Binding of the opiate-like pentapeptide methionine-enkephalin to a particulate fraction from rat brain

Abstract The characteristics of stereospecific binding of [ 3 H] met-enkephalin (15 Ci/mmole) were studied in a particulate fraction from rat brain. The binding assay was performed for 70 min at 0°C and the bound radioactivity separated by filtration through glass fiber filters (Whatman, GF C ). In the absence of sodium, binding of [ 3 H] met-enkephalin could be described on the basis of two independent binding sites with apparent K D s of 2.1 and 53 nM, respectively. The data are also consistent with one class of binding sites showing negative cooperativity. In the presence of 100 mM NaCl, binding of [ 3 H] met-enkephalin was 90–95% reduced, thus indicating the agonist properties of the peptide. The highly stereospecific binding of [ 3 H] met-enkephalin was evidenced by the 10,000-fold greater potency of levorphanol than its analgesically inactive enantiomer dextrorphan to compete for [ 3 H] met-enkephalin binding. Similar conclusions could be reached using levallorphan, (+)-3-hydro-N-allyl-morphinan, (−) methadone and (+) methadone. The apparent affinity of various opiate agonists and antagonists for the binding sites was closely correlated with their known pharmacological activity.