n(6)-cyclopentyladenosine and 8-(4-((2-aminoethyl)aminocarbonylmethyloxy)phenyl)-1-3-dipropylxanthine

Although it is well established that adenosine exerts antinociceptive effects at the spinal level in various species including human, the mechanisms responsible for such effects are still a matter of debate. We presently investigated whether adenosine-induced antinociception might possibly be related to an inhibitory influence of this neuromodulator on the spinal release of neuropeptides implicated in the transfer and/or control of nociceptive signals. For this purpose, the K(+)-evoked overflow of substance P-, calcitonin gene-related peptide (CGRP)- and cholecystokinin-like materials was measured from slices of the dorsal half of the rat lumbar enlargement superfused with an artificial cerebrospinal fluid supplemented with increasing concentrations of various adenosine receptor ligands. The data showed that stimulation of adenosine A(1) and (possibly) A(3) receptors, but not A(2A) receptors, exerted an inhibitory influence on the spinal release of CGRP-like material. In contrast, none of the adenosine A(1), A(2A) and A(3) receptor agonists tested within relevant ranges of concentrations significantly affected the release of substance P- and cholecystokinin-like materials. These results support the idea that adenosine-induced antinociception at the spinal level might possibly be caused, at least partly, by the stimulation of inhibitory adenosine A(1) receptors located presynaptically on primary afferent fibres containing CGRP but not substance P.

Topics: Adenosine; Adenosine-5'-(N-ethylcarboxamide); Animals; Calcitonin Gene-Related Peptide; Cholecystokinin; Dose-Response Relationship, Drug; In Vitro Techniques; Male; Neuropeptides; Pain; Phenethylamines; Potassium; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P1; Spinal Cord; Substance P; Xanthines

Four adenosine receptor subtypes of the family of G protein-coupled receptors, designated A1, A2A, A2B and A3 are currently known. In this study all human subtypes were stably transfected into Chinese hamster ovary (CHO) cells in order to be able to study their pharmacological profile in an identical cellular background utilizing radioligand binding studies (A1, A2A, A3) or adenylyl cyclase activity assays (A2B). The A1 subtype showed the typical pharmacological profile with 2-chloro-N6-cyclopentyladenosine (CCPA) as the agonist with the highest affinity and a marked stereoselectivity for the N6-phenylisopropyladenosine (PIA) diastereomers. In competition with antagonist radioligand biphasic curves were observed for agonists. In the presence of GTP all receptors were converted to a single low affinity state indicating functional coupling to endogenous G proteins. For A2A adenosine receptors CGS 21680 (2-[p-(2-carboxyethyl)phenylethylamino]-5'-N-ethylcarboxamidoadeno sine) and N-ethylcarboxamidoadenosine (NECA) were found to be the most potent agonists followed by R- and S-PIA with minor stereoselectivity. The relative potencies of agonists for the A2B adenosine receptor could only be tested by measurement of receptor-stimulated adenylyl cyclase activity. NECA was the most potent agonist with an EC50-value of 2.3 microM whereas all other compounds tested were active at concentrations in the high micromolar range. Inhibition of NECA-stimulated adenylyl cyclase identified xanthine amino congener (XAC; 8-[4-[[[[(2-aminoethyl)amino]-carbonyl]methyl]oxy]phenyl]-1,3-dipropylxa nthine) as the most potent antagonist at this receptor subtype. The A3 receptor was characterized utilizing the nonselective agonist [3H]NECA. The N6-benzyl substituted derivatives of adenosine-5'-N-methyluronamide (MECA) turned out to be the most potent agonists. The notion of xanthine-insensitivity of the A3 receptor should be dropped at least for the human receptor as xanthines with submicromolar affinity were found. Overall, the pharmacological characteristics of the human receptors are similar to other species with some species-specific characteristics. In this study we present for the first time the comparative pharmacology of all known human adenosine receptor subtypes. The CHO cells with stably transfected adenosine receptors provide an identical cellular background for such a pharmacological characterization. These cells are valuable systems for further characterization of specifi

Topics: Adenosine; Adenosine-5'-(N-ethylcarboxamide); Animals; Base Composition; Binding, Competitive; CHO Cells; Cricetinae; Guanylate Cyclase; Humans; Phenethylamines; Phenylisopropyladenosine; Receptors, Purinergic P1; Stereoisomerism; Structure-Activity Relationship; Transfection; Xanthines