RESUMEN
We employed an ex vivo [(3)H]rolipram binding experiment to elucidate the mechanism of emetic activity of phosphodiesterase 4 inhibitors. In Suncus murinus (an insectivore used for evaluation of emesis), emetic potential as well as ability to occupy the high-affinity rolipram binding site in brain membrane fraction in vivo were determined for phosphodiesterase 4 inhibitors. In vitro, [(3)H]rolipram bound to the membrane fraction of S. murinus brain with high affinity and its value was comparable to that for rat brain (K(d)=3.6 nM and 3.5 nM, respectively). The test compounds included denbufylline, rolipram, piclamilast, CDP840 and KF19514, each of which possessed similar affinities for the rolipram binding sites in both S. murinus and rat brain. In S. murinus, these compounds induced emesis via intraperitoneal administration. Their ED(50) values were as follows: denbufylline (1.4 mg/kg), rolipram (0.16 mg/kg), piclamilast (1.8 mg/kg), CDP840 (20 mg/kg), and KF19514 (0.030 mg/kg). In addition, these compounds occupied the high-affinity rolipram binding site in vivo as detected by dose-dependent reduction in capacity of ex vivo [(3)H]rolipram binding in brain membrane fractions. A clear correlation was observed between dose required to induce emesis and that to occupy the high-affinity rolipram binding site for individual phosphodiesterase 4 inhibitors. We conclude that the emetic effect of phosphodiesterase 4 inhibitors is caused at least in part via binding to the high-affinity rolipram binding site in brain in vivo.
Asunto(s)
Encéfalo/efectos de los fármacos , Inhibidores de Fosfodiesterasa 4 , Rolipram/toxicidad , Vómitos/inducido químicamente , Animales , Benzamidas/administración & dosificación , Benzamidas/toxicidad , Sitios de Unión , Unión Competitiva/efectos de los fármacos , Encéfalo/metabolismo , Relación Dosis-Respuesta a Droga , Inyecciones Intraperitoneales , Cinética , Masculino , Estructura Molecular , Naftiridinas/química , Naftiridinas/toxicidad , Inhibidores de Fosfodiesterasa/administración & dosificación , Inhibidores de Fosfodiesterasa/química , Inhibidores de Fosfodiesterasa/toxicidad , Piridinas/administración & dosificación , Piridinas/química , Piridinas/toxicidad , Ensayo de Unión Radioligante , Ratas , Ratas Sprague-Dawley , Rolipram/administración & dosificación , Rolipram/metabolismo , Musarañas , Tritio , Xantinas/administración & dosificación , Xantinas/toxicidadRESUMEN
1. We previously reported a presynaptic facilitatory action of A(2A) receptors on GABAergic synaptic transmission in the rat globus pallidus (GP). In the present study we identify the intracellular signalling mechanisms responsible for this facilitatory action of A(2A) receptors, using biochemical and patch-clamp methods in rat GP slices. 2. The adenosine A(2A) receptor selective agonist CGS21680 (1, 10 microM) and the adenylyl cyclase activator forskolin (1, 10 microM) both significantly increased cyclic AMP accumulation in GP slices. The CGS21680 (1 microM)-mediated increase in cyclic AMP was inhibited by the A(2A) receptor selective antagonist KF17837 (10 microM). 3. In an analysis of miniature inhibitory postsynaptic currents (mIPSCs), forskolin (10 microM) increased the mIPSC frequency without affecting their amplitude distribution, a result similar to that previously reported with CGS21680. 4. The adenylyl cyclase inhibitor 9-(tetrahydro-2-furanyl)-9H-purin-6-amine (SQ22,536, 300 microM) abolished the CGS21680-induced enhancement in the frequency of mIPSCs. 5. H-89 (10 microM), a selective inhibitor for cyclic AMP-dependent protein kinase (PKA), blocked the CGS21680-induced enhancement of the mIPSC frequency. 6. The calcium channel blocker CdCl(2) (100 microM) did not prevent CGS21680 from increasing the frequency of mIPSCs. 7. These results indicate that A(2A) receptor-mediated potentiation of mIPSCs in the GP involves the sequential activation of the A(2A) receptor, adenylyl cyclase, and then PKA, and that this facilitatory modulation could occur independently of presynaptic Ca(2+) influx.
Asunto(s)
AMP Cíclico/fisiología , Globo Pálido/metabolismo , Receptores Presinapticos/metabolismo , Receptores Purinérgicos P1/metabolismo , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/metabolismo , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Animales , Globo Pálido/efectos de los fármacos , Técnicas In Vitro , Masculino , Agonistas del Receptor Purinérgico P1 , Ratas , Ratas Sprague-Dawley , Receptor de Adenosina A2A , Receptores Presinapticos/agonistasRESUMEN
We investigated the biochemical and pharmacological properties of a new adenosine A(3) receptor antagonist, KF26777 (2-(4-bromophenyl)-7,8-dihydro-4-propyl-1H-imidazo[2,1-i]purin-5(4H)-one dihydrochloride). This compound was characterized using N(6)-(4-amino-3-iodobenzyl)adenosine-5'-N-methyluronamide ([125I]AB-MECA) or [35S]guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) binding to membranes from human embryonic kidney 293 (HEK293) cells expressing human adenosine A(3) receptors. KF26777 showed a K(i) value of 0.20+/-0.038 nM for human adenosine A(3) receptors labeled with [125I]AB-MECA and possessed 9000-, 2350- and 3100-fold selectivity vs. human adenosine A(1), A(2A) and A(2B) receptors, respectively. The inhibitory mode of binding was competitive. KF26777 inhibited the binding of [35S]GTPgammaS stimulated by 1 microM 2-chloro-N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide (Cl-IB-MECA). The IC(50) value was 270+/-85 nM; the compound had no effect on basal activity. Dexamethasone treatment for HL-60 cells, human promyelocytic leukemia, up-regulated functional adenosine A(3) receptors expression, and resulted in the enhanced elevation of intracellular Ca(2+) concentration ([Ca(2+)](i)) via the adenosine A(3) receptor. KF26777 antagonized this [Ca(2+)](i) mobilization induced by Cl-IB-MECA, with a K(B) value of 0.42+/-0.14 nM. These results indicate that KF26777 is a highly potent and selective antagonist of the human adenosine A(3) receptor.
Asunto(s)
Antagonistas de Receptores Purinérgicos P1 , Purinas/farmacología , Receptores Purinérgicos P1/metabolismo , Animales , Células CHO , Línea Celular , Cricetinae , Células HL-60 , Humanos , Receptor de Adenosina A3RESUMEN
The synthesis of a series of 4-(6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolin-2-yl)methyl-2-arylbenzofuran and 4-(6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolin-2-yl)methylbenzofuran-2-carboxamide derivatives as novel alpha(2C)-adrenergic receptor antagonists are described. Their affinity at three different human alpha(2)-adrenergic receptors is reported, and some of these compounds exhibited high affinity for the alpha(2C)-adrenergic receptor with high subtype selectivity. Among them, compound 10e has been found to show the anti-L-dopa-induced dyskinetic activity in marmosets. The structure-activity relationship of these compounds is also discussed.