Eurt,pean Journal of Pharmacology, 200 ( 1991 ) 361-364 ~3 1991 Elsevier Science Publishers B.V. 0014-2999/91/$03.50 ADONIS 001429999100560G 2~i EJP 20882 Short communication Influence of thiorphan and phosphoramidon on the relaxant effect of vasoactive intestinal polypeptide and non-adrenergic non-cholinergic neurone stLmulation in the rat gastric fundus Geert J. Smits and Romain A. Lefebvre Heymans h~stitute of Pharmacology, Unicersity of Gent Medical School. De Pintelaan 185, B-9000 Gent, Belgium Received 29 May 1991, accepted 18 June 1991 Relaxations were induced in longitudinal muscle strips of the rat gastric fundus by exogenous administration of vasoactive intestinal polypeptide (VIP) and by transmural stimulation in the presence of atropine. These responses were not influenced by two neutral endopeptidase inhibitors, thiorphan (3 × l0 -5 M) and phosphoramidon (10 -5 M). This suggests that neutral endopeptidase is not involved in the breakdown of exogenous VIP and non-adrenergic non-cholinergic inhibitory transmitters of the rat gastric fundus. Gastric fundus (rat); Non-adrenergic, non-cholinergic (NANC) neurotransmitzers; VIP (vasoactive intestinal peptide); Neutral endopeptidase; Thiorphan; Phosphoramidon 1. Introduction Tissue degradation of peptides by proteases may influence their effect. For example, in isolated guinea- pig trachea, both the contractile effect of substance P and the relaxant effect of vasoactive intestinal polypep- tide (VIP) are potentiated by thiorphan and phospho- ramidon, two inhiUitors of neutral endopeptidase (NEP; EC 3.4.24.11; Stimler-Gerard, 1987; Farmer and Togo, 19901. NEP is a widely dispersed membrane-bound enzyme, cleaving peptides at the amino group of hy- drophobic residues. VIP has been proposed as an inhibitory non-adrenergic non-cholinergic (NANC) neurotransmitter in the rat gastric fundus (De Beurme and Lefebvre, 1988). We therefore investigated the influence of thiorphan and phosphoramidon on the relaxation induced by V|P and transmural stimulation of the inhibitory NANC neurones in the rat gastric fundus. The influence of thiorphan on the substance P-elicited contractions in the guinea-pig gallbladder and in the rat gastric fundus was studied as control. Correspondence to: R.A. Lefebvre, Heymans Institute of Pharmacol- ogy, University of Gent Medical School, De Pintelaan 185, B-9000 Gent, Belgium. Tel. 32.91.40.33.97, fax 32.91.40.49.88. 2. Materials and methods Rats (either sex, 200-300 g) were reserpinized (5 mg/kg i.p.) 24 h before killing and fasted from then on. The gastric fundus was rapidly dissected out and two longitudinal muscle strips (15 mm long × 3 mm wide) were prepared; the mucosa was removed in some experiments. The strips were mounted under a load of l g between two parallel platinum electrodes (25 mm long × 9 mm wide) in 5-ml organ baths. The bathing medium, Krebs solution (composition in raM: NaCl 118.5, KCI 4.8, CaC! 2 1.9, KH2PO 4 1.2, MgSO4 1.2, NaHCO 3 25.0 and glucose 10.11 containing 10 -6 M atropine, was maintained at 37°C and bubbled with 95% 0 2 and 5% CO 2. Strips were equilibrated for 1 h, with rinsing every 15 rain and changes in length were recorded isotonically (Hugo Sachs Lever transducer B type 368) on a Graphtec Linearcorder mark 8 WR 3500. Electrical stimulation of the strips (supramaximal voltage, 1 ms duration) was done with a Grass $88 stimulator over a constant voltage unit. Tone was raised by means of 10 -6 M prostaglandin F2~ (PGF:a) and VIP (10-~°-10 -7 M) added cumula- tively or of transmural stimulation (supramaximal volt- age, 1 ms duration) performed at cumulatively increas- ing frequencies (0.25-8 Hz). After rinsing for at least 40 rain, the tissue was incubated with thiorphan (3 × 10 -5