Autoradiographic localization of δ opioid receptors in the rat brain using a highly selective bis-penicillamine cyclic enkephalin analog

Abstract

A pioneering approach to the design of a ligand selective for the 8 opioid receptors was implemented by Mosberg et al. (1983) who demonstrated that the cyclic conformationally restricted enkephalin analog (2-D-penicillamine,5-D-penicillamine)enkephalin (DPDPE) displayed extremely high specificity toward the 8 opioid receptors. Recently Akiyama et al. (1985) provided the first in vitro characterization of tritium labeled DPDPE binding to 8 opioid receptors in the rat brain and neuroblastoma-glioma hybrid (NG 108-15) cells. In the present study we examined the light microscopic autoradiographic distribution of opioid receptors in the rat central nervous system employing the highly selective 8 agonist [3H]DPDPE (S.A.= 40 Ci /mmol , Amersham Corp, Arlington Heights, IL.). Male Sprague-Dawley rats (150-200 g) were killed by decapitation, and the brain was removed and coated with plastic embedding medium (OCT Compound, Lab-Tek Products) onto microtome chucks and frozen by immersion into liquid nitrogen. Saggital and coronal sections (20 #m) were cut, thawed and moun ted onto chromealum/gelatin coated glass slides and air dried at room temperature. Slide-mounted sections were preincubated for 15 min at 25°C in 50 mM TrisHCI buffer (pH 7.4 at 25°C) containing 5 mM MgC12, 2 m g /ml BSA, 20 g g / m l bacitracin and 100 mM NaC1, to reduce any endogenous opioids