Biochemical characterization of high-affinity 3H-opioid binding. Further evidence for Mu1 sites

SL Nishimura, LD Recht and GW Pasternak

In saturation studies with [3H]dihydromorphine, unlabeled D-Ala2-D-Leu5- enkephalin (1 nM) inhibited the high-affinity binding component far more potently than the lower-affinity one. Similarly, morphine (1 nM) inhibited the higher-affinity binding of 3H-D-Ala2-D-Leu5-enkephalin to a greater extent than its lower-affinity binding component, consistent with a common high-affinity binding site for opiates and enkephalins. Treatment of tissue with either trypsin (1 microgram/ml) or N- ethylmaleimide (25 microM) effectively eliminated the high-affinity binding component of a series of 3H-opiates and opioid peptides. Competition studies following both treatments were consistent with a common high-affinity binding site. Both treatments also eliminated the ability of low morphine concentrations (less than 1 nM) to inhibit 3H-D- Ala2-D-Leu5-enkephalin binding and of low D-Ala2-D-Leu5-enkephalin concentrations (less than 1 nM) to inhibit [3H]dihydromorphine binding. Protection experiments examining N-ethylmaleimide (25 microM) inhibition of [3H]dihydromorphine binding showed significant protection (p less than 0.002) by both unlabeled D-Ala2-D-Leu5-enkephalin and morphine (both at 1 nM). When studied together, both naloxonazine and N- ethylmaleimide inhibited [3H]dihydromorphine binding to a similar extent. Equally important, tissue previously treated with naloxonazine was far less sensitive to N-ethylmaleimide than was untreated control tissue, consistent with the possibility that both treatments affected the same site. Together, these results support the concept of a common high-affinity binding site for opiates and opioid peptides.

Volume 25, Issue 1, pp. 29-37, 01/01/1984
Copyright © 1984 by American Society for Pharmacology and Experimental Therapeutics

This article has been cited by other articles:

				Y. Jinsmaa, Y. Okada, Y. Tsuda, K. Shiotani, Y. Sasaki, A. Ambo, S. D. Bryant, and L. H. Lazarus Novel 2',6'-Dimethyl-L-Tyrosine-Containing Pyrazinone Opioid Mimetic {micro}-Agonists with Potent Antinociceptive Activity in Mice J. Pharmacol. Exp. Ther., April 1, 2004; 309(1): 432 - 438. [Abstract] [Full Text]

				G. Ling, J. MacLeod, S Lee, S. Lockhart, and G. Pasternak Separation of morphine analgesia from physical dependence Science, October 26, 1984; 226(4673): 462 - 464. [Abstract] [PDF]