Structural diversity of NADPH diaphorase-reactive enteral networks in Stylommatophora (Gastropoda, Pulmonata)

Abstract. In this work we investigated the involvement of putative nitric oxide (NO)-forming neurons in enteric plexuses of stylommatophoran gastropods. The nitric oxide synthase (NOS)-containing cells were detected by NADPH diaphorase (NADPHd) histochemistry in the entreral nervous systems of several stylommatophoran species (Achatinacea: Achatina fulica , Helicacea: Cepaea hortensis, Cepaea nemoralis, Discus rotundatus, Helicella obvia, Helix lucorum, Helix lutescens, Monachoides umbrosa, Trichia hispida, Zebrina detrita , Succineacea: Succinea putris , Vertiliginacea: Clausilia dubia , Zonitacea: Arion ater, Arion subfuscus, Limax maximus ). We detected the NO synthesis of isolated midintestinal segments by Griess's quantification of nitrite, one end product of NO. Effects of the NOS substrate L-arginine and the NOS inhibitor Nω-nitro-L-arginine (NOARG) were also tested on nitrite production. We found NADPHd-reactive neurons and extrinsic nerves with NADPHd-stained fibers within the myenteric and submucosal networks of the midintestine of investigated members of Helicacea, Succineacea, and Vertiliginacea families. These networks innervated the midintestinal musculature and several nerve cells of the myenteric and submucosal plexi. In investigated members of Achatinacea and Zonitacea, NADPHd-stained networks were not detectable within the digestive tract. Administration of 1 mM L-arginine elevated, whereas 2 mM of NOARG diminished, the nitrite levels of the NADPHd-stained networks containing midintestine in C. nemoralis and H. lucorum . Enteral NADPHd staining was not detected in A. ater and L. maximus , and the nitrite production was not affected by L-arginine. Our results indicate a possible, but evolutionarily not conserved, NO-mediated enteral transmission in stylommatophoran gastropods.