In view of recent pharmacological studies suggesting the existence of delta-kappa opioid receptor heterodimers/oligomers in the spinal cord, we have synthesized and evaluated (intrathecally in mice) a series of bivalent ligands (KDN series) containing kappa and delta antagonist pharmacophores. Pharmacological and binding data have provided evidence for the bridging of spinal delta-kappa receptor heterodimers by KDN-21 and for their identification as delta(1) and kappa(2). The selectivity profile of KDN-21 and the apparent absence of coupled delta(1)-kappa(2) phenotypes in the brain suggest a new approach for targeting receptors.

To characterize delta- and kappa-opioid receptor phenotypes, bivalent ligands (KDAN series) containing delta-antagonist (naltrindole) and kappa(1)-agonist (ICI-199,441) pharmacophores were synthesized and evaluated by the intrathecal route using the mouse tail-flick assay and binding studies. The data have suggested that KDAN-18 (2) bridges phenotypic delta(2)- and kappa(1)-receptors. A conceptual model is presented to explain the organizational differences between the opioid receptors that give rise to the phenotypes (delta(1), delta(2), kappa(1), kappa(2)).

KDN21 is a bivalent ligand that contains ␦ and opioid antagonist pharmacophores linked through a 21-atom spacer. It has been reported that KDN21 bridges ␦ and receptors that are organized as heterodimers. We have shown previously that when using [ 3 H]diprenorphine as radioligand, KDN21 displayed greatly enhanced affinity in this series for coexpressed ␦ and opioid receptors (CDK). The present study used in vitro expression systems to investigate interactions of members of the KDN series with ␦-heterodimers through competition binding using selective ligands and the mitogen-activated protein kinase (MAPK) assay. In this regard, the use of the selective radioligands [ 3 H]naltrindole and [ 3 H]norbinaltorphimine (nor-BNI) in competition binding studies revealed that KDN21 has much higher affinity than other KDN members for CDK and bound to CDK more selectively relative to mixed ␦ and opioid receptors or singly expressed ␦ and opioid receptors. Other experiments revealed that the binding of naltrindole to ␦ opioid receptors could increase the binding of nor-BNI to opioid receptors and vice versa, suggesting reciprocal allosteric modulation of receptors in the heterodimer. Regarding the selectivity of KDN21 for phenotypic ␦ and opioid receptors, we investigated the effect of KDN21 on the activation of MAPKs [extracellular signal-regulated kinases 1 and 2 (ERK1/2)] by ␦-or -selective agonists.

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