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Computer modeling of carbachol docked in the human m 1 receptor binding pocket has been used to discover a series of carbamate and thiocarbamate chiral, conformationally restricted analogues of carbachol based on azabicyclo[2.2.1]heptan-3-ol. These molecules have been evaluated for affinity and efficacy at human muscarinic receptors (m 1 -ms) transfected into a CHO cell line. None of these compounds was selective in binding. Thiocarbamate analogues had greater affinity for the m 1 receptor subtype, but lower efficacy based on comparison of their ability to induce phosphoinositide (PI) turnover. Carbamate analogues had lower affinity for m 1 receptors than thiocarbamates and varied in efficacy from 10% to 100% of the carbachol response in phosphoinositide (PI) turnover. One of these analogues 3S,4R-azabicyclo[2.2.1]heptan-3-methylcarbamate, WAY-131256, (VI) has been characterized as an m 1 /m 2 agonist in vitro. (Vl) was equi-efficacious to the standard m i agonist, xanomeline (Phase III) in vivo in a scopolamine-impaired radial arm maze paradigm (MED 1 mg/kg, 5.88 mmol/kg for VI and MED 1 mg/kg, 3.55 mmoles/kg for xanomeline) and was approximately equal to xanomeline in an AF64A-impaired radial arm maze paradigm. Despite its lack of m 1 selectivity in vitro, in vivo experiments on (VI) indicated no significant effect on blood pressure or heart rate at 10 mg/kg (58.78 mmol/kg) (i.p.), and no peripheral side effects attributed to stimulation of either the m 2 or m 3 receptors (salivation, lacrimation, and chromodacryorrhea) up to doses of 30 mg/kg, 1 76.2 mmol/kg. These results may be explained by different receptor densities in various brain regions not accounted for in a transfected cell line or by metabolism of (VI) to a m 1 selective agonist in vivo.

A series of 2-amino-(phosphonoalkyl)-1H-benzimidazole-2-alkanoic acids was synthesized and evaluated for NMDA receptor affinity using a [3H]CPP binding assay. Functional antagonism of the NMDA receptor complex was evaluated in vitro using a stimulated [3H]TCP binding assay and in vivo by employing an NMDA-induced seizure model. Several compounds of the AP-6 type demonstrated potent and selective NMDA antagonistic activity both in vitro and in vivo. In particular, [R(-)]-2-amino-3-(5-chloro-1-phosphonomethyl-1H-benzoimidazol-2-yl)-propionic acid (1) displayed an IC(50) value of 7.1 nM in the [3H]CPP binding assay and an ED(50) value of 0.13 mg/kg (ip) in the NMDA lethality model. Compound 1, when administered intravenously as a single bolus dose of 3 mg/kg following permanent occlusion of the middle cerebral artery in the rat, reduced the volume of infarcted brain tissue by 45%. These results support a promising therapeutic potential for compound 1 as a neuroprotective agent.

In this report, a novel bioisostere of the alpha-amino acid, 3,4-diamino-3-cyclobutene-1,2-dione, has been incorporated into a series of compounds which are NMDA antagonists. These compounds, which are achiral and easily prepared, demonstrated good affinity at the NMDA receptor by their ability to displace [3H]CPP binding in vitro. In particular, the phosphonic acid 24 provided protection against NMDA-induced lethality in mice equivalent to 2-amino-7-phosphonoheptanoic acid (5). This was considered an encouraging result in lieu of the fact that 24, like 5, lacks the conformational rigidity of the more potent NMDA antagonists. In addition, analogs that incorporate the 1,2,4-oxadiazolidine-3,5-dione heterocycle of quisqualic acid and the unsaturation of kainic acid were prepared to explore selectivity at the non-NMDA receptor subtypes.

In an effort to develop an orally active, non-sedating H1-antagonist, compound Wy-49,051 was synthesized and examined in various in vivo and in vitro assays. In vitro, the compound possessed high affinity for H1 receptors (IC50 = 44 nM) and was found to competitively antagonize histamine-induced guinea pig ileum contraction. At a concentration of 1 × 10−10 M, Wy-49,051 shifted the histamine dose-response curve to the right in a parallel fashion. Wy-49,051 demonstrated H1 antagonism in a variety of in vivo tests. Wy-49,051 (s.c.) protected against histamine-induced death (guinea pig) in a dose-dependent manner and with a longer duration of action than terfenadine. In addition, Wy-49,051 effectively antagonized histamine-induced wheal formation (administered up to 18 hr prior to challenge) and was more potent than astemizole and chlorpheniramine. At a dose of 5 mg/kg (p.o.), Wy-49,051 produced less inhibition of in vivo brain [3H]pyrilamine binding than ebastine (5 mg/kg, p.o.), terfenadine (20 mg/kg, p.o.), and chlorpheniramine (100 mg/kg, p.o.). Importantly, Wy-49,051 did not exhibit H2-antagonist activity (as indicated by a lack of effect on gastric acid secretion in the pylorus-ligated rat) and did not affect histamine-induced changes in blood pressure and heart rate in the anesthetized guinea pig. Although Wy-49,051 displayed significant alpha1-receptor affinity in vitro (IC50 =8 nM), it did not alter blood pressure or heart rate in SHR rats at doses examined above its projected therapeutic dose for H1-receptor antagonism. Wy-49,051 did not affect carbachol-induced guinea pig ileum contraction and possessed little affinity for M2 muscarinic receptors (IC50 = 7.3 μ). Taken together with the previously mentioned in vivo [3H]pyrilamine binding data, these data indicate that Wy-49,051 is a long-acting, potent H1-antagonist with less potential for sedative and anticholinergic side effects than many common antihistamines including chlorpheniramine.

A series of 2-amino-(phosphonoalkyl)-1H-benzimidazole-2-alkanoic acids was synthesized and evaluated for NMDA receptor affinity using a [3H]CPP binding assay. Functional antagonism of the NMDA receptor complex was evaluated in vitro using a stimulated [3H]TCP binding assay and in vivo by employing an NMDA-induced seizure model. Several compounds of the AP-6 type demonstrated potent and selective NMDA antagonistic activity both in vitro and in vivo. In particular, [R(-)]-2-amino-3-(5-chloro-1-phosphonomethyl-1H-benzoimidazol-2-yl)-propionic acid (1) displayed an IC(50) value of 7.1 nM in the [3H]CPP binding assay and an ED(50) value of 0.13 mg/kg (ip) in the NMDA lethality model. Compound 1, when administered intravenously as a single bolus dose of 3 mg/kg following permanent occlusion of the middle cerebral artery in the rat, reduced the volume of infarcted brain tissue by 45%. These results support a promising therapeutic potential for compound 1 as a neuroprotective agent.

Rational drug design utilizing a receptor homology model of the human muscarinic M1 receptor led to the discovery of the highly potent (Ki = 2 nM), efficacious, and in vivo functionally-selective M1 agonist, WAY-132983.

The diazabicyclic amino acid phosphonate 15, [2-(8,9-dioxo-2,6-diazabicyclo[5.2.0]non-1(7)-en-2-yl)ethyl]phosphonic acid, was identified as a potent NMDA antagonist. It contains the alpha-amino acid bioisostere 3,4-diamino-3-cyclobutene-1,2-dione and an additional ring for conformational rigidity. Compound 15 was as potent as CGS-19755 (5) in the [3H]CPP binding assay, the stimulated [3H]TCP binding assay, and the NMDA-induced lethality model in mice. A single bolus dose of compound 15, administered intravenously following permanent occlusion of middle cerebral artery (MCA) in the rat, reduced the size of infarcted tissue by 57%. Structure-activity relationship (SAR) studies have indicated that the six- and eight-membered ring derivatives had diminished activity and that the two-carbon side chain length was optimum for NMDA receptor affinity. Substitution on the ring was found to be counterproductive in the case of sterically demanding dimethyl groups and of no consequence in the case of an H-bonding hydroxyl group. Replacement of the phosphonic acid group by either a carboxylic acid or a tetrazole group was unproductive. The potent bicyclic NMDA antagonists were synthesized efficiently by virture of their achiral nature and the ease of vinylgous amide formation from squaric acid esters. Compound 15, being a unique NMDA antagonist structural type with a favorable preclinical profile, may offer advantages over existing NMDA antagonists for the treatment of neurological disorders such as stroke and head trauma. Compound 15 is currently under clinical evaluation as a neuroprotective agent for stroke.

Les auteurs decrivent l'electrocardiogramme de la Daphnie enregistre a l'aide de microelectrodes. La forme des ondes varie. La frequence du cœur varie entre 3 et 9 battements par seconde, et l'amplitude entre 100 et 500 μV. Les effects de l'acide gamma-aminobutyrique (GABA) et de la picrotoxine ont ete enregistres. L'administration de GABA a diminue la frequence des battements, tandis que la picrotoxine a semble l'augmenter.