Pharmacological activities of optically pure enantiomers of the κ opioid agonist, U50,488, and its cis diastereomer: evidence for three κ receptor subtypes

De Costa et al. (FEBS Lett. 223, 335; 1987) recently described the synthesis of optically pure enantiomers of (±)-trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide (U50,488). In the present study we examined the in vitro opioid receptor selectivity of (-)-(1S,2S)-U50,488, (+)-(1R,2R)-U50,488 and (±)-cis-3,4-di-chloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide (the cis diasteromers of U50,488), as well as their pharmacological activities in rhesus monkeys. Using [³H]5α,7α,8β-(-)-N-methyl-N-[7-1-pyrrolidinyl)-1-oxaspiro (4,5)dec-8-yl]-phenyl-benzeneacetamide([³H]U69,593) to label κ binding sites of guinea pig membranes, the apparent dissociation constants of the enantiomers hadU50,488 were 0.89 and 299 nM, for the (S,S) and (R,R) enantiomers, respectively. The (-)-cis and (+)-cis diastereomers had apparent K_ds of 167 and 2715 nM, respectively. Binding surface analysis of the interaction of (-)-(1S,2S)-U50,488 with κ binding sites labeled by [³H]bremazocine resolved two binding sites at which (-)-(1S,2S)-U50,488 had K_ds of 30 and 10485 nM, respectively. The (±)-cis, (-)-cis and (+)-cis diastereomers of U50,488 (1 μM) did not inhibit [³H]bremazocine binding. Rhesus monkeys were trained to discriminate ethylketocyclazocine (EKC) and saline. All compounds tested substituted completely for EKC. The order of potency was (-)-(1S,2S)-U50,488 > (±)-U50,488 > (±)-cisdiastereomer of U50,488 > (+)-(1R,2R)-U50,488. In tests of analgesia, (-)-(1S,2S)-U50,488 was 2-4 times more potent than (±)-U50,488, while (±)-cis diastereomer of U50,488 and (+)-(1R,2R)-U50,488 were inactive at the highest doses tested (32 mg/kg). Taken collectively, these data indicate that the pharmacologically active enantiomer of U50,488 is (-)-(1S,2S)-U50,488, and provide preliminary evidence for three subtypes of κ binding sites in guinea pig brain.