Functional vagal input to gastric myenteric plexus as assessed by vagal stimulation-induced Fos expression

Immunohistochemical detection of c-Fos expression was used to identify gastric myenteric plexus neurons that receive excitatory input from vagal efferent neurons activated by electrical stimulation of the cervical vagi in anesthetized rats. Vagal stimulation-induced Fos expression increased with higher pulse frequency, so that with 16 Hz (rectangular pulses of 1 mA/0.5 ms for 30 min) approximately 30% and with 48 Hz 90% of all neurons near the lesser curvature were Fos positive. In sham-stimulated rats there was no Fos expression. The percentage of Fos-activated neurons was only slightly smaller (85% with 48 Hz) near the greater curvature. Prior atropine administration (1 mg/kg ip) had little effect on vagal stimulation-induced Fos expression, and in unilaterally stimulated rats there was no Fos expression on the contralateral (noninnervated) side of the stomach, ruling out mediation by gastric motility or secretory responses. However, polysynaptic recruitment of third- and higher-order neurons cannot be ruled out completely. These results support the idea that, at least in the stomach, functional excitatory innervation of myenteric plexus neurons by the efferent vagus is profuse and widespread, refuting the notion of only a few vagal "command neurons."     

The longitudinal smooth muscle layer of the pig small intestine is innervated by both myenteric and submucous neurons

Originally, intestinal motility was thought to be exclusively regulated by myenteric neurons. Some years ago, however, it was demonstrated in large mammals that submucous neurons also participate in the innervation of the circular smooth muscle layer. To date, no information is available about the submucous innervation of the longitudinal smooth muscle layer (LM). This study provides evidence that in the small intestine of large mammals, the LM is innervated not only by the myenteric plexus, but also by the inner and outer submucous plexuses (ISP and OSP). In the porcine small intestine, the involved neurons can be subdivided into the following neurochemically distinct populations: leu-enkephalin (ENK)- and/or substance P (SP)-IR neurons and nitric oxide synthase (NOS)- and/or vasoactive intestinal polypeptide (VIP)-IR neurons. In the myenteric plexus, the majority of VIP- and/or NOS-IR neurons and ENK(+)/SP(-)-IR neurons exhibit descending projections, whereas ENK(+)/SP(+)-IR neurons preferentially have ascending projections. The ENK(-)/SP(+)-IR neurons do not show a polarized pattern. In the OSP, only ENK(+)/SP(-)- and VIP(+)/NOS(-)-IR neurons display a polarized (descending) projection pattern, whereas no polarization can be noted in the ISP. Morphological analysis of the traced neurons revealed that, in general, myenteric descending LM motor neurons have larger cell bodies than ascending ones and, in addition, myenteric descending VIP- and/or NOS-IR neurons have longer projections than ENK and/or SP-IR neurons. In conclusion, the present study demonstrates the involvement of not only myenteric, but also submucous neurons in the innervation of the LM. The two major populations are descending nitrergic neurons and ascending tachykinergic motor neurons, but also other subpopulations with specific projection patterns and neurochemical features have been identified.     

Choline acetyltransferase immunoreactivity of putative intrinsic primary afferent neurons in the rat ileum

The colocalisation of choline acetyltransferase (ChAT) with markers of putative intrinsic primary afferent neurons was determined in whole-mount preparations of the myenteric and submucosal plexuses of the rat ileum. In the myenteric plexus, prepared for the simultaneous localisation of ChAT and nitric oxide synthase (NOS), all nerve cells were immunoreactive (IR) for ChAT or NOS, but seldom for both; only 1.6 +/- 1.8% of ChAT-IR neurons displayed NOS-IR and, conversely, 2.8 +/- 3.3% of NOS-IR neurons were ChAT-IR. In preparations double labelled for NOS-IR and the general nerve cell marker, neuron-specific enolase, 24% of all nerve cells were immunoreactive for NOS, indicating that about 75% of all nerve cells have ChAT-IR. All putative intrinsic primary afferent neurons in the myenteric plexus, identified by immunoreactivity for the neurokinin 1 (NK1) receptor and the neurokinin 3 (NK3) receptor, were ChAT-IR. Conversely, of the ChAT-IR nerve cells, about 45% were putative intrinsic primary afferent neurons (this represents 34% of all nerve cells). The cell bodies of putative intrinsic primary afferent neurons had Dogiel type II morphology and were also immunoreactive for calbindin. All, or nearly all, nerve cells in the submucosal plexus were immunoreactive for ChAT. About 46% of all submucosal nerve cells were immunoreactive for both neuropeptide Y (NPY) and calbindin; 91.8 +/- 10.5% of NPY/calbindin cells were also ChAT-IR and 99.1 +/- 0.7% were NK3 receptor-IR. Of the nerve cells with immunoreactivity for ChAT, 44.3 +/- 3.8% were NPY-IR, indicating that about 55% of submucosal nerve cells had ChAT but not NPY-IR. Only small proportions of the ChAT-IR, non-NPY, nerve cells had NK3 receptor or calbindin-IR. It is concluded that about 45% of submucosal nerve cells are ChAT/calbindin/NPY/VIP/NK3 receptor-IR and are likely to be secretomotor neurons. Most of the remaining submucosal nerve cells are immunoreactive for ChAT, but their functions were not deduced. They may include the cell bodies of intrinsic primary afferent neurons.     

Central vagal stimulation activates enteric cholinergic neurons in the stomach and VIP neurons in the duodenum in conscious rats

The influence of central vagal stimulation induced by 2h cold exposure or intracisternal injection of thyrotropin-releasing hormone (TRH) analog, RX-77368, on gastro-duodenal enteric cholinergic neuronal activity was assessed in conscious rats with Fos and peripheral choline acetyltransferase (pChAT) immunoreactivity (IR). pChAT-IR was detected in 68%, 70% and 73% of corpus, antrum and duodenum submucosal neurons, respectively, and in 65% of gastric and 46% of duodenal myenteric neurons. Cold and RX-77368 induced Fos-IR in over 90% of gastric submucosal and myenteric neurons, while in duodenum only 25-27% of submucosal and 50-51% myenteric duodenal neurons were Fos positive. In the stomach, cold induced Fos-IR in 93% of submucosal and 97% of myenteric pChAT-IR neurons, while in the duodenum only 7% submucosal and 5% myenteric pChAT-IR neurons were Fos positive. In the duodenum, cold induced Fos in 91% of submucosal and 99% of myenteric VIP-IR neurons. RX-77368 induces similar percentages of Fos/pChAT-IR and Fos/VIP-IR neurons. These results indicate that increased central vagal outflow activates cholinergic neurons in the stomach while in the duodenum, VIP neurons are preferentially stimulated.     

Chemical profile of vagal preganglionic motor cells innervating the airways in ferrets: the absence of noncholinergic neurons.

In ferrets, we investigated the presence of choline acetyltransferase (ChAT), vasoactive intestinal peptide (VIP), and markers for nitric oxide synthase (NOS) in preganglionic parasympathetic neurons innervating extrathoracic trachea and intrapulmonary airways. Cholera toxin beta-subunit, a retrograde axonal transganglionic tracer, was used to identify airway-related vagal preganglionic neurons. Double-labeling immunohistochemistry and confocal microscopy were employed to characterize the chemical nature of identified airway-related vagal preganglionic neurons at a single cell level. Physiological experiments were performed to determine whether activation of the VIP and ChAT coexpressing vagal preganglionic neurons plays a role in relaxation of precontracted airway smooth muscle tone after muscarinic receptor blockade. The results showed that 1) all identified vagal preganglionic neurons innervating extrathoracic and intrapulmonary airways are acetylcholine-producing cells, 2) cholinergic neurons innervating the airways coexpress ChAT and VIP but do not contain NOS, and 3) chemical stimulation of the rostral nucleus ambiguus had no significant effect on precontracted airway smooth muscle tone after muscarinic receptor blockade. These studies indicate that vagal preganglionic neurons are cholinergic in nature and coexpress VIP but do not contain NOS; their stimulation increases cholinergic outflow, without activation of inhibitory nonadrenergic, noncholinergic ganglionic neurons, stimulation of which induces airway smooth muscle relaxation. Furthermore, these studies do not support the possibility of direct inhibitory innervation of airway smooth muscle by vagal preganglionic fibers that contain VIP.     

消化管括约肌部VIP免疫活性神经细胞分布

应用免疫组织化学方法研究了食管下部,幽门和回盲部肌间神经丛内ＶＩＰ免疫活性神经细胞的分布。ＶＩＰ免疫活性神经细胞在括约肌部比相邻部位数量多。并用Ｏｐｅｎ－ｔｉＰ法测量了刺激迷走神经后食管下段括约肌部压力的变化。用高阈值参数电刺激迷走神经引起预先投给阿托品的狗食管下段括约肌部压力的降低;这样条件下延长迷走神经刺激引起肌间神经丛内ＶＩＰ免疫活性神经细胞数量明显增加。由此结果提示含有或产生ＶＩＰ的神经细胞可能接受迷走神经的控制。由于刺激节前迷走神经纤维可能作用到这些细胞。     

Intracisternal TRH analog induces Fos expression in gastric myenteric neurons and glia in conscious rats

Activation of gastric myenteric cells by intracisternal injection of the stable thyrotropin-releasing hormone (TRH) analog RX-77368, at a dose inducing near maximal vagal cholinergic stimulation of gastric functions, was investigated in conscious rats. Fos immunoreactivity was assessed in gastric longitudinal muscle-myenteric plexus whole mount preparations 90 min after intracisternal injection. Fos-immunoreactive cells were rare in controls (~1 cell/ganglion), whereas intracisternal RX-77368 (50 ng) increased the number to 24.8 +/- 1.8 and 26.8 +/- 2.2 cells/ganglion in the corpus and antrum, respectively. Hexamethonium (20 mg/kg sc) prevented Fos expression by 90%, whereas atropine (2 mg/kg sc) had no effect. The neuronal marker protein gene product 9.5 and the glial markers S-100 and glial fibrillary acidic proteins showed that RX-77368 induced Fos in both myenteric neurons and glia. Vesicular ACh transporter and calretinin were detected around the activated myenteric neurons. These results indicated that central vagal efferent stimulation by intracisternal RX-77368 activates gastric myenteric neurons as well as glial cells mainly through nicotinic ACh receptors in conscious rats.     

Canine vagus nerve stores cholecystokinin-58 and -8 but releases only cholecystokinin-8 upon electrical vagal stimulation

Cholecystokinin-58 has been shown to be the major form of cholecystokinin (CCK) released to the circulation upon lumenal stimulation of the small intestine in humans and dogs. In anesthetized dogs, electrical vagal stimulation evokes pancreatic exocrine secretion that is in part mediated through the release of CCK. We studied the molecular form of CCK stored in canine vagus nerves and that released into circulation upon electrical vagal stimulation. Gel filtration and radioimmunoassay of the water and acid extracts of canine vagus nerves indicated CCK-8 (35%) and CCK-58 (65%) as the major molecular forms in the vagus nerve. Both forms of CCK isolated from the vagal extracts were equally bioactive as the standard CCK-8 and CCK-58, respectively, in stimulation of amylase release from isolated rat pancreatic acini. Analysis of plasma collected after electrical vagal stimulation indicated that CCK-8 is the only form released into the circulation. The release of CCK-8 upon electrical vagal stimulation was not affected by application of lidocaine to the upper small intestinal mucosa, suggesting that it was released from vagal nerve terminals.     

Distribution of nitric oxide synthase and vasoactive intestinal polypeptide immunoreactivity in the sphincter of Oddi and duodenum of the possum

The nitrergic innervation of the sphincter of Oddi (SO) and duodenum in the Australian brush-tailed possum and the possible association of this innervation with the neuropeptide vasoactive intestinal polypeptide (VIP) were investigated by using immunohistochemical localisation of nitric oxide synthase (NOS) and VIP, together with the general neuronal marker, protein gene product 9.5 (PGP9.5). Whole-mount preparations of the duodenum and attached SO without the mucosa, submucosa and circular muscle (n=12) were double- and triple-labelled. The density of myenteric nerve cell bodies of the SO in the more distal region (duodenal end) was significantly higher than that in the more proximal region. In the SO, approximately 50% of all cells were NOS-immunoreactive (IR), with 27% of the NOS-IR cells being VIP-IR. Within the duodenal myenteric plexus, NOS immunoreactivity was present in about 25% of all neurons, with 27% of these NOS-IR neurons also being VIP-IR, a similar proportion to that in the SO. Varicose nerve fibres with NOS and VIP immunoreactivity were present within the myenteric and submucous plexuses of the SO and duodenum, and in the circular and longitudinal muscle layers. The NOS-positive cells within both the SO and duodenum were unipolar, displaying a typical Dogiel type I morphology. The myenteric plexuses of the SO and duodenum were in direct continuity, with many interconnecting nerve trunks, some of which showed NOS and VIP immunoreactivity. Thus, the possum possesses an extensive NOS innervation of the SO and duodenum, with a significantly higher proportion of NOS-IR neurons within the SO, a subset of which contains VIP.     

Spatial organization of neurons in the dorsal motor nucleus of the vagus synapsing with intragastric cholinergic and nitric oxide/VIP neurons in the rat

The dorsal motor nucleus of the vagus (DMV) contains preganglionic neurons that control gastric motility and secretion. Stimulation of different parts of the DMV results in a decrease or an increase in gastric motor activities, suggesting a spatial organization of vagal preganglionic neurons in the DMV. Little is known about how these preganglionic neurons in the DMV synapse with different groups of intragastric motor neurons to mediate contraction or relaxation of the stomach. We used pharmacological and immunohistochemical methods to characterize intragastric neural pathways involved in mediating gastric contraction and relaxation in rats. Microinjections of L-glutamate (L-Glu) into the rostral or caudal DMV produced gastric contraction and relaxation, respectively, in a dose-related manner. Intravenous infusion of hexamethonium blocked these actions, suggesting mediation via preganglionic cholinergic pathways. Atropine inhibited gastric contraction by 85.5 +/- 4.5%. Gastric relaxation was reduced by intravenous administration of N(G)-nitro-L-arginine methyl ester (L-NAME; 52.5 +/- 11.9%) or VIP antagonist (56.3 +/- 14.9%). Combined administration of L-NAME and VIP antagonist inhibited gastric relaxation evoked by L-Glu (87.8 +/- 4.3%). Immunohistochemical studies demonstrated choline acetyltransferase immunoreactivity in response to L-Glu microinjection into the rostral DMV in 88% of c-Fos-positive intragastric myenteric neurons. Microinjection of L-Glu into the caudal DMV evoked expression of nitric oxide (NO) synthase and VIP immunoreactivity in 81 and 39%, respectively, of all c-Fos-positive intragastric myenteric neurons. These data indicate spatial organization of the DMV. Depending on the location, microinjection of L-Glu into the DMV may stimulate intragastric myenteric cholinergic neurons or NO/VIP neurons to mediate gastric contraction and relaxation.     

The distribution and co-localization of immunoreactivity to nitric oxide synthase,vasoactive intestinal polypeptide and substance P within nerve fibres supplying bovine and porcine female genital organs

The distribution of nitric oxide synthase-immunoreactive (NOS-IR) axons and their relationship to structures immunoreactive to vasoactive intestinal polypeptide (VIP), substance P (SP) and tyrosine hydroxylase (TH) were studied by means of the nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) technique or double-labelling immunofluorescence in the genital organs of cow and pig. Relevant neurons were also investigated in the pig. NOS-containing neural structures were TH-immunonegative in bovine or porcine genital organs, or in the studied ganglia. In the bovine ovary, NOS-IR nerves were neither VIP-IR nor SP-IR, whereas in the pig, most NOS-containing axons were also VIP-IR. The oviduct was supplied by single NOS/VIP- or NOS/SP-containing nerves, whereas in the uterus, NOS-IR axons were moderate in number, often being immunoreactive for VIP or SP. Numerous NOS/VIP-IR and NOS/SP-IR nerves were found in the vagina of both species. In all tissues studied, NOS-IR axons were mainly related to vascular smooth muscle. Most of the neurons of the paracervical ganglia and some neurons in dorsal root ganglia exhibited strong NOS activity. Only single neurons in sympathetic ganglia were NADPH-d-positive. Most nitrergic neurons in the autonomic ganglia were VIP-IR but SP-immunonegative. The sensory neurons were mostly NOS/SP-IR, whereas only single neurons co-expressed NOS and VIP immunoreactivity.     

大鼠下丘脑室旁核神经元对电刺激迷走神经的反应

用玻璃微电极记录了93只大鼠的1059个PVH单位的电活动,观察了电刺激颈部迷走神经对PVH单位自发放电的效应和所引起的PVH单位的诱发反应。电刺激迷走神经分别使46个及10个PVH单位呈诱发兴奋和抑制反应。给予迷走神经以不同强度的刺激时,发现PVH神经元对激活A和C两类纤维的强刺激反应,而对仅激活A类纤维的弱刺激则不反应。PVH单位对电刺激坐骨神经或迷走神经的反应有以下几种:对迷走神经和坐骨神经刺激均作出兴奋或抑制反应;仅对迷走刺激作出兴奋或兴奋-抑制反应,而对坐骨神经刺激不反应;对坐骨神经刺激作出兴奋反应,而对迷走神经刺激不反应。讨论了迷走神经到室旁核的中枢传导特点以及内脏传入和躯体传入信息在PVH单位会聚的可能意义。     

Vanilloid receptor (VR1) expression in vagal afferent neurons innervating the gastrointestinal tract

The vanilloid receptor VR1 is a nonselective cation channel activated by capsaicin as well as increases in temperature and acidity, and can be viewed as molecular integrator of chemical and physical stimuli that elicit pain. The distribution of VR1 receptors in peripheral and central processes of rat primary vagal afferent neurons innervating the gastrointestinal tract was investigated by immunohistochemistry. Forty-two percent of neurons in the nodose ganglia retrogradely labeled from the stomach wall expressed low to moderate VR1 immunoreactivity (VR1-IR). VR1-IR was considerably lower in the nodose ganglia as compared to the jugular and dorsal root ganglia. In the vagus nerve, strongly VR1-IR fibers ran in separate fascicles that supplied mainly cervical and thoracic targets, leaving only weakly VR1-IR fibers in the subdiaphragmatic portion. Vagal afferent intraganglionic laminar endings (IGLEs) in the gastric and duodenal myenteric plexus did not express VR1-IR. Similarly, VR1-IR was contained in fibers running in perfect register with vagal afferents, but was not colocalized with horseradish peroxidase in the same varicosities of intramuscular arrays (IMAs) and vagal afferent fibers in the duodenal submucosa anterogradely labeled from the nodose ganglia. Only in the gastric mucosa did we find evidence for colocalization of VR1-IR in vagal afferent terminals. In contrast, many nerve fibers coursing through the myenteric and submucosal plexuses contained detectable VR1-IR, the majority of which colocalized calcitonin gene-related peptide immunoreactivity. In the dorsal medulla there was a dense plexus of VR1-IR varicose fibers in the commissural, dorsomedial and gelatinosus subnuclei of the medial NTS and the lateral aspects of the area postrema, which was substantially reduced, but not eliminated on the ipsilateral side after supranodose vagotomy. It is concluded that about half of the vagal afferents innervating the gastrointestinal tract express low levels of VR1-IR, but that presence in most of the peripheral terminal structures is below the immunohistochemical detection threshold.     

Neuro-Anatomical Evidence Indicating Indirect Modulation of Macrophages by Vagal Efferents in the Intestine but Not in the Spleen

Background

Electrical stimulation of the vagus nerve suppresses intestinal inflammation and normalizes gut motility in a mouse model of postoperative ileus. The exact anatomical interaction between the vagus nerve and the intestinal immune system remains however a matter of debate. In the present study, we provide additional evidence on the direct and indirect vagal innervation of the spleen and analyzed the anatomical evidence for neuroimmune modulation of macrophages by vagal preganglionic and enteric postganglionic nerve fibers within the intestine.

Methods

Dextran conjugates were used to label vagal preganglionic (motor) fibers projecting to the small intestine and spleen. Moreover, identification of the neurochemical phenotype of the vagal efferent fibers and enteric neurons was performed by immunofluorescent labeling. F4/80 antibody was used to label resident macrophages.

Results

Our anterograde tracing experiments did not reveal dextran-labeled vagal fibers or terminals in the mesenteric ganglion or spleen. Vagal efferent fibers were confined within the myenteric plexus region of the small intestine and mainly endings around nNOS, VIP and ChAT positive enteric neurons. nNOS, VIP and ChAT positive fibers were found in close proximity of intestinal resident macrophages carrying α7 nicotinic receptors. Of note, VIP receptors were found on resident macrophages located in close proximity of VIP positive nerve fibers.

Conclusion

In the present study, we show that the vagus nerve does not directly interact with resident macrophages in the gut or spleen. Instead, the vagus nerve preferentially interacts with nNOS, VIP and ChAT enteric neurons located within the gut muscularis with nerve endings in close proximity of the resident macrophages.     

Catecholaminergic neurons in rat dorsal motor nucleus of vagus project selectively to gastric corpus

Nitric oxide synthase-immunoreactive (NOS-IR) neurons in the rat caudal dorsal motor nucleus of the vagus (DMV) project selectively to the gastric fundus and may be involved in vagal reflexes controlling gastric distension. This study aimed to identify the gastric projections of tyrosine hydroxylase-immunoreactive (TH-IR) DMV neurons, whether such neurons colocalize NOS-IR, and if they are activated after esophageal distension. Gastric-projecting neurons were identified after injection of retrograde tracers into the muscle wall of the gastric fundus, corpus, or antrum/pylorus before removal and processing of the brain stems for TH- and NOS-IR. A significantly higher proportion of corpus- compared with fundus- and antrum/pylorus-projecting neurons were TH-IR (14% compared with 4% and 2%, respectively, P < 0.05). Colocalization of NOS- and TH-IR was never observed in gastric-projecting neurons. In rats tested for c-Fos activation after intermittent esophageal balloon distension, no colocalization with TH-IR was observed in DMV neurons. These findings suggest that TH-IR neurons in the caudal DMV project mainly to the gastric corpus, constitute a subpopulation distinct from that of nitrergic vagal neurons, and are not activated on esophageal distension.     

Brain-gut interactions between central vagal activation and abdominal surgery to influence gastric myenteric ganglia Fos expression in rats

We previously showed that medullary thyrotropin-releasing hormone (TRH) or the stable TRH agonist, RX-77368 administered intracisternally induces vagal-dependent activation of gastric myenteric neurons and prevents post surgery-induced delayed gastric emptying in rats. We investigated whether abdominal surgery alters intracisternal (ic) RX-77368 (50 ng)-induced gastric myenteric neuron activation. Under 10 min enflurane anesthesia, rats underwent an ic injection of saline or RX-77368 followed by a laparotomy and a 1-min cecal palpation, or no surgery and were euthanized 90 min later. Longitudinal muscle/myenteric plexus whole-mount preparations of gastric corpus and antrum were processed for immunohistochemical detection of Fos alone or double labeled with protein gene-product 9.5 (PGP 9.5) and vesicular acetylcholine transporter (VAChT). In the non surgery groups, ic RX-77368 induced a 17 fold increase in Fos-expression in both gastric antrum and corpus myenteric neurons compared to saline injected rats. PGP 9.5 ascertained the neuronal identity of myenteric cells expressing Fos. In the abdominal surgery groups, ic RX-77368 induced a significant increase in Fos-expression in both the corpus and antrum myenteric ganglia compared with ic saline injected rats which has no Fos in the gastric myenteric ganglia. However, the response was reduced by 73-78% compared with that induced by ic RX 77368 without surgery. Abundant VAChT positive nerve fibers were present around Fos positive neurons. These results indicate a bidirectional interaction between central vagal stimulation of gastric myenteric neurons and abdominal surgery. The modulation of gastric vagus-myenteric neuron activity could play an important role in the recovery phase of postoperative gastric ileus.     

Unit responses of the cerebellar cortex to stimulation of the vagus nerve and nerves of the limbs

Responses of 74 cerebellar cortical neurons to electrical stimulation of the vagus nerve and its gastric branches and also of the limb nerves were recorded extracellularly in experiments on rats anesthetized with chloralose and pentobarbital. Phasic and tonic unit responses were similar to all types of stimulation, but fewer neurons responded to stimulation of the vagus nerve than to stimulation of the limb nerves. Monosensory neurons responding only to visceral stimulation, were not found. Among cells giving a phasic response, some had a short and others a long latent period. The latent period of responses of all types was longer to vagal stimulation than to stimulation of the limb nerves. The maximal frequency of rhythmic responses to vagal stimulation was 8/sec, compared with 16/sec for stimulation of the limb nerves. Tonic responses of the neurons were excitatory, inhibitory, and mixed. The type of unit response could vary depending on the frequency and repetitiveness of the stimuli but was otherwise independent of the type of stimulation.N. I. Pirogov Vinnitsa Medical Institute. I. M. Sechenov Institute of Evolutionary Biochemistry and Physiology, Academy of Sciences of the USSR, Leningrad. Translated from Neirofiziologiya, Vol. 4, No. 5, pp. 471–479, September–October, 1972.     

Fos expression in rat celiac ganglion: an index of the activation of postganglionic sympathetic nerves

To develop an index of the activation of abdominal sympathetic nerves, we used Fos immunostaining of the celiac ganglion (CG) taken from rats receiving nicotine, preganglionic nerve stimulation, or glucopenic agents. Subcutaneous nicotine injection moderately increased Fos expression in the principal ganglionic cells of the CG (17 +/- 4 Fos+ per mm(2), approximately 12% of all principal CG cells), whereas subcutaneous saline had no effect (0 +/- 0 Fos+ per mm(2); n = 7; P < 0.01). Greater Fos expression was obtained by applying nicotine topically to the CG (71 +/- 8 Fos+ per mm(2); 52% of all principal CG cells, n = 5; P < 0.01 vs. topical saline, n = 4) and by preganglionic nerve stimulation (126 +/- 9 Fos+ per mm(2); 94% of all principal CG cells, n = 11; P < 0.01 vs. nerve isolation, n = 7). Moderate Fos expression was also observed in the CG after intraperitoneal 2-deoxy-D-glucose (2DG) injection (21 +/- 2 Fos+ per mm(2); 16% of all principal CG cells, n = 5; P < 0.01 vs. saline ip) or insulin injection (16 +/- 2 Fos+ per mm(2); 12% of all principal CG cells, n = 6; P < 0.01 vs. saline ip). Furthermore, Fos expression induced by 2DG was dose and time dependent. These data demonstrate significant Fos expression in the CG in response to chemical, electrical, and reflexive stimulation. Thus Fos expression in the CG may be a useful index to describe various levels of activation of its postganglionic sympathetic neurons.     

Influence of peripheral targets on the expression of calcitonin gene-related peptide immunoreactivity in rat cranial motoneurones

Calcitonin gene-related peptide-like immunoreactivity (CGRP-ir) is displayed by motoneurons that innervate striated muscle but is absent from preganglionic parasympathetic motoneurons. One hypothesis to explain this is that CGRP gene expression in motoneurons is, in part, dependent on influences from the innervated organ. To test this hypothesis, we cross-anastomosed the right hypoglossal and cervical vagal nerves of rats so that the vagal motoneurons grew to innervate the musculature of the tongue. Following a recovery period of 17 to 52 weeks, the distribution of CGRP-ir in the dorsal motor vagal nucleus was determined in both cross-anastomosed animals and self-anastomosed control animals. Successful reinnervation of the tongue musculature by vagal motoneurons was demonstrated by showing that electrical stimulation of the central vagus/peripheral hypoglossal nerve produced a twitch of the tongue muscles. Motoneurones of the dorsal motor vagal nucleus, which now innervated the tongue were found to express CGRP-ir, which was evident from the double labeling of neurons with both horseradish peroxidase and CGRP-ir. Motoneurones of the dorsal motor vagal nucleus contralateral to the cross-anastomosis remained CGRP negative. Similarly, motoneurons of the dorsal motor vagal nucleus in control animals where the vagus nerve was self-anastomosed remained CGRP negative, showing that an induction of CGRP expression is not a result of nerve section itself. We suggest that a signal from the striated muscle transported retrogradely via the motor axon regulates expression of CGRP-ir in motoneurons. © 1995 John Wiley & Sons, Inc.     

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.