CA2+ DEPENDENCE OF MOTILIDE-INDUCED CONTRACTIONS IN RABBIT DUODENAL MUSCLE STRIPS INVITRO

Recent studies suggested that certain erythromycin A (EM-A) derivatives are motilin receptor agonists. As proposed by Itoh they may be called "motilides". We have investigated the Ca2+-dependence of contractions induced by two potent motilides, ME-34 [de(N-methyl) 8,9-anhydroeryhtromycin A 6,9-hemiacetal] and EM-523 [de(N-methyl)-N-ethyl-8,9-anhydroerythromycin A 6,9-hemiacetal], in duodenal tissues and compared the results with those previously obtained with motilin. Isometric and isotonic contractile responses of isolated longitudinal muscle sheets from the rabbit duodenum were tested under normal, Ca2+-free and depolarizing conditions. Prior to stimulation with motilides, the maximal response to acetylcholine was recorded and all responses were always expressed as a percentage of this response. Both motilides induced contractions in normally polarized tissue, with an EC50 of 26 +/- 5 nM for ME-34 (n = 7), and 27 +/- 5 nM for EM-523 (n = 16) and maximal responses of respectively 88 +/- 4% and 80 +/- 3%. Like motilin, both compounds induced an 'extra'-contraction in depolarized tissues. The EM-523 response in 140 mM K+under isotonic conditions was 84 +/- 3% (n = 5) at 10(-5) M, with an EC50 that was shifted to 65 +/- 18 nM. Similar figures were obtained for ME-34. When Ca2+ was added to Ca2+-depleted strips, half-maximal Ca2+ values (in mM) were 1.10 +/- 0.11 (n = 9) for EM-523 and 1.13 +/- 0.12 (n = 3) for ME-34, as compared with 1.12 +/- 0.13 (n = 7) for motilin and 2.8 +/- 1.1 (n = 9) for K+. Both ME-34 and EM-523 also induced a transient contraction in Ca2+-free solutions under isometric conditions. The response to EM-523 (5 x 10(-6) M) was 49 +/- 15% (n = 4) after 3 min. A maximal EM-523 stimulation reduced a subsequent ACh response by 78 +/- 7%, whereas EM-523 and ME-34 could not induce a contraction after ACh. We conclude that motilides depend upon external Ca2+ to a similar extent to motilin. Like motilin, they are also able to mobilize intracellullar Ca2+ stores. Our results support the hypothesis that motilides act on motilin receptors.