The beta3-adrenoceptor agonist 4-[[(hexylamino)carbonyl]amino]-N-[4-[2-[[(2S)-2-hydroxy-3-(4-hydroxyphenoxy)propyl]amino]ethyl]-phenyl]-benzenesulfonamide (L755507) and antagonist [...]

This study identifies signaling pathways activated by the beta2-/beta3-AR agonist zinterol, the selective beta3-AR agonist L755507 and the selective beta3-AR antagonist L748337 in CHO-K1 cells expressing human beta3-adrenoceptors. Zinterol and L755507 caused a robust concentration-dependent increase in cAMP accumulation (pEC50 8.5 and 12.3 respectively), while L748337 had low efficacy. Maximal cAMP accumulation with zinterol and L755507 was increased after pre-treatment with pertussis toxin, indicating that the human beta3-AR couples to Gi as well as Gs. In contrast to cAMP, zinterol, L755507 and L748337 increased phosphorylation of Erk1/2 with very high potency (pEC50 10.9, 11.7 and 11.6). These compounds also stimulated phosphorylation of p38 MAPK, but with much lower potency than Erk1/2 (pEC50 5.9, 5.5 and 5.7 respectively). Pertussis toxin completely blocked Erk1/2 and p38 MAPK phosphorylation in response to L748337, demonstrating a requirement for Gi/o coupling, whereas L755507-stimulated p38 MAPK phosphorylation was not inhibited by pertussis toxin, and Erk1/2 phosphorylation was inhibited by only 30 . We found that high levels of cAMP interfered with agonist-activated p38 MAPK phosphorylation. L748337 increased extracellular acidification rate (ECAR) in the cytosensor microphysiometer with efficacy similar to zinterol and L755507, albeit with lower potency (pEC50 7.2 compared to zinterol, 8.1, and L755507, 8.6). The ECAR response to L748337 was largely via activation of p38 MAPK, demonstrated by 65 inhibition with RWJ67657. We conclude that the beta3-AR agonist L755507 couples to both Gs and Gi to activate adenylate cyclase and MAPK signaling, whereas the beta3-AR antagonist L748337 couples predominantly to Gi to activate MAPK signaling.