413 zyxwvut European Journal of Pharmacology , 144 (1987) 413-415 Elsevier EZJP 096RC Rapid communication Receptor autoradiography with [ 3H]L-glutamate reveals the presence and axonal transport of glutamate receptors in vagal afferent neurones of the rat Stephen J. Lewis ‘, Marion Cincotta, Anthony J.M. Verberne, Bevyn Jarrott, David Lodge 2 and Philip M. Beart * University of Melbourne, Clinical Pharmacology and Therapeutics Unii, Austin Hospital, Heidelberg, Victoria 3084, Australia and 2 Department of Physiology, Royal Veterinary College, London N W I OTU, U.K. Received 9 November 1987, accepted 11 November 1987 The perikarya of vagal afferent neurones are located within the inferior vagal (nodose) ganglia (Palkovits and Zaborsky, 1977). Recent evidence suggests that receptors for a variety of putative neurotransmitters/neuromodulators may be syn- thesized within these perikarya and then delivered by axonal transport mechanisms in the vagus nerve to their central and/or peripheral processes where they are incorporated into the terminal mem- branes (Laduron, 1984; Young et al., 1980). In this study we provide evidence from receptor autoradiography with [ 3H]L-glutamate that bind- ing sites for the excitatory transmitter L-gluta- mate, are associated with the perikarya of vagal afferent neurones and that these receptors un- dergo axonal transport in the peripheral vagal trunks. Female Sprague-Dawley rats (225-250 g) were anaesthetized with an amylobarbitone-methohexi- tone (30 and 16.7 mg/kg i.p. respectively) mix- ture. In 4 rats the left nodose ganglion was ex- posed and the vagus nerve was tightly ligatured (siliconised silk thread, 6/O; Dynek Pty. Ltd., Australia) approximately 1 cm distal to the gan- ’ Present address: Department of Pharmacology and Cardio- vascular Center, University of Iowa, Iowa City, IA 52242, U.S.A. * To whom all correspondence should be addressed. glion. In control rats (n = 4) a loosely fitting ligature was placed at the same distance from the left ganglion. Upon conclusion of the surgery, the wound was closed and the animals maintained in a warm, sterile environment. After 24 h, the animals were reanaesthetized and the vagi were sectioned a few millimetres distal to the ligature or sham-ligature, and proximal to the nodose gangli- on. The pharyngeal and superior laryngeal nerves were cut near their point of connection with the ganglion (see fig. la). Each vagus nerve and asso- ciated nodose ganglion were then removed and immediately frozen in mounting medium (Tissue- Tek, Miles Scientific, U.S.A.) at -20°C over- night. Longitudinal sections (10 pm) including the ganglion and vagus nerve, were then cut using a cryostat and thaw-mounted on to glass micro- scope slides for receptor autoradiography with [ 3H]L-glutamate (Cincotta et al., 1987). Tissue sections were air dried, preincubated in 50 mM Tris-citrate pH 7.0 for 20 min and dried in a stream of cool air before incubation for 10 min in 50 mM Tris-citrate pH 7.0, containing 2.5 mM Ca*’ and 20 mM Cl-, with [3H]L-glutamate (1 PM; 42 Ci/mmol, Amersham, U.K.). Rapid wash- ing for 0.5 min in ice-cold Tris-citrate followed. Full details of this procedure will be published at a later date. Parallel sections incubated with 1 mM L-glutamate served to define non-specific binding. Thorough drying in air and over silica gel was carried out, before sections were apposed to 0014-2999/87/$03.50 0 1987 Elsevier Science Publishers B.V. (Biomedical Division)