Opioid ligand binding sites in the spinal cord of the guinea-pig

The properties of opioid binding sites in membranes from the spinal cord of the guinea-pig were analyzed in experiments employing radiolabeled opioid ligands, selective or partially selective for μ, δ and k-type binding sites. Incubation was conducted at 37°C in a quasi-physiological modified Krebs medium, containing sodium and magnesium. The types of binding sites were discriminated on the basis of their affinities for [3H]-d-Ala2-MePhe4-Gly5-ol]enkephalin ([3H]DAGO), [3H-d-Ala2-d-Leu5-enkephalin, and [3H]ethyketocyclazocine and the relative potencies of the displacing ligands, DAGO, [d-Ser2-Leu-5]enkephalyl-Thr and tran-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzene-acetamide methanesulfonate hydrate (U50488H), which are selective for μ, δ and k type binding sites respectively. In membranes from whole spinal cord, k type sites comprised about 60%, μ about 30% and δ about 10% of the total of μ, δ and k binding sites. Binding sites of the μ type were also found in the lumbo-sacral region of guinea-pig spinal cord, in contrast to earlier reports of their absence from this tissue. Morphine showed a better than 500-fold selectivity for μ over k sites in spinal cord, while nalbuphine and (-)1-cyclopentyl-5-(1,2,3,4,5,6-hexahydro-8-hydroxy-3,6,11-trimethyl-2,6-methano-3-benzazocin-11-yl)3-pentanone methanesulfonate (WIN 44441-3) showed about a 10-fold selectivity for μ sites. The drug U50488H had about a 150-fold greater affinity for k than μ-type binding sites. © 1986

Opioid binding to rat and guinea-pig neural membranes in the presence of physiological cations at 37°C

We have identified mu, delta and kappa opioid binding sites in four types of neural membranes under conditions which include physiological concentrations of ions and an incubation temperature of 37°C. We hypothesize that binding parameters determined under these conditions should be more directly comparable with physiological experiments than parameters obtained under conditions of low ionic concentration and at low temperature. By using either a radioligand which is selective for a single type of opioid binding site or a relatively nonselective radioligand in the presence of an unlabeled selective ligand, we have isolated binding to single populations of sites. Saturation and displacement data were analyzed with the aid of a computerized nonlinear curve fitting program. [3H]Tyr-D-Ala-Gly-(Me)Phe-Gly-ol bound to a single population of sites with the characteristics of mu receptors, as determined by saturation and displacement analysis. Binding to the mu site represented 70% of the total specific opioid binding in rat brain, but only 20 to 30% in guinea pig tissues. [3H][D-Ala2-D-Leu5]enkephalin bound almost equally well to mu and delta sites, but the delta site could be examined by the inclusion of unlabeled Tyr-D-Ala-Gly-(Me)Phe-Gly-ol in the incubations. [3H]Ethylketocyclazocine bound mu and kappa sites, and Tyr-D-Ala-Gly-(Me)Phe-Gly-ol was also used to block the mu component in experiments in which we studied kappa binding. Binding to kappa sites represented 50 to 60% of the total in guinea pig tissues, but less than 20% in rat brain

International comparison of guarded hot plate apparatus using national and regional reference materials

Peer reviewed: NoNRC publication: Ye