fraction the number of binding sites (1.47 pmoles/mg protein) is rather similar to that found with haloperidol and spiperone, two neuroleptics of the butyrophenone type. Subcellular distribution of thioproperazine binding reveals a pattern similar to that of 5'-nucleotidase, a plasma membrane marker. This indicates that the receptor sites of [ 3 H]thioproperazine are associated with membrane-like structures. Inhibition studies indicate that all neuroleptic compounds, belonging to different chemical families, are capable of displacing thioproperazine from its binding sites. The most potent drugs are spiperone, thioproperazine and pipotiazine. Dopamine and the two dopamine agonists, apomorphine and bromocryptine, present affinity for thioproperazine binding sites. It is concluded that thioproperazine, a phenothiazine neuroleptic, can be used, like the butyrophenone derivatives haloperidol and spiperone, as a ligand for studying neuroleptic receptors.

The action of suriclone (R.P. 31,264), a new non-benzodiazepine compound of the cyclopyrrolone family with clinical anxiolytic activity was examined using biochemical and electrophysiological models supposed to be capable of revealing central GABAergic activity. Suriclone, which does not act directly on the γ-aminobutyric acid (GABA) receptor (muscimol binding assay), markedly reduced the increase of striatal HVA level induced in the rat by a neuroleptic and decreased the cerebellar vermis cGMP content. Moreover, in the cat, suriclone was able to enhance dorsal root potential amplitude which reflects an increase of the presynaptic inhibition. In view of these results, a central GABAergic mechanism of action may be proposed for suriclone.

Up to now the only drugs known to be able to inhibit the binding of benzodiazepines to rodent brain receptors are members of this chemical family. Zopiclone (RP 27 267), a new drug with a pharmacological profile similar to that of chlordiazepoxide and nitrazepam but entirely different chemically from benzodiazepines, has been tested for its ability to inhibit benzodiazepine binding. In vitro and in vivo studies have shown that zopiclone is able to inhibit the binding of [3H] diazepam and [3H] flunitrazepam to brain receptors. The potency of zopiclone is quite comparable to that of diazepam and nitrazepam in vitro and to that of chlordiazepoxide in vivo . These results confirm the pharmacological similarities existing between zopiclone and the benzodiazepines.

Zopiclone (ZPC; RP 27, 267) is a new hypnotic which belongs to the new family of cyclopyrrolones chemically unrelated to benzodiazepines (BZD). Psychopharmacological studies on ZPC have shown that it exhibits the five main types of activities —anti-convulsant, anti-agressive, myorelaxant, sedative-hypnotic and anti-conflict (anxiolytic) — which have been considered as being partly those of barbiturates and more specifically those of minor tranquilizers of the BZD type. ZPC possesses in rat brain a high affinity for BZD receptors (Ki values of 36 nM in hippocampus and 22.5 nM in cerebellum). ZPC does not recognize other brain receptors even the peripheral BZD receptor sites. Studies performed with [3H]-ZPC have confirmed the high affinity of ZPC (KD = 13 4 nM in hippocampus). ZPC binding is not enhanced by GABA, or barbiturates and is less sensitive to photoaffinity labelling (low photoshift) than BZD agonists. These results could indicate that ZPC acts on a site distinct from BZD.Centre de Recherches de V...