Stretch-activated neuronal pathways to longitudinal and circular muscle in guinea pig distal colon

The role of the longitudinal muscle (LM) layer during the peristaltic reflex in the small and large intestine is unclear. In this study, we have made double and quadruple simultaneous intracellular recordings from LM and circular muscle (CM) cells of guinea pig distal colon to correlate the electrical activities in the two different muscle layers during circumferential stretch. Simultaneous recordings from LM and CM cells (<200 microm apart) at the oral region of the colon showed that excitatory junction potentials (EJPs) discharged synchronously in both muscle layers for periods of up to 6 h. Similarly, at the anal region of the colon, inhibitory junction potentials (IJPs) discharged synchronously in the two muscle layers. Quadruple recordings from LM and CM orally at the same time as from the LM and CM anally revealed that IJPs occurred synchronously in the LM and CM anally at the same time as EJPs in LM and CM located 20 mm orally. Oral EJPs and anal IJPs were linearly related in amplitude between the two muscle layers. Spatiotemporal maps generated from simultaneous video imaging of the movements of the colon, combined with intracellular recordings, revealed that some LM contractions orally could be correlated in time with IJPs in CM cells anally. N(omega)-nitro-L-arginine (L-NA; 100 microM) abolished the IJP in LM, whereas a prominent L-NA-resistant "fast" IJP was always observed in CM. In summary, in stretched preparations, synchronized EJPs in both LM and CM orally are generated by synchronized firing of many ascending interneurons, which simultaneously activate excitatory motor neurons to both muscle layers. Similarly, synchronized IJPs in both LM and CM anally are generated by synchronized firing of many descending interneurons, which simultaneously activate inhibitory motor neurons to both muscle layers. This synchronized motor activity ensures that both muscles around the entire circumference are excited orally at the same time as inhibited anally, thus producing net aboral propulsion.     

Structural characterization of interstitial cells of Cajal in myenteric plexus and muscle layers of canine colon

We have carried out a detailed ultrastructural study of the interstitial cells near the myenteric plexus of the canine colon and defined the structural characteristics which distinguish them from other resident non-neural cells. We have also examined the interconnections of these interstitial cells with nerves, the longitudinal muscle, and the circular muscle. In addition, we sought connections between interstitial cells of the myenteric plexus and those described earlier at the inner border of the circular muscle in proximal and distal colon. The interstitial cells of the myenteric plexus were structurally distinctive, and made gap junctions with one another and occasionally with smooth muscle. There seemed to be two subsets of these interstitial cells, one associated with the longitudinal muscle and the other with the circular muscle. Cells of both subsets were often close (less than or equal to 20 nm) to nerve profiles. The interstitial cells near the longitudinal muscle layer penetrated slightly into the muscle layer, but those near the circular muscle did not and neither set contacted the other. Moreover, interstitial cells of Cajal located near the myenteric plexus were never observed to contact those at the inner border of circular muscle. The interstitial cells of Cajal at the canine colon myenteric plexus are structurally organized to provide independent pacemaking activities for the longitudinal and adjacent circular muscle. Their dense innervation suggests that they mediate neural modulation of intestinal pacemaker activities. Moreover, they lack direct contacts with the interstitial cell network at the inner border of circular muscle, which is essential for the primary pacemaking activity of circular muscle. The structural organization of interstitial cells in canine colon is consistent with their proposed role in pacemaking activity of the two muscle layers.     

Projections of 5-hydroxytryptamine-immunoreactive neurons in guinea-pig distal colon

The presence of 5-hydroxytryptamine in enteric neurons of the guinea-pig distal colon was demonstrated by immunohistochemistry and the projections of the neurons were determined. 5-Hydroxytryptamine-containing nerve cells were observed in the myenteric plexus but no reactive nerve cells were found in submucous ganglia. Varicose reactive nerve fibres were numerous in the ganglia of both the myenteric and submucous plexuses, but were infrequent in the longitudinal muscle, circular muscle, muscularis mucosae and mucosa. Reactivity also occurred in enterochromaffin cells. Lesion studies showed that the axons of myenteric neurons projected anally to provide innervation to the circular muscle and submucosa and to other more anally located myenteric ganglia. The results suggest that a major population of 5-hydroxytryptamine neurons in the colon is descending interneurons, most of which extend for 10 to 15 mm in the myenteric plexus and innervate both 5-hydroxytryptamine and non-5-hydroxytryptamine neurons.     

Inhibitory innervation of colonic smooth muscle cells and interstitial cells of Cajal

The effect of neural inhibition on the electrical activities of circular and longitudinal colonic smooth muscle was investigated. In addition, a comparative study was carried out between circular muscle preparations with and without the "submucosal" and "myenteric plexus" network of interstitial cells of Cajal (ICC) to study innervation of the "submucosal" ICC and to investigate whether or not the ICC network is an essential intermediary system for inhibitory innervation of smooth muscle cells. Electrical stimulation of intrinsic nerves in the presence of atropine caused inhibitory junction potentials (ijps) throughout the circular and longitudinal muscle layers. The ijp amplitude depended on the membrane potential and not on the location of the muscle cells with respect to the ICC network. Neurally mediated inhibition of the colon resulted in a reduction in amplitude and duration of slow wave type action potentials in circular and abolishment of spike-like action potentials in longitudinal smooth muscle, both resulting in a reduction of contractile activity. With respect to mediation by ICC, the study shows (i) "submucosal" ICC receive direct inhibitory innervation and (ii) circular smooth muscle cells can be directly innervated by inhibitory nerves without ICC as necessary intermediaries. The reversal potential of the ijp in colonic smooth muscle was observed to be approximately -76 mV, close to the estimated potassium equilibrium potential, suggesting that the nerve-mediated hyperpolarization is caused by increased potassium conductance.     

The source of the nerve fibres forming the deep muscular and circular muscle plexuses in the small intestine of the guinea-pig

Summary A quantitative ultrastructural study was made of the neuntes forming the deep muscular and circular muscle plexuses of the guinea-pig small intestine following microsurgical lesions designed to interrupt intrinsic and extrinsic nerve pathways within the intestinal wall. Removal of a collar of longitudinal muscle with attached myenteric plexus from the circumference of a segment of small intestine resulted in the subsequent disappearance of 99.3% of neurites in the underlying circular muscle. The few surviving neurites in the deep muscular plexus and circular muscle disappeared completely from lesioned segments that were, in addition, extrinsically denervated surgically. These results indicate that the majority of nerve fibres in the deep muscular and circular muscle plexuses of the guinea-pig small intestine is intrinsic to the intestine and originates from nerve cell bodies located in the overlying myenteric plexus. At the light-microscopic level, nerve bundles were traced from the myenteric plexus to the circular muscle.     

Distribution and projections of neurons with immunoreactivity for both gastrin-releasing peptide and bombesin in the guinea-pig small intestine

Summary Bombesin-like and gastrin-releasing peptide (GRP)-like immunoreactivities were localized in nerves of the guinea-pig small intestine and celiac ganglion with the use of antibodies raised against the synthetic peptides. The anti-bombesin serum (preincubated to avoid cross reactivity with substance P) and the anti-GRP serum revealed the same population of neurons. Preincubation of the antibombesin serum with bombesin abolished the immunoreactivity in nerves while absorption of the anti-GRP serum with either bombesin or the 14–27 C-terminal of GRP only reduced the immunoreactivity. The immunoreactivity was abolished by incubation with GRP 1–27.Immunoreactive nerves were found in the myenteric plexus, circular muscle, submucous plexus and in the celiac ganglion. Faintly reactive nerve cell bodies were found in the myenteric ganglia (3.2% of all neurons) but not in submucous ganglia. After all ascending and descending pathways in the myenteric plexus had been cut, reactive terminals disappeared in the myenteric plexus, circular muscle (including the deep muscular plexus) and the submucous plexus on the anal side. After the mesenteric nerves were cut no changes were observed in the intestinal wall but the reactive fibres in celiac ganglia disappeared. It is deduced that GRP/bombesin-immunoreactive nerve cell bodies in myenteric ganglia project from the myenteric plexus to other myenteric ganglia situated further anally (average length 12 mm), anally to the circular muscle (average length 9 mm), anally to submucous ganglia (average length 13 mm) and external to the intestine to the celiac ganglia.It is concluded that the GRP/bombesin-reactive neurons in the intestinal wall represent a distinct population of enteric neurons likely to be involved in controlling motility and in the coordination of other intestinal functions.     

Three-dimensional structures of c-Kit-positive cellular networks in the guinea pig small intestine and colon

Cryosections and whole-mount preparations of the guinea pig small intestine and colon were single or double immunolabeled using the anti-c-Kit and protein gene product 9.5 antibodies. Immunolabeled specimens were observed under a confocal laser scanning microscope. The main findings of the present study are: (1) the distribution and profiles of three-dimensional structures of c-Kit-positive cellular networks in the small intestine and colon, and (2) the anatomical relations of c-Kit-positive cells to the enteric nerves in the layers. In the small intestine, c-Kit-positive cellular networks were observed at levels of the deep muscular plexus and myenteric plexus. The c-Kit-positive cellular networks ran along or overlay the nerve fibers at the deep muscular plexus, while they showed the reticular structures intermingled with the nerve elements at the myenteric plexus. In the colon, c-Kit-positive cellular networks were observed at levels of the submuscular plexus and myenteric plexus, and were further identified within the circular and longitudinal muscle layers as well as in the subserosal layer. In the circular muscle layer, c-Kit-positive cells surrounded the associated nerve fibers and extended several long processes toward the adjacent c-Kit-positive cells. The c-Kit-positive cellular networks within the longitudinal muscle layer as well as in the subserosal layer were not associated with the nerve fibers. In the layers of the intestinal wall with c-Kit-positive cells, the cellular networks of the interstitial cells were identified in ultrastructure. The characteristic profiles of c-Kit-positive cellular networks provide a morphological basis upon which to investigate the mechanisms regulating intestinal movement. Received: 14 July 1998 / Accepted: 2 September 1998     

Circumferential,not longitudinal,colonic stretch increases synaptic input to mouse prevertebral ganglion neurons

The relationship between longitudinal and circular muscle tension in the mouse colon and mechanosensory excitatory synaptic input to neurons in the superior mesenteric ganglion (SMG) was investigated in vitro. Electrical activity was recorded intracellularly from SMG neurons, and muscle tension was simultaneously monitored in the longitudinal, circumferential, or both axes. Colonic intraluminal pressure and volume changes were also monitored simultaneously with muscle tension changes. The results showed that the frequency of fast excitatory postsynaptic potentials (fEPSPs) in SMG neurons increased when colonic muscle tension decreased, when the colon relaxed and refilled with fluid after contraction, and during receptive relaxation preceding spontaneous colonic contractions. In contrast, fEPSP frequency decreased when colonic muscle tension increased during spontaneous colonic contraction and emptying. Manual stretch of the colon wall to 10-15% beyond resting length in the circumferential axis of flat sheet preparations increased fEPSP frequency in SMG neurons, but stretch in the longitudinal axis to 15% beyond resting length in the same preparations did not. There was no increase in synaptic input when tubular colon segments were stretched in their long axes up to 20% beyond their resting length. The circumferential stretch-sensitive increase in the frequency of synaptic input to SMG neurons persisted when the colonic muscles were relaxed pharmacologically by nifedipine (2 microM) or nicardipine (3 microM). These results suggest that colonic mechanosensory afferent nerves projecting to the SMG function as length or stretch detectors in parallel to the circular muscle layer.     

Role of l-Ca(2+) channels in intestinal pacing in wild-type and W/W(V) mice

Rhythmic contractions generating transit in the digestive tract are paced by a network of cells called interstitial cells of Cajal (ICC) found in the myenteric plexus (MP). ICC generate cyclic depolarizations termed "slow waves" that are passively transmitted to the smooth muscle to initiate contractions. The opening of l-Ca(2+) channels are believed to be primarily responsible for the influx of calcium generating a contraction in smooth muscle. However, l-Ca(2+) channels are not thought to be important in generating the pacing current found in ICC. Using intact segments of circular (CM) and longitudinal (LM) muscle from wild-type mice and mice lacking c-kit kinase (W/W(V)), we found that l-Ca(2+) channel currents are required for pacing at normal frequencies to occur. Application of 1 muM nicardipine caused a significant decrease in contraction amplitude and frequency in LM and CM that was successfully blocked with BAY K 8644. Nicardipine also abolished the pacing gradient found throughout the intestines, resulting in a uniform contraction frequency of 30-40/minute. Stimulating l-Ca(2+) channels with BAY K 8644 neither removed nor recovered the pacing gradient. W/W(V) mice, which lack ICC-MP, also exhibited a pacing gradient in LM. Application of nicardipine to LM segments of W/W(V) mouse intestine did not reduce pacing frequency, and in jejunum, resulted in a slight increase. BAY K 8644 did not affect pacing frequency in W/W(V) tissue. In conclusion, we found that l-Ca(2+) channel activity was required for normal pacing frequencies and to maintain the pacing frequency gradient found throughout the intestines in wild-type but not in W/W(V) mouse intestine.     

Nitric oxide synthase in the enteric nervous system of the guinea-pig: a quantitative description

The distribution and abundance of nitric oxide synthase (NOS)-containing neurons and their terminals in the gastrointestinal tract of the guinea-pig were examined in detail using NADPH diaphorase histochemistry and NOS immunohistochemistry. NOS-containing cell bodies were found in the myenteric plexus throughout the gastrointestinal tract and in the submucous plexus of the stomach, colon and rectum. NOS-containing neurons comprised between 12% (in the duodenum) and 54% (in the esophagus) of total myenteric neurons. In the ileum, NOS neurons represented 19% of total myenteric neurons. Most of the NOS neurons throughout the gastrointestinal tract possessed lamellar dendrites and a single axon. NOS-containing terminals were abundant in the circular muscle, including that of the sphincters, but were rare in the longitudinal muscle, except for the taeniae of the caecum. The muscularis mucosae of the esophagus, stomach, colon and rectum received a medium to dense innervation by NOS terminals. Within myenteric ganglia, NOS-containing terminals were extremely sparse in the esophagus, stomach and duodenum, common in the ileum and distal colon and extremely dense in the proximal colon and rectum. The submucous plexus in the ileum and large intestine contained a sparse plexus of NOS-containing terminals. NOS terminals were not observed in the mucosa of any region. We conclude that throughout the gastrointestinal tract of the guinea-pig, NOS neurons are inhibitory motor neurons to the circular muscle; in the ileum and large intestine, NOS neurons may also function as interneurons.     

Ruminal muscle of sheep is innervated by non-polarized pathways of cholinergic and nitrergic myenteric neurones

The motility patterns of the reticulorumen evoke mainly mixing of the ingesta. So far unknown, intrinsic neural circuits of the enteric nervous system are involved in the control of these motility patterns. The aim of the study was to characterize neurochemically sheep ruminal myenteric neurones, in particular the neural pathways innervating the ruminal muscle layers. Cell bodies within the myenteric plexus projecting to the longitudinal or circular muscle layer were retrogradely labelled by direct application of the fluorescent tracer 1,1'-didodecyl-3,3,3',3'-tetramethyl indocarbocyanine perchlorate (DiI) onto the circular or longitudinal muscle. The neurochemical code of myenteric neurones was identified by their immunoreactivity for choline acetyltransferase (ChAT), nitric oxide synthase (NOS), substance P (SP) and vasoactive intestinal peptide (VIP). According to their neurochemical code, ruminal myenteric neurones were divided into three populations: ChAT/SP (68% of all myenteric neurones), NOS/VIP (26% of all myenteric neurones) and ChAT/- (5% of all myenteric neurones). Application of DiI onto the circular or longitudinal muscle revealed on average 64 or 44 labelled cell bodies in the myenteric plexus, respectively. DiI-labelled neurones expressed the code ChAT/SP or NOS/VIP. In the pathways to circular or longitudinal muscle, ChAT/SP-positive neurones outnumbered NOS/VIP-immunoreactive neurones by 5:1 and 2:1. Pathways to the circular or longitudinal muscle did not exhibit any pronounced polarized innervation patterns. This study demonstrated specific projections of myenteric neurones to the ruminal muscle. Neurones expressing the code ChAT/SP might function as excitatory muscle motor neurones, whereas NOS/VIP neurones are likely to act as inhibitory muscle motor neurones.     

Shapes and projections of tertiary plexus neurons of the guinea-pig small intestine

The axons of neurons that innervate the longitudinal muscle of the small intestine in small mammals such as rabbit, rat, guinea pig and mouse form a network, the tertiary plexus, against the inner surface of the muscle. In general, because of their substantial overlap, it has not been possible to follow the ramifications of individual axons in the tertiary plexus. In the present work, the longitudinal muscle motor neurons were filled with marker dyes through an intracellular microelectrode, and their morphologies and projections were examined in whole-mount preparations of longitudinal muscle and myenteric plexus. Most neurons that were examined were in the small intestine (ileum and duodenum), but a few were examined in the distal colon. Neurons in all regions had similar morphologies and projections. The cell bodies were amongst the smallest in myenteric ganglia, with major and minor axes of 14 microns and 25 microns (mean, n = 40) in the plane of the myenteric plexus. Each neuron had a single axon that branched extensively in the tertiary plexus, most had multiple lamellar dendrites and a few had filamentous dendrites or a mixture of filamentous and lamellar dendrites. The mean area of muscle covered by an axon and its branches extended 1.6 mm orally to anally and 1.7 mm circumferentially. The area covered was 2.8 +/- 1.9 mm2 (mean +/- SD, n = 23). From the density of occurrence of cell bodies, it can be calculated that each point in the longitudinal muscle is innervated by the processes of about 100 motor neurons and is influenced by electrotonic conduction of signals through the muscle by about 300 motor neurons.     

Presynaptic nicotinic acetylcholine receptors in the myenteric plexus of guinea pig intestine

Presynaptic nicotinic acetylcholine receptors (nAChRs) were studied in myenteric plexus preparations from guinea pig ileum using intracellular electrophysiological methods. Microapplication of nicotine (1 mM) caused a biphasic depolarization in all AH neurons (n = 30) and in 36 of 49 S neurons. Cytisine (1 mM) caused fast depolarizations in S neurons and no response in AH neurons. Mecamylamine (10 microM) blocked all responses caused by nicotine and cytisine. TTX (0.3 microM) blocked slow excitatory synaptic potentials in S and AH neurons but had no effect on fast depolarizations caused by nicotine. Nicotine-induced slow depolarizations were reduced by TTX in two of twelve AH neurons (79% inhibition) and four of nine S neurons (90+/-12% inhibition). Slow nicotine-induced depolarizations in the remaining neurons were TTX resistant. TTX-resistant slow depolarizations were inhibited after neurokinin receptor 3 desensitization caused by senktide (0.1 microM); senktide desensitization inhibited the slow nicotine-induced depolarization by 81+/-5% and 63+/-15% in AH and S neurons, respectively. A low-calcium and high-magnesium solution blocked nicotine-induced slow depolarizations in AH neurons. In conclusion, presynaptic nAChRs mediate the release of substance P and/or neurokinin A to cause slow depolarizations of myenteric neurons.     

Calbindin neurons of the guinea-pig small intestine: quantitative analysis of their numbers and projections

Summary The distribution of nerve cells with immunoreactivity for the calcium-binding protein, calbindin, has been studied in the small intestine of the guinea-pig, and the projections of these neurons have been analysed by tracing their processes and by examining the consequences of nerve lesions. The immunoreactive neurons were numerous in the myenteric ganglia; there were 3500±100 reactive nerve cells per cm² of undistended intestine, which is 30% of all nerve cells. In contrast, reactive nerve cells were extremely rare in submucous ganglia. The myenteric nerve cells were oval in outline and gave rise to several long processes; this morphology corresponds to Dogiel's type-II classification. Processes from the cell bodies were traced through the circular muscle in perforating nerve fibre bundles. Other processes ran circumferentially in the myenteric plexus. Removal of the myenteric plexus, allowing time for subsequent fibre degeneration, showed that reactive nerve fibres in the submucous ganglia and mucosa came from the myenteric cell bodies. Operations to sever longitudinal or circumferential pathways in the myenteric plexus indicated that most reactive nerve terminals in myenteric ganglia arise from myenteric cell bodies whose processes run circumferentially for 1.5 mm, on average. It is deduced that the calbindin-reactive neurons are multipolar sensory neurons, with the sensitive processes in the mucosa and with other processes innervating neurons of the myenteric plexus.     

Interstitial cells of Cajal: mediators of communication between circular and longitudinal muscle layers of canine colon

The network of interstitial cells of Cajal associated with Auerbach’s (myenteric) plexus in the canine colon was investigated to determine its role in facilitating communication between circular and longitudinal muscle layers. Electrical coupling between the muscle layers was demonstrated by propagating extracellularly evoked electrotonic pulses from circular muscle cells to nearby longitudinal muscle cells. The likelihood of cytoplasmic continuity across Auerbach’s plexus was further demonstrated by the ability of neurobiotin to spread between the interstitial cells and the circular and longitudinal muscle cells. Importantly, direct neurobiotin spread between circular and longitudinal muscle cells was not observed even when they were in close proximity as determined by confocal microscopy. When neurobiotin did spread across the two muscle layers, the intervening interstitial cells were always neurobiotin-positive. In regions where circular and longitudinal muscle cells approach each other closely, electron microscopy revealed the presence of close appositions between interstitial cells and smooth muscle cells. Gap junctions between interstitial cells and smooth muscle cells of both layers, as judged by electron microscopy, were extremely rare. Neither gap junctions nor close appositions were observed between longitudinal and circular muscle cells. The special arrangement for electrotonic coupling across Auerbach’s plexus through interstitial cells of Cajal suggests controlled coupling between the two muscle layers, explaining the preservation of their distinct electrical activities. Received: 21 July 1995 / Accepted: 22 April 1998     

Interactions of neurogenic responses of longitudinal and circular muscle in the guinea-pig ileum

The relationship between neurogenic responses of longitudinal and circular muscle was studied by measuring contractions and EMG or nonadrenergic, non-cholinergic (NANC) relaxations and NANC inhibitory junction potentials in different preparations of the guinea-pig ileum. NANC relaxation of longitudinal muscle was observed also without any preceding or concomitant circular muscle contraction ruling out the possibility that the latter might be the cause of the NANC relaxation. Circular muscle twitches or powerful contractions were absent if there was no preceding neurogenic or myogenic excitation of longitudinal muscle; in preparations with myenteric plexus-longitudinal muscle layers removed only small residual responses were seen although still under neurogenic influences. Thus excitation of longitudinal muscle seemed a prerequisite for synchronized and powerful contractions of circular muscle to occur. Cholinergic contraction and NANC relaxation of longitudinal muscle evoked by field stimulation were partly inhibited if the submucous plexus was also present suggesting the involvement of a more complex neuronal circuitry in these responses.     

Projections of excitatory and inhibitory motor neurones to the circular and longitudinal muscle of the guinea pig colon

The aim of this study was to identify myenteric pathways to the circular and longitudinal muscle of the guinea pig proximal colon. To identify excitatory and inhibitory muscle motoneurones, we applied the neuronal retrograde tracer DiI onto the circular or longitudinal muscle layer and performed additional immunohistochemistry for nitric oxide synthase (NOS) and choline acetyltransferase (ChAT). On average 166 +/- 81 circular muscle motoneurones (CMMN) and 100 +/- 74 longitudinal muscle motoneurones (LMMN) were labelled by DiI tracing. Myenteric pathways innervating the muscle were either ascending (DiI-labelled neurones with oral projections) or descending (DiI-labelled neurones with anal projections). The circular muscle was preferentially innervated by ascending pathways (66.0 +/- 9.1%). Most ascending CMMN were ChAT-positive (87.2 +/- 8.5%), whereas descending CMMN were mainly NOS-positive (82.3 +/- 14.6%). Most ascending (62.2 +/- 11.1%) and descending (82.0 +/- 12.5%) CMMN had circumferential projection preferences (circumferential projections were longer than projections along the longitudinal gut axis). In contrast to the polarised projections to the circular muscle, the longitudinal muscle was equally innervated by ascending (46.2 +/- 15.1%) and descending (53.9 +/- 15.1%) neurones. Ascending and descending pathways to the longitudinal muscle consisted predominantly of ChAT-positive neurones (98.1 +/- 1.9% and 68.0 +/- 8.5%, respectively), and both pathways had prominent longitudinal projection preferences. Only 25.5% of the descending LMMN were NOS-positive. In conclusion, the circular muscle in the proximal colon is innervated by descending inhibitory (NOS-positive neurones) and ascending excitatory (ChAT-positive neurones) pathways. In contrast, the longitudinal muscle is primarily innervated by ascending and descending excitatory motoneurones, and only a small proportion of the descending pathway consisted of inhibitory motoneurones.     

Evidence that myenteric neurons of the gastric corpus project to both the mucosa and the external muscle: myectomy operations on the canine stomach

Summary The distribution of nerve cell bodies and fibres in the canine stomach was investigated using antibodies to the general neuronal marker, neuron-specific enolase. Prominent ganglia containing many reactive nerve cells were found in the myenteric plexus of the gastric corpus and antrum. Nerve cells were absent from the submucosa of the corpus and were extremely rare in the antrum. Renoval of areas of longitudinal muscle and myenteric plexus from the corpus (myectomy), with 7 days allowed for axon degeneration, resulted in the loss of fibres reactive for galanin, gastrin-releasing peptide, substance P and vasoactive intestinal peptide from both the circular muscle and mucosa in the area covered by the lesion. Combined vagotomy and sympathetic denervation did not significantly affect these fibres, but did cause fibres reactive for calcitonin gene-related peptide to degenerate. It is concluded that the myenteric plexus of the gastric corpus, like the myenteric plexus of the small intestine and colon, is the source of nerve fibres innervating the circular muscle, but, in contrast to other regions of the gastrointestinal tract, myenteric ganglia, not submucous ganglia, are the major, or sole, source of the intrinsic innervation of the mucosa.     

Vasoactive intestinal polypeptide induces neurogenic contraction of guinea-pig ileum. Involvement of acetylcholine and substance P.

The effect and mode of action of vasoactive intestinal polypeptide (VIP), a peptidergic neuromodulator in the gastrointestinal nervous system, were investigated in isolated muscle strips of the guinea-pig ileum. VIP induced concentration-dependent (20 nM-1 microM) contractions of longitudinal ileal strips. TTX (1 microM), a mixture of atropine (3 microM) and spantide (30 microM), a mixture of atropine (3 microM) and omega-conotoxin GVIA (100 nM), somatostatin (60 nM) and dynorphin (100 nM) abolished the effect of VIP. In most cases a small relaxation became evident. Desensitization to substance P in the presence of atropine prevented VIP-induced contraction. A partial inhibition was observed in the presence of atropine (3 microM), spantide (30 microM), omega-conotoxin GVIA (100 nM), beta-endorphin (265 nM), met-enkephalin (1100 nM) and a mixture of spantide (30 microM) and omega-conotoxin GVIA (100 nM). The action of VIP was not significantly modified by guanethidine (3 microM) or hexamethonium (150 microM). In circular ileal strips VIP (10-300 nM) caused concentration-dependent relaxations through a direct myogenic effect. These results indicate that the VIP produced contractions of the guinea-pig ileum are exclusively neurally mediated and involve a cholinergic as well as a noncholinergic-nonadrenergic (NANC) pathway. It is concluded that besides acetylcholine (Ach) VIP releases the peptidergic transmitter substance P from postganglionic nerve fibers of myenteric plexus. Opioid peptides and somatostatin modulate the activity of cholinergic and peptidegic nerves in the guinea-pig ileum. The release of substance P appears to depend completely on N-type voltage sensitive calcium channels.