Antiplatelet agents are proven efficacious treatments for cardiovascular and cerebrovascular diseases. However, the existing drugs are compromised by unwanted and sometimes life-threatening bleeding that limits drug usage or dosage. There is a substantial unmet medical need for an antiplatelet drug with strong efficacy and low bleeding risk. Thrombin is a potent platelet agonist that directly induces platelet activation via the G protein (heterotrimeric guanine nucleotide-binding protein)-coupled protease-activated receptors PAR1 and PAR4. A PAR1 antagonist is approved for clinical use, but its use is limited by a substantial bleeding risk. Conversely, the potential of PAR4 as an antiplatelet target has not been well characterized. Using anti-PAR4 antibodies, we demonstrated a low bleeding risk and an effective antithrombotic profile with PAR4 inhibition in guinea pigs. Subsequently, high-throughput screening and an extensive medicinal chemistry effort resulted in the discovery of BMS-986120, an orally active, selective, and reversible PAR4 antagonist. In a cynomolgus monkey arterial thrombosis model, BMS-986120 demonstrated potent and highly efficacious antithrombotic activity. BMS-986120 also exhibited a low bleeding liability and a markedly wider therapeutic window compared to the standard antiplatelet agent clopidogrel tested in the same nonhuman primate model. These preclinical findings define the biological role of PAR4 in mediating platelet aggregation. In addition, they indicate that targeting PAR4 is an attractive antiplatelet strategy with the potential to treat patients at a high risk of atherothrombosis with superior safety compared with the current standard of care.

Potassium channels have been identified as targets for a large number of therapeutic indications. The ability to use a highthroughput functional assay for the detection and characterization of small-molecule modulators of potassium channels is very desirable. However, present techniques capable of screening very large chemical libraries are limited in terms of data quality, temporal resolution, ease of use, and requirements for specialized instrumentation. To address these issues, the authors have developed a fluorescence-based thallium flux assay. This assay is capable of detecting modulators of both voltageand ligand-gated potassium channels expressed in mammalian cells. The thallium flux assay can use instruments standard to most high-throughput screening laboratories, and using such equipment has been successfully employed to screen large chemical libraries consisting of hundreds of thousands

The present study demonstrated that large lesions of the amygdala disrupt the maintenance of reflex facilitation of the unconditioned nictitating membrane (NM) response and slow the acquisition of conditioned NM responses in rabbit. Before behavioral training, the central nucleus of the amygdala and adjacent areas were lesioned electrolytically. In the 1st experiment, the lesioned animals exhibited no reflex facilitation of the unconditioned NM response at conditioned stimulus (CS)-unconditioned stimulus (US) intervals of 125-8,000 ms. In the 2nd and 3rd experiments in which one CS-US interval (500 ms) was used, the lesions disrupted the maintenance of reflex facilitation but did not alter the facilitation exhibited in the 1st block of training. The lesions retarded the acquisition of conditioned NM responses when the 1000-Hz tone CS intensity was 65 dB but not when the intensity was 85 dB.

The administration of L-dihydroxyphenylalanine (L-DOPA) to patients with Parkinson's disease is known to produce acute effects that include the reduction of rigidity as well as delayed therapeutic actions involving the resumption of complex motor behavior. In order to examine the potential role of dopamine (DA) cell activity in mediating these responses, the effects of acute and repeated L-DOPA administration on the electrophysiological activity of the residual dopamine (DA) neurons were examined in rats that had received partial 6-hydroxydopamine (6-OHDA)-induced DA lesions. DA cell activity was assessed along three dimensions: (1) the relative proportion of DA neurons exhibiting spontaneous spike firing, (2) their basal firing rate, and (3) their firing pattern. Following 6-OHDA-induced DA depletion, rats were treated for 1 month with saline or L-DOPA. In addition, rats from each group received either an acute injection of L-DOPA or saline on the day of recording. In rats receiving repeated saline treatment, the DA neurons recorded following acute L-DOPA administration were firing at significantly slower basal firing rates and exhibited less burst firing when compared to saline-pretreated rats given acute saline. In contrast, DA cells recorded from rats that had received repeated L-DOPA administration for 4 weeks followed by an acute saline injection did not exhibit any significant differences from DA cells of intact control rats with respect to basal firing rate or firing pattern; however, there was a substantial increase in the proportion of DA neurons exhibiting spontaneous spike firing after correcting for 6-OHDA-induced cell loss. In addition, in rats receiving repeated L-DOPA treatment, the DA cells recorded following acute administration of L-DOPA showed significantly less of a reduction in firing rate when compared to the cells recorded following acute L-DOPA in the saline treatment group. These results show that: (1) acute L-DOPA administration appears to exert its actions by DA autoreceptor stimulation, whereas (2) repeated L-DOPA administration increases the proportion of spontaneously active DA neurons in partially lesioned rats. As a result, repeated L-DOPA administration would be expected to cause an increase in spike-dependent DA release as a consequence of the greater proportion of DA cells showing spontaneous activity. This may be the major factor underlying the delayed therapeutic benefits of L-DOPA therapy in the treatment of Parkinson's disease.

(S)-N-[1-(3-Morpholin-4-ylphenyl)ethyl]-3-phenylacrylamide (2) was synthesized as an orally bioavailable KCNQ2 potassium channel opener. In a rat model of migraine, 2 demonstrated significant oral activity in reducing the total number of cortical spreading depressions induced by potassium chloride.

BL-1249 [(5,6,7,8-tetrahydro-naphthalen-1-yl)-[2-(1H-tetrazol-5-yl)-phenyl]-amine] produced a concentration-dependent membrane hyperpolarization of cultured human bladder myocytes, assessed as either a reduction in fluorescence of the voltage-sensitive dye bis-(1,2-dibutylbarbituric acid)trimethine oxonol (EC 50 ϭ 1.26 Ϯ 0.6 M) or by direct electrophysiological measurement (EC 50 ϭ 1.49 Ϯ 0.08 M). BL-1249 also produced a membrane hyperpolarization of acutely dissociated rat bladder myocytes. Voltage-clamp studies in human bladder cells revealed that BL-1249 activated an instantaneous, noninactivating current that reversed near E K . The BL-1249-evoked outward K ϩ current was insensitive to blockade by glyburide, tetraethylammonium, iberiotoxin, 4-aminopyridine, apamin, or Mg 2ϩ . However, the current was inhibited by extracellular Ba 2ϩ

NMDA receptor hypofunction is hypothesized to contribute to cognitive deficits associated with schizophrenia. Since direct activation of NMDA receptors is associated with serious adverse effects, modulation of the NMDA co-agonists, glycine or D-serine, represents a viable alternative therapeutic approach. Indeed, clinical trials with glycine and D-serine have shown positive results, although concerns over toxicity related to the high-doses required for efficacy remain. Synaptic concentrations of D-serine and glycine are regulated by the amino acid transporter alanine serine cysteine transporter-1 (asc-1). Inhibition of asc-1 would increase synaptic D-serine and possibly glycine, eliminating the need for high-dose Schizophrenia is a complex neuropsychiatric disorder characterized by positive symptoms, negative symptoms, and cognitive deficits (Brown et al. 2010). While current antipsychotic medications are effective at alleviating positive symptoms, treatment of negative symptoms and cognitive deficits remain an unmet medical need. Cognitive deficits are hypothesized to results from an under-activation or hypofunction of NMDA receptors. This hypothesis is based on data in human subjects, demonstrating that the pharmacological blockade of NMDA receptors produces cognitive deficits that parallel those seen in schizophrenia patients (Malhotra et al. 1996;Adler et al. 1999). These findings are supported by pre-clinical data demonstrating that inhibition of NMDA receptor function disrupts measures of cognition (Jentsch et al. 1997a,b;Stefani and Moghaddam 2005). Based on these data, development of newer medications has largely focused on improving NMDA receptor function. However, since direct pharmacological activation of NMDA receptors leads to over-excitation and seizures, indirect modulation of NMDA receptors at one of the many regulatory sites may provide an attractive alternative to addressing NMDA receptor hypofunction.Activation of NMDA receptors requires the binding of both glutamate as an agonist and either glycine or D-serine as a coagonist. Recent clinical trials have shown beneficial effects of adjunctive D-serine and glycine treatment on both cognitive and negative symptom domains of schizophrenia (Javitt et al. 1994;Heresco-Levy et al. 1996 Abbreviations used: NMDA, N-methyl-D-aspartate; asc-1, Na + -independent alanine-serine-cysteine transporter 1; ASCT-1 and -2, Na + -dependent alanine-serine-cysteine transporter 1 and 2; LAT-1 and 2, large amino acid transporter; AIB, a-aminoisobutyric acid.

The formation of a reactive intermediate was found to be responsible for CYP3A4 metabolism-dependent inhibition (MDI) observed with (S)-N-[1-(3-morpholin-4-ylphenyl)ethyl]-3-phenyl-acrylamide (1). Structure-3A4 MDI relationship studies culminated in the discovery of a difluoro analogue, (S)-N-[1-(4-fluoro-3-morpholin-4-ylphenyl)ethyl]-3-(4-fluoro-phenyl)acrylamide (2), as an orally bioavailable KCNQ2 opener free of CYP3A4 MDI.