Specific activation of the μ opioid receptor (MOR) by endomorphin 1 and endomorphin 2

The recently discovered endomorphin 1 (Tyr‐Pro‐Trp‐Phe‐NH 2 ) and endomorphin 2 (Tyr‐Pro‐Phe‐Phe‐NH 2 ) were investigated with respect to their direct receptor‐binding properties, and to their ability to activate G proteins and to inhibit adenylyl cyclase in both cellular and animal models. Both tetrapeptides activated G proteins and inhibited adenylyl cyclase activity in membrane preparations from cells stably expressing the μ opioid receptor, an effect reversed by the μ receptor antagonist CTAP ( d ‐Phe‐Cys‐Tyr‐ d ‐Trp‐Arg‐Thr‐Pen‐Thr‐NH 2 ), but they had no influence on cells stably expressing the δ opioid receptor. To further establish the selectivity of these peptides for the μ opioid receptor, brain preparations of mice lacking the μ opioid receptor gene were used to study their binding and signalling properties. Endomorphin 2, tritiated by a dehalotritiation method resulting in a specific radioactivity of 1.98 TBq/mmol (53.4 Ci/mmol), labelled the brain membranes of wild‐type mice with a K d value of 1.77 n m and a B max of 63.33 fmol/mg protein. In membranes of mice lacking the μreceptor gene, no binding was observed, and both endomorphins failed to stimulate [ 35 S]guanosine‐5′‐O‐(3‐thio)triphosphate ([ 35 S]GTPγS) binding and to inhibit adenylyl cyclase. These data show that endomorphins are capable of activating G proteins and inhibiting adenylyl cyclase activity, and all these effects are mediated by the μ opioid receptors.