PTH-043 Characterisation of motor nerve responses to electrical field stimulation in human isolated colon

Abstract

Introduction The ability of drugs to affect gastrointestinal (GI) motility may be predicted in vitro by studying how they modify changes in muscle tension evoked by motor nerve activation with electrical field stimulation (EFS). In human tissues, variations in types of response to EFS make the technique difficult unless only one type is studied, ignoring the others.1 This study aimed to characterise the variations so all tissues may be used. Methods Human ascending/ transverse colon were obtained at surgery for cancer, following informed consent. After removing the mucosa, 20 strips (∼4 mm wide, 10 mm long) were cut parallel to the circular muscle from each colon. They were suspended between 2 ring electrodes in tissue baths (Kreb9s; 5% CO 2 in O 2 ; 37°C; 1 g tension) for isometric recording. EFS was applied (5 Hz, 0.5 ms pulse width, 50 V, 10 s) every 1 min. Results Responses to EFS in strips from the same patient were inconsistent but prevented by tetrodotoxin 1 μM (n=24 strips; four patients). During EFS the muscle contracted (54% of 65 strips; seven patients), relaxed (18%) or did not respond (28%). After terminating EFS a large “after-contraction” usually occurred (86%), decaying quickly (duration 13.5±0.7 s, type 1; 59%) or slowly (35.1±1.5 s, type 2; 41%). Atropine 10 μM prevented contraction during EFS (n=6) revealing muscle relaxation, and attenuated type 1 after-contraction (by 59.3±13.4%; n=8); the remaining contraction was abolished by neurokinin (NK) NK 1,2,3 receptor antagonists L732138 1 μM, MDL29913 1 μM+SB235375 0.1 μM. Atropine less effectively reduced type 2 after-contraction (by 25.1±16.9%, n=3) with NK 1,2,3 antagonism further reducing the contraction to 45.3±15.9% of control. NK 1,2,3 antagonists alone decreased type 1 and 2 after-contraction (by 38.4±34.8%, n=3 total). The NO synthase inhibitor L-NAME 0.3 mM prevented relaxation during EFS, revealing an atropine-sensitive contraction, and enhanced type 1 (by 164.0±98.6%, n=5) or attenuated type 2 after-contraction (by 27.4±26.7%, n=8). The 5-HT 4 receptor agonist prucalopride 10 μM increased contraction during EFS (by 90.6±52.5%, n=4) and variably decreased type 1 and 2 after-contraction (by 22.5±22.4 and 38.1±13.0%, respectively; n=2–4). In the presence of L-NAME prucalopride increased contraction during EFS by 30.8 ±30.2%, n=8. Conclusion The results suggest interplay between cholinergic, nitrergic and tachykinergic systems, so agents changing GI motility exert both inhibitory and excitatory actions on neurotransmission. Understanding these interactions now allows a more complete characterisation of the mechanisms by which agents affect colonic motility.