Non-peptidic CRF1 receptor antagonists for the treatment of anxiety, depression and stress disorders.

Anxiety and depression are psychiatric disorders that constitute a major health concern worldwide, and new pharmacological approaches with the potential for improved efficacy and decreased side effect profiles relative to currently marketed drugs are desired. Since the identification of corticotropin releasing factor (CRF) by Vale and colleagues in 1981, an extensive research effort has solidified the importance of this 41 amino acid peptide in mediating the body's behavioral, endocrine, and autonomic responses to stress. The further identification of CRF receptor subtypes has provided compelling targets for novel pharmaceutical agents. The present review focuses on the potential of non-peptidic antagonists of the CRF(1) receptor subtype as a novel therapeutic approach for the treatment of anxiety and depression. The first section reviews preclinical and clinical evidence implicating CRF, in general, and CRF(1) receptors, in particular, in anxiety and depression. Clinical studies have demonstrated a dysfunctional hypothalamic-pituitary-adrenal (HPA) axis and/or elevated CRF levels in depression and in some anxiety disorders. Preclinical data utlilizing correlational methods, genetic models, and exogenous CRF administration techniques in rodents and non-human primates supports a link between hyperactive CRF pathways and anxiogenic and depressive-like symptoms. Studies employing the use of receptor knockouts and selective, non-peptidic antagonists of the CRF(1) receptor have demonstrated anxiolytic and antidepressant effects under certain types of laboratory conditions. A Phase II, open-label, clinical trial in major depressive disorder has reported that a CRF(1) receptor antagonist was safe and effective in reducing symptoms of anxiety and depression. In the second section, a topological approach is used to describe the design strategies employed to produce potent, non-peptidic CRF(1) receptor antagonists. Two main topologies, featuring a center core, a top side-chain, and a pending aromatic ring, can be used to characterize the vast majority of currently known CRF(1) receptor antagonists. By exploiting some of these structural elements, pharmacological, physicochemical, and pharmacokinetic properties can be modulated and optimized. However, as a result of a relatively conservative iteration process during the structural optimization, the chemical space presently defined by the existing CRF(1) receptor antagonists still remains fairly narrow. Expanding these structural and topological boundaries, while optimizing the "drug-like" properties of the CRF(1) receptor antagonists, seems to be a common objective across pharmaceutical companies to maximize the chances for a clinical success in the near future.

Effects of the CRF(1) receptor antagonist, CP 154,526, in the separation-induced vocalization anxiolytic test in rat pups.

CRF(1) receptor antagonists have been proposed as novel pharmacological treatments for depression, anxiety and stress disorders. The primary goal of the present study was to evaluate the effects of the CRF(1) receptor antagonist, CP 154,526, in the separation-induced vocalization (SIV) model of anxiety. Nine- to 11-day-old rat pups were separated from their litter and the effects of intraperitoneally administered test compounds on the elicited ultrasonic vocalizations were measured. Side-effect potential was assessed using a modified inclined plane test ('time on an inclined plane', or TIP), and using negative geotaxis. SIV was reduced by CP 154,526 at doses that did not affect TIP or negative geotaxis, a profile like that of the 5-HT(1A) partial agonist buspirone. The benzodiazepine anxiolytic, diazepam, decreased SIV but also produced significant side effects at one to three-fold higher doses. Additional pharmacological characterization of SIV demonstrated anxiolytic-like effects of the atypical antipsychotic, clozapine, but not the typical antipsychotic, haloperidol, and of the serotonin reuptake inhibitor, zimelidine, but not the norepinephrine reuptake inhibitor, desipramine. In summary, the data support the conclusion that selective CRF(1) receptor antagonists may have utility in anxiety and stress disorders. The data further support the use of separation-induced vocalizations for identifying mechanistically diverse compounds with anxiolytic actions in man.

Mixed D2/5-HT2A antagonism of cocaine-induced facilitation of brain stimulation reward.

Previous behavioral, neurochemical and neurophysiological experiments have shown that selective 5-HT2A and mixed D2/5-HT2A antagonists can attenuate some, but not all, responses to amphetamine. The generality of these findings were determined in the present experiment by assessing the effect of mixed D2/5-HT2A antagonists on cocaine-induced facilitation of ventral tegmental area self-stimulation in rats. Although amphetamine and cocaine influence activity in monoaminergic neurons through different mechanisms, our previous research has shown that selective D2 and 5-HT2A antagonists have similar effects on behavioral responses to these psychostimulants. Therefore, we expected a similar pattern of results using mixed D2/5-HT2A antagonists. As shown previously, cocaine decreased self-stimulation threshold in a dose-dependent manner. Haloperidol and the mixed D2/5-HT2A antagonists risperidone and MDL 28, 133A antagonized cocaine-induced facilitation of self-stimulation, but only at doses that increased baseline self-stimulation threshold. There was a significant correlation (r = 0.87, p < 0.001) between antagonist-induced change in baseline threshold and attenuation of cocaine's effect on threshold. Taken together, the results of this and previous experiments support the importance of D2 receptors in the mechanisms of brain stimulation reward. 5-HT2A receptors appear not to be involved in mediation of both brain stimulation reward and amphetamine- and cocaine-induced facilitation of brain stimulation reward.

Mixed D2/5-HT2 antagonism differentially affects apomorphine- and amphetamine-induced stereotyped behavior.

Evidence supports the hypothesis that psychostimulant stereotypy is mediated through postsynaptic dopamine receptors. Given the recent findings of behavioral, neurochemical and electrophysiological studies showing 5-HT2 modulation of dopamine systems, a series of experiments were undertaken to assess the ability of D2 and 5-HT2 antagonists to reverse apomorphine and amphetamine stereotypy in the rat. Haloperidol reduced stereotyped behavior induced by d-amphetamine (50% reduction with 0.162 mg/kg) and apomorphine (50% reduction with 0.112 mg/kg) MDL 28,133A, a mixed D2/5-HT2 antagonist, also reduced stereotypy in the apomorphine group (50% reduction with 3.89 mg/kg) but was much less effective in antagonizing the effects of d-amphetamine (not even a 25% reduction with 9.0 mg/kg). MDL 100,907, a selective 5-HT2 antagonist, was ineffective at reducing stereotyped behavior induced by either stimulant. Thus, 5-HT2 modulation of dopaminergic activity was not demonstrated in the case of psychostimulant stereotypy. Furthermore, 5-HT2 antagonism did not induce stereotypy, as has been proposed in some models. These findings provide further support for the hypothesis that antipsychotic medications with high affinity for 5-HT2 receptors do not interfere with the regulation of the nigrostriatal dopaminergic system and, therefore, would be less likely to produce extrapyramidal side effects.

Pharmacological characterization of MDL 105,519, an NMDA receptor glycine site antagonist.

MDL 105,519, (E)-3-(2-phenyl-2-carboxyethenyl)-4,6-dichloro-1 H-indole-2-carboxylic acid, is a potent and selective inhibitor of [3H]glycine binding to the NMDA receptor. MDL 105,519 inhibits NMDA (N-methyl-D-aspartate)-dependent responses including elevations of [3H]N-[1,(2-thienyl)cyclohexyl]-piperidine ([3H]TCP) binding in brain membranes, cyclic GMP accumulation in brain slices, and alterations in cytosolic CA2+ and NA(+)-CA2+ currents in cultured neurons. Inhibition was non-competitive with respect to NMDA and could be nullified with D-serine. Intravenously administered MDL 105,519 prevented harmaline-stimulated increases in cerebellar cyclic GMP content, providing biochemical evidence of NMDA receptor antagonism in vivo. This antagonism was associated with anticonvulsant activity in genetically based, chemically induced, and electrically mediated seizure models. Anxiolytic activity was observed in the rat separation-induced vocalization model, but muscle-relaxant activity was apparent at lower doses. Higher doses impair rotorod performance, but were without effect on mesolimbic dopamine turnover or prepulse inhibition of the startle reflex. This pattern of activities differentiates this compound from (5R,10S)-(+)-5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) and indicates a lower psychotomimetic risk.

Preclinical characterization of MDL 27,192 as a potential broad spectrum anticonvulsant agent with neuroprotective properties.

The compound 5-(4-chlorophenyl)-2,4-dihydro-4-ethyl-3H-1,2,4-triazol-3-one (MDL 27,192) was evaluated in a variety of rodent models to assess its anticonvulsant profile and its potential neuroprotective activity. MDL 27,192 demonstrated anticonvulsant activity in a wide range of epilepsy models that are genetically-based (audiogenic seizures in the seizure susceptible DBA/2J or Frings mouse; spike wave seizures in genetic absence epilepsy rats of Strasbourg (GAERS), electrically-based (MES seizures in mice and rats, corneally-kindled seizures in rats) and chemically-based (bicuculline, PTZ, picrotoxin, 3-mercaptopropionic acid, quinolinic acid and strychnine). When compared to valproate, orally administered MDL 27,192 was 17-48-fold more potent as an anticonvulsant and showed a safety index one to three-fold greater. Following a timed intravenous administration of PTZ to mice, MDL 27,192, but not phenytoin or carbamazepine, consistently increased the latencies to first twitch and clonus. MDL 27,192 was active in a genetic model of absence epilepsy, the GAERS rat model. These data indicate that MDL 27,192 likely exerts its anticonvulsant action by affecting seizure spread and by raising seizure threshold. MDL 27,192 did not display any signs of tolerance following subchronic (15 day) administration. In tests of neuroprotective potential, MDL 27,192 reduced infarct volume in a permanent middle cerebral artery occlusion model of focal cerebral ischemia in rats and reduced the loss of hippocampal dentate hilar neurons in an animal model of unilateral head injury. In summary, MDL 27,192 possesses a broad-spectrum anticonvulsant profile. The potential for reduced tolerance and neuroprotective activity are additional positive features of MDL 27,192's preclinical profile.

The peripheral NK-1/NK-2 receptor antagonist MDL 105,172A inhibits tachykinin-mediated respiratory effects in guinea-pigs.

1. Stimulation of sensory nerves causes release of tachykinins, including substance P (SP) and neurokinin A (NKA), which produce a variety of respiratory effects via NK-1 and NK-2 receptors, respectively. Hence, development of a compound which could potently and equivalently antagonize both receptors was pursued. 2. MDL 105,172A ((R)-1-[3-(3,4-dicholorophenyl)-1-(3,4,5-trimethoxybenzoyl)- 3-pyrrolidinyl]-4- phenyl-piperidine-4-morpholinecarboxamide) exhibited high affinity for NK-1 (4.34 nM) and NK-2 (2.05 nM) receptors. In vitro, the compound antagonized SP (pA2 = 8.36) or NKA (pA2 = 8.61)-induced inositol phosphate accumulation in tachykinin monoreceptor cell lines. 3. In anaesthetized guinea-pigs, MDL 105,172A inhibited SP-induced plasma protein extravasation (ED50 = 1 mg kg-1, i.v.) and [beta-Ala8]NKA 4-10-induced bronchoconstriction (ED50 = 0.5 mg kg-1, i.v.) indicating NK-1 and NK-2 antagonism, respectively. 4. Capsaicin was used to elicit respiratory effects in anaesthetized and conscious guinea-pigs; the latter were inhibited by MDL 105,172A following i.v. (ED50 = 1 mg kg-1) or oral (ED50 = 20 mg kg-1) administration. Hence, MDL 105,172A can inhibit pulmonary responses to tachykinins released endogenously in the airways. 5. At doses up to 200 mg kg-1, p.o., MDL 105,172A failed to inhibit repetitive hind paw tapping induced by i.c.v GR 73632, and NK-1 selective agonist, in gerbils, whereas CP-99,994 (0.87 mg kg-1, s.c.) completely ablated the effect. These data suggest that MDL 105,172A does not penetrate the central nervous system (CNS) and its tachykinin antagonism is restricted to the periphery. 6. MDL 105,172A is a non-peptide, potent, equivalent antagonist of NK-1 and NK-2 receptors. Its ability to inhibit both exogenously administered as well as endogenously released tachykinins support its use in examining the role of sensory neuropeptides in diseases associated with neurogenic inflammation including asthma.

Cyclic nitrone free radical traps: isolation, identification, and synthesis of 3,3-dimethyl-3,4-dihydroisoquinolin-4-ol N-oxide, a metabolite with reduced side effects.

A C-4 hydroxylated metabolite (2, 3,3-dimethyl-3,4-dihydroisoquinolin-4-ol N-oxide) of the previously described cyclic nitrone free radical trap 1 (3,3-dimethyl-3,4-dihydroisoquinoline N-oxide, a cyclic analog of phenyl-tert-butylnitrone (PBN)) was isolated, identified, and synthesized. The metabolite (2), though a less potent antioxidant than 1 in an in vitro lipid peroxidation assay, showed greatly reduced acute toxicity and sedative properties. Several analogs of 2 were prepared in attempts to improve on its weak antioxidant activity while retaining the desirable side effect profile. Effective structural changes included replacement of the gem-dimethyl moiety with spirocycloalkane groups and/or oxidation of the alcohols to the corresponding ketones. All of the analogs were more lipophilic (log k'(w)) and more active in the standard lipid peroxidation assay than 2. In addition, some of the compounds were able to protect cerebellar granule cells against oxidative damage (an in vitro model of oxidative brain injury) with IC50 values well below the value of the lead compound 1. The ketones, as predicted, were much more potent than 2 (and 1) in both of the above assays (up to ca. 200-fold). However, only compounds with a hydroxyl or an acetate group at C-4 displayed significantly reduced acute toxicity and sedative properties relative to those of 1. Importantly, the diminishment of toxicity and sedation were not the result of a lack of brain penetration as both 2 and the corresponding ketone (3,3-dimethyl-3,4-dihydro-3H-isoquinolin-4-one N-oxide) achieved equal or greater brain levels than those of 1 when administered to rats i.p.

Effects of the selective 5-HT2A receptor antagonist MDL 100,907 on MDMA-induced locomotor stimulation in rats.

(+/-)3,4-Methylenedioxymethamphetamine (MDMA) releases dopamine and serotonin in vivo and stimulates locomotor activity. Previous work demonstrated that MDMA-stimulated dopamine release could be reduced by the selective 5-HT2A receptor antagonist [R-(+)-a- (2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidinem ethanol] (MDL 100,907). In the present study MDL 100,907 significantly reduced MDMA-stimulated locomotion without affecting basal levels of locomotion. Other agents with 5-HT2A antagonist activity (ritanserin, clozapine, MDL 28,133A, or methiothepin), as well as agents that block 5-HT1A-(propranolol), D2-(haloperidol), or D1 receptors (SCH 23390) also reduced MDMA-stimulated locomotion. Intraventricularly administered 5,7-dihydroxytryptamine decreased regional 5-HT levels and attenuated MDMA-stimulated locomotion. These data support the conclusion that serotonin released onto 5-HT2A receptors contributes to MDMA-stimulated locomotion and suggest that MDMA-stimulated locomotion may be useful as an in vivo behavioral measure of 5-HT2A antagonism. The data also support previous reports of contributions of 5-HT1A, D1 and D2 receptors to MDMA-stimulated locomotion. A preliminary time-course analysis indicating time-dependent contributions of different receptors to MDMA-stimulated locomotion suggests the potential utility of this model for characterizing potential atypical antipsychotic compounds.

Purification of the CIC-0 chloride channel from Torpedo california electroplax identification of a phosphorylation site for cAMP-dependent protein kinase.

The voltage-gated chloride channel (CIC-0) from the electric organ of Torpedo californica was purified by immunoaffinity chromatography. A polyclonal antibody was shown to specifically recognize the CIC-0 channel (M(r) 85,000-90,000) in a Western blot of total membrane proteins. As monitored by immunoprecipitation, the formation of antibody-antigen complexes in solution strongly depends on the detergent composition. The highest yield of precipitated CIC-0 was obtained from an incubation mixture containing both an anionic detergent, cholate or lauryl sulfate, and the zwitterionic detergent CHAPS. In contrast, immuno-precipitation of CIC-0 was largely reduced when cholate was exchanged for the nonionic detergent Triton x-100, suggesting that the efficient formation of immuno-complexes is favored by negatively charged detergent. In initial immunopurification experiments, in addition to CIC-0 a major contaminating polypeptide of M(r) approximately 115,000 was copurified, which represents the SITS-binding protein [Jentsch et al. (1989) Biochem. J. 261, 155]. Purification of CIC-0 could be increased from about 35% up to 60% homogeneity when immunoaffinity chromatography was performed in the presence of N-acetylglucosamine. Therefore the highly glycosylated SITS-binding protein most likely interacts with the CIC-0 protein via its carbohydrate parts. The purified CIC-0 channel was found to be phosphorylated by PKA in vitro to a level of 0.35-0.4 mol of phosphate incorporated per mol of CIC-0. Proteolytic digestion with endoproteinase GluC and HPLC-separation revealed two major phosphopeptides, which could be identified by amino acid sequence analysis as different size fragments of the same consensus phosphorylation site. Comparison of the peptide sequences with the deduced protein sequence of CIC-0 [Jentsch et al. (1990) Nature 348, 510; O'Neill et al. (1991) Biochem. Biophys. Acta 1129, 131] indicates serine 600 as the phosphorylated residue. Therefore, our results provide strong evidence that CIC-0 is phosphorylated at this single site by PKA in vitro.

Preclinical characterization of the potential of the putative atypical antipsychotic MDL 100,907 as a potent 5-HT2A antagonist with a favorable CNS safety profile.

In preclinical studies, [R-(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenyl)ethyl]-4- piperidinemethanol] [formula: see text] (MDL 100,907), a putative atypical antipsychotic, was characterized in vitro as a potent and selective ligand for the serotonin2A (5-HT2A) receptor and was evaluated in vitro and in vivo as a potent 5-HT2A receptor antagonist. Furthermore, MDL 100,907's potential CNS safety profile and selectivity as a potential antipsychotic agent were evaluated and compared with benchmark compounds. MDL 100,907 demonstrated low nanomolar or subnanomolar binding in vitro at the 5-HT2A receptor and showed a > 100-fold separation from all other receptors measured. MDL 100,907 had subnanomolar potency as a 5-HT2A antagonist in vitro in reversing 5-HT-stimulated inositol phosphate accumulation in NIH 3T3 cells transfected with the rat 5-HT2A receptor. In vivo, MDL 100,907 potently inhibited 5-methoxy-N, N-dimethyltryptamine-induced head twitches in mice or 5-hydroxytryptophan-induced head twitches in rats. In vivo functional tests in mice revealed a > 500-fold separation between doses that produced 5-HT2A antagonism and doses that produced alpha 1-adrenergic or striatal D2 antagonism. Using inhibition of D-amphetamine-stimulated locomotion in mice as a measure of potential antipsychotic efficacy, MDL 100,907 showed a superior CNS safety index relative to the reference compounds, haloperidol, clozapine, risperidone, ritanserin, and amperozide, in each of five tests for side effect potential, including measures of ataxia, general depressant effects, alpha 1-adrenergic antagonism, striatal D2 receptor antagonism, and muscle relaxation. MDL 100,907 did not antagonize apomorphine-induced stereotypes in rats, suggesting that it potentially lacks extrapyramidal side effect liability. MDL 100,907 showed selectivity as a potential antipsychotic in that it lacked consistent activity in selected rodent models of anticonvulsant, antidepressant, analgesic, or anxiolytic activity. In summary, these preclinical data indicate that MDL 100,907 is a potent and selective ligand at the 5-HT2A receptor. MDL 100,907's potent 5-HT2A antagonist activity might account for its activity in preclinical models of antipsychotic potential. Ongoing clinical evaluation with MDL 100,907 will test the hypothesis that 5-HT2A receptor antagonism is sufficient for antipsychotic activity in humans.

5-HT modulation of auditory and visual sensorimotor gating: I. Effects of 5-HT releasers on sound and light prepulse inhibition in Wistar rats.

Increasing evidence suggests an important role for serotonin (5-HT) neurons in the etiology and treatment of schizophrenia. The prepulse inhibition paradigm is used as a model for sensorimotor gating processes that are disrupted in schizophrenia. The present study assessed the general role of 5-HT in modulating auditory and visual prepulse inhibition in Wistar rats. A general overactivation of central serotonerigic pathways was produced pharmacologically by four different agents which all shared the common property of releasing 5-HT, i.e., p-chloroamphetamine, 3,4-methylenedioxymethamphetamine, N-ethyl-3,4-methylenedioxymethamphetamine, or fenfluramine. Within each test session, both sound and light prepulses were used to obtain a cross-modal assessment of auditory and visual sensory gating processes. All four 5-HT releasing agents produced dose-related disruptions of auditory and visual prepulse inhibition, with p-chloroamphetamine being the most potent. The releasers depressed baseline to varying degrees. The alpha 2-adrenergic agonist clonidine decreased baseline startle without substantially disrupting prepulse inhibition, demonstrating that the two effects were dissociable. Using fenfluramine as the most selective 5-HT releaser, two approaches were used to demonstrate 5-HT mediation of its disruptive effect on prepulse inhibition. In the first approach, the selective 5-HT uptake blocker MDL 28,618A was used to prevent fenfluramine-induced 5-HT release. In the second approach, prior exposure to a neurotoxic dose of p-chloroamphetamine (10 mg/kg) was used to produce a substantial, sustained depletion of cortical 5-HT, presumably reflecting the loss of 5-HT terminals. Both approaches reduced the disruptive effect of fenfluramine on auditory and visual prepulse inhibition, thereby demonstrating 5-HT mediation of these effects. Neither manipulation significantly affected the depressant effect of fenfluramine on startle baseline, demonstrating that the baseline-reducing and prepulse inhibition-reducing effects of fenfluramine could be dissociated. MDL 28,618A alone did not affect prepulse inhibition or basal startle levels, demonstrating an important functional difference between pharmacologically induced 5-HT uptake blockade and 5-HT release. In summary, these data indicate that serotonergic overactivation can disrupt auditory and visual sensorimotor gating as measured using sound and light prepulse inhibition in rats. These data support a potential role of excessive 5-HT activity as a contributing factor to disrupted sensory gating processes seen in schizophrenia and possibly other neuropsychiatric disorders.

5-HT modulation of auditory and visual sensorimotor gating: II. Effects of the 5-HT2A antagonist MDL 100,907 on disruption of sound and light prepulse inhibition produced by 5-HT agonists in Wistar rats.

Increasing evidence suggests an important role of 5-HT, and 5-HT2A receptors in particular, in the etiology and treatment of schizophrenia. The prepulse inhibition paradigm is used as a model for sensorimotor gating processes that are disrupted in schizophrenia. The present study used the selective serotonin2A (5-HT2A) antagonist and putative antipsychotic agent MDL 100,907 to evaluate the contribution of 5-HT2A receptors to the disruptions of prepulse inhibition produced by several 5-HT agonists. The D2 antagonist haloperidol was used to evaluate a possible interaction with dopamine neurons. Sound or light prepulses were used to measure the generality of these drug effects on cross-modal prepulse inhibition. In the first study, MDL 100,907 antagonized the disruptions of auditory prepulse inhibition produced by the 5-HT releasing agents fenfluramine and 3,4-methylenedioxymethamphetamine (MDMA). These effects on prepulse inhibition were modality-specific in that MDL 100,907 did not reverse the effects of the 5-HT releasers on visual prepulse inhibition. Haloperidol did not alter the disruptive effects of MDMA or fenfluramine on either auditory or visual prepulse inhibition. In the second study, the direct acting 5-HT2A/2C receptor agonist/hallucinogen (+)1-4-iodo-2,5-dimethoxyphenyl-2-aminopropane (DOI) consistently disrupted auditory prepulse inhibition, and this effect was blocked by MDL 100,907 but not by haloperidol. A dose-response analysis demonstrated that MDL 100,907 potently antagonized DOI disrupted auditory prepulse inhibition, with an ED50 of 0.04 mg/kg, IP. DOI did not consistently disrupt visual prepulse inhibition. In summary, these data indicate that, at least under the conditions of the present studies, the disruptions of auditory prepulse inhibition produced by fenfluramine, MDMA, and DOI result from stimulation of 5-HT2A receptors. Furthermore, these disruptions do not involve direct or indirect stimulation of D2 receptors. The identity of the 5-HT receptor(s) underlying the disruptive effects of fenfluramine or MDMA on visual prepulse inhibition has not yet been identified. MDL 100,907 may be generally useful in CNS disorders in which excessive 5-HT2A receptor tone disrupts sensory gating processes.

Reversal of amphetamine-induced behaviours by MDL 100,907, a selective 5-HT2A antagonist.

MDL 100,907 is a potent and selective antagonist of the 5-HT2A receptor which, unlike other antagonists at this receptor, has little affinity for the 5-HT2C receptor. We have investigated the antipsychotic potential of MDL 100,907 by examining its ability to antagonise different behavioural effects of amphetamine in rats. MDL 100,907 reversed the locomotor stimulant effects of amphetamine in rats without itself having any effect on locomotor activity. It also antagonised the disruptive effects of amphetamine on the development of latent inhibition. In contrast, MDL 100,907 had no effect on the discriminative stimulus properties of amphetamine, nor did it affect the ability of amphetamine to reduce the threshold required to sustain rewarding brain stimulation in the ventral tegmental area. This profile is different from that of typical and atypical neuroleptics, and also from other 5-HT2 receptor antagonists, which lack the selectivity of MDL 100,907. These results suggest that MDL 100,907 may have a unique interaction with dopaminergic systems and support the further development of selective 5-HT2 receptor antagonists as a novel therapeutic strategy for schizophrenia.

Mixed D2/5-HT2A antagonism of amphetamine-induced facilitation of brain stimulation reward.

Recent experiments have demonstrated that 5-HT2A antagonists can modify electrophysiological, neurochemical, and behavioral responses to psychostimulants. These findings led to an interest in using 5-HT2A antagonists to block the effects of psychostimulants on brain reward mechanisms. The present experiments assessed the ability of mixed D2/5-HT2A antagonists to reverse amphetamine-induced facilitation of self-stimulation. The D2/5-HT2A antagonists MDL 28,133A and risperidone attenuated the effects of cocaine and amphetamine, but only at antagonist doses that elevated baseline self-stimulation thresholds. A comparison of the effects of the mixed antagonists to those of haloperidol and eticlopride revealed that all four antagonists produced similar anti-stimulant effects when the influence of the drugs on baseline responding was considered. The D2 activity of the antagonists appears to account for their ability to reduce the effects of psychostimulants on self-stimulation. 5-HT2A antagonism makes a negligible contribution to the anti-amphetamine effects.

MDL 100,458 and MDL 102,288: two potent and selective glycine receptor antagonists with different functional profiles.

Glycine receptor antagonists have been proposed to have multiple therapeutic applications, including the treatment of stroke, epilepsy, and anxiety. The present study compared the biochemical and behavioral profiles of two strychnine-insensitive glycine receptor antagonists, MDL 100,458 (3-(benzoylmethylamino)-6-chloro-1H-indole-2- carboxylic acid) and MDL 102,288 (5,7-dichloro-1,4-dihydro-4-[[[4- [(methoxycarbonyl)amino]phenyl]sulfonyl]imino]-2-quinolinecarboxylic acid monohydrate). Both compounds potently inhibited [3H]glycine binding to rat cortical/hippocampal membranes (Ki = 136, 167 nM, respectively) without showing significant activity in 18 other receptor binding assays. In an in vitro functional assay, both compounds completely antagonized N-methyl-D-aspartate (NMDA)-stimulated cGMP accumulation in rat cerebellar slices. However, in contrast to their equipotency in the glycine receptor assay, MDL 100,458 was approximately 6-fold more potent than MDL 102,288 in the cGMP assay (IC50 values = 1.25, 7.8 microM, respectively). Behavioral tests demonstrated that MDL 102,288 and MDL 100,458 exhibited strikingly different in vivo profiles. MDL 100,458 antagonized audiogenic seizures in DBA/2J mice (ED50 = 20.8 mg/kg i.p.), whereas MDL 102,288 was without effect in the dose range tested (ED50 > 300 mg/kg i.p.). Central nervous system penetration did not appear to account for this difference. For example, MDL 102,288 was not active following direct intracerebroventricular administration (ED50 > 16 micrograms; vs. 0.78 microgram for MDL 100,458). In a test of anxiolytic activity, MDL 102,288 reduced separation-induced ultrasonic vocalizations in rat pups (ED50 = 6.3 mg/kg i.p.) whereas MDL 100,458 was only weakly active (ED50 = 80.8 mg/kg i.p.). Furthermore, the anxiolytic effect of MDL 102,288 was selective in that it occurred at doses that did not produce motoric disruption as measured by an inclined-plane test (ED50 > 160 mg/kg; therapeutic index > 25.4). In contrast, the anxiolytic activity of MDL 100,458 was non-selective in that it occurred at doses that also produced motoric disruption (ED50 = 57.7 mg/kg; therapeutic index = 0.7). Thus, two glycine receptor antagonists which have similar in vitro binding profiles as selective ligands for the strychnine-insensitive glycine receptor, demonstrate different in vitro and in vivo functional profiles. The reason for these differences is not clear, though one possibility could be that the compounds may act on different NMDA receptor subtypes. These data support the possibility that different glycine receptor antagonists may have different therapeutic targets.

The role of 5-HT2A receptors in antipsychotic activity.

The correlation between the clinical activity of antipsychotic agents and their affinity for the D2 dopamine receptor has been the mainstay of the hypothesis that schizophrenia is due to excessive dopaminergic function. More recently, the unique clinical profile of the atypical antipsychotic clozapine has been proposed to involve actions on additional receptor systems. In particular, the high affinity of clozapine for the 5HT2A receptor subtype has been suggested to contribute to its reduced side-effect liability, greater efficacy and its activity in therapy-resistant schizophrenia. We have used the highly selective 5-HT2A antagonist MDL 100,907 to explore the contribution of 5-HT2A receptor blockade to antipsychotic activity. Biochemical, electrophysiological and behavioral studies reveal that selective 5HT2A receptor antagonists have the preclinical profile of an atypical antipsychotic. The limited clinical evidence available also suggests that compounds producing 5-HT2A receptor blockade are effective, in particular, against the negative symptoms of schizophrenia.

5-Aryl-3-(alkylthio)-4H-1,2,4-triazoles as selective antagonists of strychnine-induced convulsions and potential antispastic agents.

Selected examples from three series of isomeric (alkylthio)-1,2,4-triazoles were prepared and examined for anticonvulsant activity versus strychnine-, maximal-electroshock-, pentylenetetrazole-, and 3-mercaptopropionic-acid-induced seizures in mice. A number of 5-aryl-3-(alkylthio)-4H-1,2,4-triazoles were selective antagonists of strychnine-induced convulsions. The isomeric 3-aryl-5-(alkylthio)- and 5-aryl-3-(alkylthio)-1H-1,2,4-triazoles were essentially inactive as anticonvulsants. The most potent antagonist of strychnine-induced convulsions was 5-(2-fluorophenyl)-4-methyl-3-(methylthio)-4H-1,2,4-triazole (3s), while the most selective antagonist was 5-(3-fluorophenyl)-4-methyl-3-(methylsulfonyl)-4H-1,2,4-triazole (3aa). The anticonvulsant profiles of these 4H-1,2,4-triazoles suggested that they were acting functionally like glycine receptor agonists. Since it has recently been postulated that compounds possessing glycine-agonist-like properties might be useful in the treatment of spasticity, we examined 5-phenyl-4-methyl-3-(methylsulfonyl)-4H-1,2,4-triazole (3c) in an in vivo model of spasticity. In this regard, 3c reduced the occurrence of hyperreflexia in rats that had received spinal transections 5-10 weeks previously. While triazole 3c appeared to possess glycine-agonist-like properties in vivo, it did not displace [3H]strychnine binding from rat brain stem/spinal cord membranes in vitro. On the other hand, 3c enhanced muscimol-stimulated 36Cl influx in a rat cerebellar membrane preparation, indicating a possible interaction of these triazoles with the GABAA receptor.

Characterization of the 5-HT2 receptor antagonist MDL 100907 as a putative atypical antipsychotic: behavioral, electrophysiological and neurochemical studies.

Progress toward understanding the role of the 5-hydroxytryptamine (5-HT)2 receptor in the therapy for schizophrenia has been hampered by the lack of highly selective antagonists. We now report on the effects of MDL 100,907 [R(+)-alpha-(2,3-dimethoxyphenyl)-1- [2-(4-fluorophenylethyl)]-4-piperidine-methanol], a highly selective and potent 5-HT2 receptor antagonist, in behavioral, electrophysiological and neurochemical models of antipsychotic activity and extrapyramidal side-effect liability. In mice, MDL 100,907 blocked amphetamine-stimulated locomotion at doses that did not significantly affect apomorphine-stimulated climbing behavior. Neither MDL 100,907 nor clozapine reduced apomorphine-induced stereotypies or produced catalepsy in rats. MDL 100,907 blocked the slowing of ventral tegmental area (A10) dopaminergic neurons by amphetamine but, like clozapine, produced only small increases in the number of active substantia nigra zona compacta (A9) and A10 dopamine neurons after acute administration. When administered chronically, MDL 100,907 and clozapine selectively reduced the number of spontaneously active A10 neurons, whereas haloperidol reduced activity in both the A9 and A10 regions. Consistent with their acute effect on A9 and A10 activity, neither MDL 100,907 nor clozapine increased dopamine metabolism in the striatum or nucleus accumbens, whereas acute haloperidol accelerated dopamine turnover in both regions. The administration of the dopamine uptake blocker amfonelic acid with haloperidol produced a massive increase in DA metabolism characteristic of typical antipsychotics. In contrast, MDL 100,907 and clozapine were without effect on dopamine turnover when given in the presence of amfonelic acid. These data indicate that MDL 100,907 has a clozapine-like profile of potential antipsychotic activity with low extrapyramidal sid-effect liability.(ABSTRACT TRUNCATED AT 250 WORDS)