Neuroscience Letters, 119 (1990) 203-206
203
Elsevier Scientific Publishers Ireland Ltd. NSL 07288
The subserosal ganglia of the human taenia R. Crowe and G. Burnstock Department of Anatomy and DevelopmentalBiology, University CollegeLondon, London ( U.K.) (Received 23 July 1990; Accepted 25 July 1990)
Key words: Taenia; Neuropeptide; Ganglion; Histochemistry; Immunohistochemistry; Human Specimens of the taenia from the sigmoid colon of female patients undergoing surgery for carcinoma of the rectum were studied histochemically and immunohistochemically for acetylcholinesterase (ACHE) and for vasoactive intestinal polypeptide (NIP)-, substance P (SP)-, somatostatin (SOM)-, neuropeptide Y (NPY)-, calcitonin gene-related peptide (CGRP)- and Met-enkephalin (mENK)-immunoreactivity. Autonomic ganglia were observed on the serosal surface of the longitudinal muscle of the taenia. The subserosal ganglia contained SP-, mENK-, NPY-, SOM-, but not CGRP- or VIP-immunoreactive nerve fibres. In addition, they contained SP-, mENK- and NPY-, but not CGRP-, SOM- and VIP-immunoreactive nerve cell bodies (although CGRP- and VIP-immunoreactive nerve fibres were observed in the longitudinal muscle of the taenia). AChE-activity was found both in nerve fibres and nerve cell bodies in these ganglia. The greatest numbers of nerve cell bodies contained ACHE, followed in decreasing order by SP, mENK and NPY. The possible function of the subserosal ganglia of the human taenia is discussed.
It is well recognised that the enteric nervous system is the most complex system outside the central nervous system. The myenteric and submucous plexuses of the enteric nervous system supply nerve fibres to the longitudinal and circular muscle layers, muscularis mucosae, mucosa and blood vessels of the intestine, and normal motility of the intestine is dependent on the anatomical, chemical and functional integrity of the myenteric and submucous plexuses [4, 5, 11]. In addition to these plexuses, a subserosal plexus forming connections between extrinsic nerves and nerves of the deeper layer of the intestine has been reported in the stomach, near the mesenteric attachment of the intestine and on the surface of the rectum, at least in the cat [3, 15]. In the present study, a subserosal plexus is described in the taenia from the human female sigrnoid colon. The taenia was studied immunohistochemically for vasoactive intestinal polypeptide (VIP), substance P (SP) somatostatin (SOM), neuropeptide Y (NPY), calcitonin generelated peptide (CGRP), Met-enkephalin (mENK) and histochemicaUy for acetylcholinesterase (ACHE) activity. Specimens of the sigrnoid colon were obtained from female patients undergoing surgery for carcinoma of the rectum; tissues were taken from a site at a distance from the tumour. The dissected taenia from the sigmoid colon
Correspondence: G. Burnstock, Department of Anatomy and Developmental Biology, University College London, Gower Street, London WC1E 6BT, U.K. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
(n = 3) were fixed in 4 % paraformaldehyde in phosphatebuffered saline and neuropeptide immunoreactivity was visualised in sections by using the indirect fluorescence technique [8]. Acetylcholinesterase activity was localised histochemically according to the method of Karnovsky and Roots [13]. Sections of the human taenia were stained with haematoxylin and eosin and Toluidine blue to assess the morphology of this tissue. Autonomic ganglia containing 1-8 nerve cell bodies (diameter 25-32/tm) were observed in the connective tissue on the serosal surface of the longitudinal muscle of the taenia (Figs. la,b and 2a,b). The ganglia were variable in their relationship to the muscle; sometimes they were close (within 9/~m) but in other areas they might be as much as 700/tm away. The subserosal ganglia contained SP-, mENK-, NPY(Figs. 3a,b and 4a,b), SOM- but not CGRP- or VIPimmunoreactive nerve fibres which were found mainly in between or to encircle nerve cell bodies. In addition, they contained SP-, mENK-, and NPY- (Figs. 3a,b and 4a,b), but not CGRP-, VIP- and SOM-immunoreactive nerve cell bodies. AChE-activity was found both in nerve fibres and nerve cell bodies in these ganglia (Fig. la,b). The greatest number of nerve cell bodies contained ACHE, followed in decreasing order by SP, mENK and NPY. NPY-immunoreactive nerve fibres were observed around blood vessels in the serosal connective tissue. The longitudinal muscle of the taenia contained nerve fibres immunoreactive to SP, mENK, NPY, CGRP and
204
/
tm
Fig. I. Acetylcholinesterase activity in longitudinal sections o f the human taenia, a: 3 AChE-positive subserosal ganglia (short black arrows) in the connective tissue on the surface of the longitudinal muscle (lm) of the taenia. Bar = 180/~m. b: an AChE-positive subserosal ganglion (short black arrow) in the connective tissue on the surface of the longitudinal muscle (lm) o f the taenia. Bar = 30/zm.
Fig. 2. Haematoxylin and eosin stained tangential sections of the human taenia, a: a subserosal ganglion in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Bar = 100 gm. b: a higher magnification of a subserosal ganglion similar to that in (a). Bar = 30 urn.
VIP, but not SOM. AChE-positive nerves were also observed and the densities of these and neuropeptidecontaining nerves were similar. Although the subserosal plexus was observed and described in the middle of the 19th century there is very little mention of this plexus in the literature [3, 15]. The present study describes the subserosal plexus which contains AChE-positive as well as neuropeptide-containing nerve fibres and cell bodies in the human taenia for the first time. The subserosal plexus forms connections with the myenteric plexus, and vagal and mesenteric nerves form connections with the myenteric plexus via the subserosal plexus (see ref. 11). Since nerve fibres were found to envelop the nerve cell bodies, it is possible that some of these may be part of the vagal and splanchnic pathways and/or may be projected from the myenteric ganglia or nerve cell bodies within the serosal plexus modifying ganglionic transmission and forming a circuitry for enteric reflexes. The origin of AChE-positive and SP-, mENK- and
NPY-immunoreactive nerves observed in the longitudinal muscle may lie in the serosal plexus since nerve cell bodies containing these chemicals were observed within this plexus. Like the myenteric and submucous plexuses, the serosal plexus shows a heterogeneity in the chemistry of its neurones (see refs. 9-12, 16). The possible functions of the AChE-positive and SP-, mENK- and NPY-containing nerves and cell bodies in the serosal plexus are speculative. AChE-positivity does not necessarily indicate cholinergic nerves since AChE is also contained in SP neurones [7]. However, some of the AChE-positive neurons may be cholinergic and acetylcholine is known to be an excitatory neurotransmitter in the gastrointestinal tract [ 11]. The SP-containing nerve may represent sensory nerves, some of which reach the colon and rectum via the pelvic nerves [6]. mENK in the human colon inhibits the non-adrenergic non-cholinergic inhibitory neuromuscular transmission (C.H.V. Hoyle, personal communication). NPY has an inhibitory role in gut motility probably through inhi-
205
';
e
Fig. 3. Met-enkephalin- and neuropeptide Y-immunoreactivity in transverse sections of the human taenia, a: mENK-immunoreactive nerve fibres seen in a nerve bundle (long white arrow) and in a subserosal ganglion (short white arrow) in the connective tissue surrounding the longitudinal muscle (lm) of the taenia, b: a subserosal ganglion containing an NPY-immunoreactive nerve cell body (thin white arrow) and nerve fibres (short white arrow) around a non-fluorescent nerve cell body in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Note: the autofluorescence of the connective tissues typical of human tissue. Bar in a and b = 30 gm.
b i t i o n o f c h o l i n e r g i c n e u r o t r a n s m i s s i o n [1, 2, 18]. Since N P Y - i m m u n o r e a c t i v e n e r v e s were f o u n d a r o u n d b l o o d vessels in the serosa, N P Y m a y be i n v o l v e d i n g u t v a s o m o t o r c o n t r o l . N P Y h a s b e e n s h o w n to coexist w i t h n o r a d r e n a l i n e i n s y m p a t h e t i c n e r v e s [14] a n d w i t h V I P i n e n t e r i c n e r v e s [17]. C o e x i s t e n c e o f SP- a n d m E N K i m m u n o r e a c t i v e n e r v e s h a s also b e e n o b s e r v e d in the h u m a n large i n t e s t i n e [17]. T h u s s u c h c o e x i s t e n c e m a y also o c c u r in the s u b s e r o s a l g a n g l i a o f the h u m a n t a e n i a . T h e p r e s e n t s t u d y d e m o n s t r a t e s t h a t the s u b s e r o s a l plexus has a h e t e r o g e n e o u s c h e m i c a l c o m p o s i t i o n a n d f u r t h e r a d d s to the c o m p l e x i t y o f the e n t e r i c n e r v o u s syst e m in m a n .
Fig. 4. Substance P immunoreactivity in transverse sections of the human taenia, a: a SP-immunoreactive nerve cell body (white arrow) and nerve fibres in a subserosal ganglion in the connective tissue surrounding the longitudinal muscle (lm) of the taenia, b: SP-immunoreactive nerve fibres in a subserosal ganglion (short white arrow) in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Bar = 30/~m. Note: the autofluorescence of the connective tissue is typical of human tissue.
1 Allen, J.M., Adrian, T.E., Tatemoto, K., Polak, J.M., Hughes, J. and Bloom, S.R., Two novel peptides, neuropeptide Y (NPY) and peptide YY (PYY) inhibit the contraction of the electrically stimulatecl mouse vas deferens, Neuropeptides, 3 (1982) 71-77. 2 Allen, J.M., Hughes, J. and Bloom, S.R., Presence, distribution and pharmacological affects of neuropeptide Y in mammalian gastrointestinal tract, Dig. Dis. Sci., 32 (1987) 506-512. 3 Auerbach, L., Fernere vorlaufige Mitteilung iiber den Nervenapparat des Darmes, Arch Pathol Anat Physiol., 30 (1964) 457-460. 4 Burnstock, G., Cytochemical studies in the enteric nervous system. In: F. Chan-Palay and S.L. Palay (Eds.), Cytochemical Methods in Neuroanatomy, Alan Liss, New York, 1982, pp. 129-149. 5 Burnstoek, G., The changing face of autonomic transmission (The first Von Euler lecture in physiology.), Acta. Physiol. Scand., 126 (1986) 67-91. 6 Christensen, J., Stiles, M.J., Rick, G.A. and Sutherland, J., Comparative anatomy of the myenteric plexus of the distal colon in eight mammals, Gastroenterology, 86 (1984) 706-713. 7 Chubb, I.W., Hodgson, A.J. and White, G.H., Acetylcholinesterase hydrolyses substance P, Neuroscience, 5 (1980) 2065-2069. 8 Coons, A.H., Leduc, E.H. and Connolly, J.H., Studies on antibody production. 1. A method for the histochemical demonstration of specific antibody and its application to a study of the hyperimmune rabbit, J. Exp. Biol., 102 (1955)49-59.
206 9 Ekblad, E., Winther, C., Ekman, R., HAkanson, R. and Sundler, F., Projections of peptide-containingneurons in the rat small intestine, Neuroscience, 20 (1987) 169-188. 10 Furness, J.B. and Costa, M., VIP and enteric inhibitory nerves. In S. Said (Ed.), Vasoactive Intestinal Peptide, Raven, New York, 1982, pp. 391-406. 11 Furness, J.B. and Costa, M., The Enteric Nervous System, Churchill Livingstone, London, 1987. 12 Hoyle, C.H.V. and Bumstock, G., Neuronal populations in the submucous plexus of the human colon, J. Anat., 166 (1989) 7-22. 13 Karnovsky, M.J. and Roots, L., A direct colouring method for cholinesterase, J. Histochem. Cytochem., 12 (1964) 219-221. 14 Potter, E.K., Neuropeptide Y as an autonomic neurotransmitter, Pharmacol. Ther., 37 (1988) 251-273.
15 Schabadasch, A., Die Nerven des Magens der Katzo, Z. Zellforsch., I0 0930) 254-319. 16 Sundler, F., H~tkanson, R. and Leander, S., Peptidergic nervous system in the gut, Clin. Gastroenterol, 9 (1980) 517-543. 17 Wattchow, D.A., Furness, J.B. and Costa, M., Distribution and coexistence of peptides in nerve fibres of the external muscle of the human gastrointestinal tract, Gastroenterology, 95 (1988) 32-41. 18 Wiley, J. and Owyang, C., Neuropeptide Y inhibits cholinergic transmission in the isolated guinea=pig colon:-mediation through alpha=adrenergic receptors, Proc. Natl. Acad. Sci. U.S.A., 84 (1987) 2047-2051.
203
Elsevier Scientific Publishers Ireland Ltd. NSL 07288
The subserosal ganglia of the human taenia R. Crowe and G. Burnstock Department of Anatomy and DevelopmentalBiology, University CollegeLondon, London ( U.K.) (Received 23 July 1990; Accepted 25 July 1990)
Key words: Taenia; Neuropeptide; Ganglion; Histochemistry; Immunohistochemistry; Human Specimens of the taenia from the sigmoid colon of female patients undergoing surgery for carcinoma of the rectum were studied histochemically and immunohistochemically for acetylcholinesterase (ACHE) and for vasoactive intestinal polypeptide (NIP)-, substance P (SP)-, somatostatin (SOM)-, neuropeptide Y (NPY)-, calcitonin gene-related peptide (CGRP)- and Met-enkephalin (mENK)-immunoreactivity. Autonomic ganglia were observed on the serosal surface of the longitudinal muscle of the taenia. The subserosal ganglia contained SP-, mENK-, NPY-, SOM-, but not CGRP- or VIP-immunoreactive nerve fibres. In addition, they contained SP-, mENK- and NPY-, but not CGRP-, SOM- and VIP-immunoreactive nerve cell bodies (although CGRP- and VIP-immunoreactive nerve fibres were observed in the longitudinal muscle of the taenia). AChE-activity was found both in nerve fibres and nerve cell bodies in these ganglia. The greatest numbers of nerve cell bodies contained ACHE, followed in decreasing order by SP, mENK and NPY. The possible function of the subserosal ganglia of the human taenia is discussed.
It is well recognised that the enteric nervous system is the most complex system outside the central nervous system. The myenteric and submucous plexuses of the enteric nervous system supply nerve fibres to the longitudinal and circular muscle layers, muscularis mucosae, mucosa and blood vessels of the intestine, and normal motility of the intestine is dependent on the anatomical, chemical and functional integrity of the myenteric and submucous plexuses [4, 5, 11]. In addition to these plexuses, a subserosal plexus forming connections between extrinsic nerves and nerves of the deeper layer of the intestine has been reported in the stomach, near the mesenteric attachment of the intestine and on the surface of the rectum, at least in the cat [3, 15]. In the present study, a subserosal plexus is described in the taenia from the human female sigrnoid colon. The taenia was studied immunohistochemically for vasoactive intestinal polypeptide (VIP), substance P (SP) somatostatin (SOM), neuropeptide Y (NPY), calcitonin generelated peptide (CGRP), Met-enkephalin (mENK) and histochemicaUy for acetylcholinesterase (ACHE) activity. Specimens of the sigrnoid colon were obtained from female patients undergoing surgery for carcinoma of the rectum; tissues were taken from a site at a distance from the tumour. The dissected taenia from the sigmoid colon
Correspondence: G. Burnstock, Department of Anatomy and Developmental Biology, University College London, Gower Street, London WC1E 6BT, U.K. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
(n = 3) were fixed in 4 % paraformaldehyde in phosphatebuffered saline and neuropeptide immunoreactivity was visualised in sections by using the indirect fluorescence technique [8]. Acetylcholinesterase activity was localised histochemically according to the method of Karnovsky and Roots [13]. Sections of the human taenia were stained with haematoxylin and eosin and Toluidine blue to assess the morphology of this tissue. Autonomic ganglia containing 1-8 nerve cell bodies (diameter 25-32/tm) were observed in the connective tissue on the serosal surface of the longitudinal muscle of the taenia (Figs. la,b and 2a,b). The ganglia were variable in their relationship to the muscle; sometimes they were close (within 9/~m) but in other areas they might be as much as 700/tm away. The subserosal ganglia contained SP-, mENK-, NPY(Figs. 3a,b and 4a,b), SOM- but not CGRP- or VIPimmunoreactive nerve fibres which were found mainly in between or to encircle nerve cell bodies. In addition, they contained SP-, mENK-, and NPY- (Figs. 3a,b and 4a,b), but not CGRP-, VIP- and SOM-immunoreactive nerve cell bodies. AChE-activity was found both in nerve fibres and nerve cell bodies in these ganglia (Fig. la,b). The greatest number of nerve cell bodies contained ACHE, followed in decreasing order by SP, mENK and NPY. NPY-immunoreactive nerve fibres were observed around blood vessels in the serosal connective tissue. The longitudinal muscle of the taenia contained nerve fibres immunoreactive to SP, mENK, NPY, CGRP and
204
/
tm
Fig. I. Acetylcholinesterase activity in longitudinal sections o f the human taenia, a: 3 AChE-positive subserosal ganglia (short black arrows) in the connective tissue on the surface of the longitudinal muscle (lm) of the taenia. Bar = 180/~m. b: an AChE-positive subserosal ganglion (short black arrow) in the connective tissue on the surface of the longitudinal muscle (lm) o f the taenia. Bar = 30/zm.
Fig. 2. Haematoxylin and eosin stained tangential sections of the human taenia, a: a subserosal ganglion in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Bar = 100 gm. b: a higher magnification of a subserosal ganglion similar to that in (a). Bar = 30 urn.
VIP, but not SOM. AChE-positive nerves were also observed and the densities of these and neuropeptidecontaining nerves were similar. Although the subserosal plexus was observed and described in the middle of the 19th century there is very little mention of this plexus in the literature [3, 15]. The present study describes the subserosal plexus which contains AChE-positive as well as neuropeptide-containing nerve fibres and cell bodies in the human taenia for the first time. The subserosal plexus forms connections with the myenteric plexus, and vagal and mesenteric nerves form connections with the myenteric plexus via the subserosal plexus (see ref. 11). Since nerve fibres were found to envelop the nerve cell bodies, it is possible that some of these may be part of the vagal and splanchnic pathways and/or may be projected from the myenteric ganglia or nerve cell bodies within the serosal plexus modifying ganglionic transmission and forming a circuitry for enteric reflexes. The origin of AChE-positive and SP-, mENK- and
NPY-immunoreactive nerves observed in the longitudinal muscle may lie in the serosal plexus since nerve cell bodies containing these chemicals were observed within this plexus. Like the myenteric and submucous plexuses, the serosal plexus shows a heterogeneity in the chemistry of its neurones (see refs. 9-12, 16). The possible functions of the AChE-positive and SP-, mENK- and NPY-containing nerves and cell bodies in the serosal plexus are speculative. AChE-positivity does not necessarily indicate cholinergic nerves since AChE is also contained in SP neurones [7]. However, some of the AChE-positive neurons may be cholinergic and acetylcholine is known to be an excitatory neurotransmitter in the gastrointestinal tract [ 11]. The SP-containing nerve may represent sensory nerves, some of which reach the colon and rectum via the pelvic nerves [6]. mENK in the human colon inhibits the non-adrenergic non-cholinergic inhibitory neuromuscular transmission (C.H.V. Hoyle, personal communication). NPY has an inhibitory role in gut motility probably through inhi-
205
';
e
Fig. 3. Met-enkephalin- and neuropeptide Y-immunoreactivity in transverse sections of the human taenia, a: mENK-immunoreactive nerve fibres seen in a nerve bundle (long white arrow) and in a subserosal ganglion (short white arrow) in the connective tissue surrounding the longitudinal muscle (lm) of the taenia, b: a subserosal ganglion containing an NPY-immunoreactive nerve cell body (thin white arrow) and nerve fibres (short white arrow) around a non-fluorescent nerve cell body in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Note: the autofluorescence of the connective tissues typical of human tissue. Bar in a and b = 30 gm.
b i t i o n o f c h o l i n e r g i c n e u r o t r a n s m i s s i o n [1, 2, 18]. Since N P Y - i m m u n o r e a c t i v e n e r v e s were f o u n d a r o u n d b l o o d vessels in the serosa, N P Y m a y be i n v o l v e d i n g u t v a s o m o t o r c o n t r o l . N P Y h a s b e e n s h o w n to coexist w i t h n o r a d r e n a l i n e i n s y m p a t h e t i c n e r v e s [14] a n d w i t h V I P i n e n t e r i c n e r v e s [17]. C o e x i s t e n c e o f SP- a n d m E N K i m m u n o r e a c t i v e n e r v e s h a s also b e e n o b s e r v e d in the h u m a n large i n t e s t i n e [17]. T h u s s u c h c o e x i s t e n c e m a y also o c c u r in the s u b s e r o s a l g a n g l i a o f the h u m a n t a e n i a . T h e p r e s e n t s t u d y d e m o n s t r a t e s t h a t the s u b s e r o s a l plexus has a h e t e r o g e n e o u s c h e m i c a l c o m p o s i t i o n a n d f u r t h e r a d d s to the c o m p l e x i t y o f the e n t e r i c n e r v o u s syst e m in m a n .
Fig. 4. Substance P immunoreactivity in transverse sections of the human taenia, a: a SP-immunoreactive nerve cell body (white arrow) and nerve fibres in a subserosal ganglion in the connective tissue surrounding the longitudinal muscle (lm) of the taenia, b: SP-immunoreactive nerve fibres in a subserosal ganglion (short white arrow) in the connective tissue surrounding the longitudinal muscle (lm) of the taenia. Bar = 30/~m. Note: the autofluorescence of the connective tissue is typical of human tissue.
1 Allen, J.M., Adrian, T.E., Tatemoto, K., Polak, J.M., Hughes, J. and Bloom, S.R., Two novel peptides, neuropeptide Y (NPY) and peptide YY (PYY) inhibit the contraction of the electrically stimulatecl mouse vas deferens, Neuropeptides, 3 (1982) 71-77. 2 Allen, J.M., Hughes, J. and Bloom, S.R., Presence, distribution and pharmacological affects of neuropeptide Y in mammalian gastrointestinal tract, Dig. Dis. Sci., 32 (1987) 506-512. 3 Auerbach, L., Fernere vorlaufige Mitteilung iiber den Nervenapparat des Darmes, Arch Pathol Anat Physiol., 30 (1964) 457-460. 4 Burnstock, G., Cytochemical studies in the enteric nervous system. In: F. Chan-Palay and S.L. Palay (Eds.), Cytochemical Methods in Neuroanatomy, Alan Liss, New York, 1982, pp. 129-149. 5 Burnstoek, G., The changing face of autonomic transmission (The first Von Euler lecture in physiology.), Acta. Physiol. Scand., 126 (1986) 67-91. 6 Christensen, J., Stiles, M.J., Rick, G.A. and Sutherland, J., Comparative anatomy of the myenteric plexus of the distal colon in eight mammals, Gastroenterology, 86 (1984) 706-713. 7 Chubb, I.W., Hodgson, A.J. and White, G.H., Acetylcholinesterase hydrolyses substance P, Neuroscience, 5 (1980) 2065-2069. 8 Coons, A.H., Leduc, E.H. and Connolly, J.H., Studies on antibody production. 1. A method for the histochemical demonstration of specific antibody and its application to a study of the hyperimmune rabbit, J. Exp. Biol., 102 (1955)49-59.
206 9 Ekblad, E., Winther, C., Ekman, R., HAkanson, R. and Sundler, F., Projections of peptide-containingneurons in the rat small intestine, Neuroscience, 20 (1987) 169-188. 10 Furness, J.B. and Costa, M., VIP and enteric inhibitory nerves. In S. Said (Ed.), Vasoactive Intestinal Peptide, Raven, New York, 1982, pp. 391-406. 11 Furness, J.B. and Costa, M., The Enteric Nervous System, Churchill Livingstone, London, 1987. 12 Hoyle, C.H.V. and Bumstock, G., Neuronal populations in the submucous plexus of the human colon, J. Anat., 166 (1989) 7-22. 13 Karnovsky, M.J. and Roots, L., A direct colouring method for cholinesterase, J. Histochem. Cytochem., 12 (1964) 219-221. 14 Potter, E.K., Neuropeptide Y as an autonomic neurotransmitter, Pharmacol. Ther., 37 (1988) 251-273.
15 Schabadasch, A., Die Nerven des Magens der Katzo, Z. Zellforsch., I0 0930) 254-319. 16 Sundler, F., H~tkanson, R. and Leander, S., Peptidergic nervous system in the gut, Clin. Gastroenterol, 9 (1980) 517-543. 17 Wattchow, D.A., Furness, J.B. and Costa, M., Distribution and coexistence of peptides in nerve fibres of the external muscle of the human gastrointestinal tract, Gastroenterology, 95 (1988) 32-41. 18 Wiley, J. and Owyang, C., Neuropeptide Y inhibits cholinergic transmission in the isolated guinea=pig colon:-mediation through alpha=adrenergic receptors, Proc. Natl. Acad. Sci. U.S.A., 84 (1987) 2047-2051.
