The effects of endomorphin‐1 and endomorphin‐2 in CHO cells expressing recombinant μ‐opioid receptors and SH‐SY5Y cells

Endomorphin‐1 and ‐2 (E‐1/E‐2) have been proposed as endogenous ligands for the μ‐opioid receptor. The aims of this study are to characterize the binding of E‐1/E‐2 and the subsequent effects on cyclic AMP formation and [Ca 2+ ] i levels in SH‐SY5Y and Chinese hamster ovary (CHO) cells expressing endogenous and recombinant μ‐opioid receptors. E‐1 displaced [ 3 H]‐diprenorphine ([ 3 H]‐DPN) binding in CHOμ and SH‐SY5Y membranes with pK i values of 8.02±0.09 and 8.54±0.13 respectively. E‐2 displaced [ 3 H]‐DPN binding in CHOμ and SH‐SY5Y cells with pK i values of 7.82±0.11 and 8.43±0.13 respectively. E‐1/E‐2 bound weakly to CHOδ and CHOκ membranes, with IC 50 values of greater than 10 μ M . In CHOμ cells, E‐1/E‐2 inhibited forskolin (1 μ M ) stimulated cyclic AMP formation with pIC 50 values of 8.03±0.16 (I max =53.0±9.3%) and 8.15±0.24 (I max =56.3±3.8%) respectively. In SH‐SY5Y cells E1/E2 inhibited forskolin stimulated cyclic AMP formation with pIC 50 values of 7.72±0.13 (I max =46.9±5.6%) and 8.11±0.31 (I max =40.2±2.8%) respectively. E‐1/E‐2 (1 μ M ) increased [Ca 2+ ] i in fura‐2 loaded CHOμ cell suspensions in a thapsigargin sensitive and naloxone reversible manner. Mean increases observed were 106±28 and 69±6.7 n M respectively. In single adherent cells E‐1/E‐2 (1 μ M ) increased [Ca 2+ ] i with a mean 340/380 ratio change of 0.81±0.09 and 0.40±0.08 ratio units respectively. E‐1/E‐2 failed to increase intracellular calcium in CHOδ, CHOκ and SH‐SY5Y cells. These data show that E‐1/E‐2 bind with high affinity and selectivity to μ‐opioid receptors and modulate signal transduction pathways typical of opioids. This provides further evidence that these two peptides may be endogenous ligands at the μ‐opioid receptor.British Journal of Pharmacology (1999) 128 , 472–478; doi: 10.1038/sj.bjp.0702798