Mang, Christian F., Sebastian Truempler, Doris Erbelding, and Heinz Kilbinger. Modulation by NO of acetylcholine release in the ileum of wild-type and NOS gene knockout mice. Am J Physiol Gastrointest Liver Physiol 283: G1132-G1138, 2002 10.1152/ajpgi.00192.2002 inhibits the release of acetylcholine and cholinergic contractions in the small intestine of several species, but no information is available about the mouse ileum. This study examines the effects of NO on the electrically evoked release of 3 H]acetylcholine release and cholinergic contractions in preparations from wild-type mice and from eNOS knockout mice. Effects of L-NNA were specifically antagonized by L-arginine. In contrast, L-NNA and ODQ did not modify the release and contractions in preparations from nNOS knockout mice. The NO donor S-nitroso-N-acetyl-DLpenicillamine inhibited the electrically evoked release of [ 3 H]acetylcholine and longitudinal muscle contractions in a quantitatively similar manner in wild-type preparations as well as in nNOS and eNOS knockout preparations. We conclude that endogenous NO released by electrical field stimulation tonically inhibits the release of acetylcholine. Furthermore, data suggest that nNOS and not eNOS is the enzymatic source of NO-mediating inhibition of cholinergic neurotransmission in mouse ileum. neuronal nitric oxide synthase knockout mice; endothelial nitric oxide synthase knockout mice; cholinergic neurotransmission NITRIC OXIDE (NO) is a nonadrenergic, noncholinergic neurotransmitter that causes inhibition of intestinal motility (for review, see Ref. 24). The relaxant action of NO is probably not confined to its direct action on smooth muscle, because NO decreases intestinal motility also indirectly by inhibiting the release of the functionally most prominent excitatory enteric neurotransmitters acetylcholine and substance P. For example, exogenous NO (31) and NO donors (5) inhibit the electrically evoked release of acetylcholine from guinea pig ileum. Vice versa, NO synthase (NOS) inhibitors enhance the evoked release of acetylcholine from the ileum of guinea pig (10,20) and dog (7) as well as the electrically evoked cholinergic and tachykininergic contractions of guinea pig ileum (6,32). This suggests that endogenous NO, released by field stimulation, decreases the evoked release of acetylcholine from enteric neurones. There are, however, tissue and species differences in the neuromodulatory effect of NO: NO or NOS inhibitors do not modify the electrically evoked acetylcholine release from the colon of guinea pig and dog (22,29) and from stomach preparations of man (13