Central and peripheral neuronal pathways revealed by backfilling with neurobiotin in the optic, tentacular and small labial nerves of Lymnaea stagnalis Oksana P. Tuchina, 1 Valery V. Zhukov 2 and V. Benno Meyer-Rochow 3,4 1 School of Engineering and Science, Jacobs University, Campus Ring 6, Research II, Room 38, D-28759 Bremen, Germany; 2 Department of Agricultural and Soil Ecol- ogy, Faculty of Bioresources and Natural Usage, Kaliningrad State Technical Univer- sity, Sovetsky avenue, 1, 236000 Kalinin- grad, Russia; 3 School of Engineering and Science, Jacobs University, Campus Ring 6, Research II, Room 37, D-28759 Bremen, Germany; 4 Department of Biology & Physi- ology, Oulu University, FI-90014 Oulu, Finland Keywords: gastropoda, immunocytochemistry, nervous system, retrograde transport, snails, visual projections Accepted for publication: 10 May 2010 Abstract Tuchina, O.P., Zhukov, V.V. and Meyer-Rochow, V.B. 2012. Central and peripheral neuronal pathways revealed by backfilling with neurobiotin in the optic, tentacular and small labial nerves of Lymnaea stagnalis. —Acta Zoologica (Stockholm) 93: 28–47. The TOLm complex in Lymnaea stagnalis contains nerves n. tentacularis, n. opti- cus and n. labialis minor. Ligatures close to where the complex enters the central nervous system (CNS) did not prevent penetration of retrograde-transported neurobiotin into fibres of an adjacent nerve. Axonal bifurcation within the com- mon nerve trunk or tight junctions may be involved, providing a basis for periph- eral axon reflexes. Peripheral terminations of n. tentacularis, n. labialis minor and n. opticus revealed numerous cell bodies in the tentacular epithelium, some in the tentacle and lip region, and some in the retina. These cell bodies’ central pro- jections were mapped by neurobiotin and verified by dissections of the cerebro- cerebral commissure and cerebro-pleural connective. Afferent fibres of the nerves form dense sensory neuropils in the ipsilateral cerebral ganglia. Direct connections between n. tentacularis and some visceral as well as parietal nerves were demonstrated by backfillings through n. pallialis dexter internus et externus, n. pallialis sinister and n. intestinalis. Labelling of n. tentacularis revealed neuronal bodies in every ganglion and stained fibres in most of the peripheral nerves. Fewer neurons were identified through n. labialis minor and n. opticus. We dis- cuss our results in relation to different behavioural forms like defence and feed- ing reactions in L. stagnalis. V. Benno Meyer-Rochow, School of Engineering and Science, Jacobs Univer- sity, Campus Ring 6, Research II, Room 37, D-28759 Bremen, Germany. E-mail: b.meyer-rochow@jacobs-university.de Introduction Tracing retrograde transport of different dyes is a widespread method to visualize neural elements and determine their path- ways in animals, including molluscs (Winlow and Kandel 1976; Benjamin and Rose 1979, 1980; Benjamin et al. 1979; Zaitseva 1986). By using dyes like cobalt and nickel salts, luci- fer yellow, dextran and horse radish peroxidase (HRP), it is possible to identify neurons in the freshwater gastropod Lym- naea stagnalis that send their axons along different peripheral nerves and to reveal the sensory inputs and motor outputs of this snail’s CNS. This holds true for neurons that take part in controlling well-recognized motor patterns, for instance those involved in locomotion (Slade et al. 1981), the withdrawal reflex (Ferguson and Benjamin 1991a; Kononenko and Zhu- kov 2005), tentacle contraction (Lever 1977), feeding activity (Nakamura et al. 1999) and respiratory control (Lukowiak et al. 2006). Usually, neuronal networks, responsible for some specific functions, consist of sensory (afferent) neurons, then interneurons, which process the information, make connec- tions with other functionally different networks and may also act as rhythm generators, and finally efferent or motoneurons, which innervate the muscles. In case of, for example, the whole-body withdrawal network in L. stagnalis, a ‘lights-off’ Acta Zoologica (Stockholm) 93: 28–47 (January 2012) doi: 10.1111/j.1463-6395.2010.00477.x Ó 2010 The Authors 28 Acta Zoologica Ó 2010 The Royal Swedish Academy of Sciences