Comparative analysis of the merino sheep and Iberian red deer abomasum during prenatal development.

The aim of this study is to describe differences in the ontogenesis of the abomasum in sheep (domestic ruminant) and deer (wild ruminant). Histomorphometric and immunohistochemical analysis were carried out on 50 embryos and fetuses of the sheep and 50 red deer from the first prenatal stages until birth. To compare similar periods of gestation in both species, we calculate the percentages of gestation. The appearance of the abomasum was earlier in the red deer (22% gestation) than in the sheep (25% gestation). Throughout development the epithelium happened sequentially, being of the types pseudostratified to simple cylindrical. This important modification was earlier in the red deer than the sheep. At 46% gestation in red deer and 50% in sheep, gastric pits were observed on the surface of abomasal folds. Our studies suggest a close link between the initial formation of these pseudoglandular structures and the clear separation of lamina propria and submucosa separated by de muscularis mucosae. At 54% gestation in red deer and at 60% in sheep, in the bottom of these pits the first outlines of glands were distinguishable. Finally, the presence of neuroendocrine and glial cells were detected in deer at earlier stages than in sheep.

A mechanism of excessive accumulation of abomasal gas in vagotomized cattle determined using fluoroscopy.

To better understand the mechanism of excessive gas accumulation in the abomasum in bovine abomasal displacement, we performed gastric fluoroscopy in vagotomized cattle. Fifteen 6-month-old Holstein steers were divided into three groups: a non-vagotomized control group (Group C; n=5), a ventral thoraco-vagotomized group (Group V; n=5), and a dorsal and ventral thoraco-vagotomized group (Group DV; n=5). These groups were examined by fluoroscopy before and during a 5-week observation period after surgery. In Group C, no change was observed throughout the observation period. In Group DV, immediately after surgery, reticuloruminal motility was completely absent and ruminal distention was seen. Two weeks after surgery, abnormal reticulum motility and increased gas accumulation in the abomasal body were noted. Abomasal dilatation was also observed. In Group V, 1 week after surgery, gas inflow into the abomasum and relatively normal reticulum motility were observed along with a rapid increase in abomasal gas. Abomasal dilatation was also observed. In addition, left-displaced abomasum occurred in one of the steers in this group. From these results, we concluded that one of the mechanisms of excessive gas accumulation in the abomasum is reticulum-mediated gas inflow from the rumen combined with vagotomy-induced hypomotility.

Role of nitrergic nerves in the regulation of motility of the omasum and abomasum in healthy sheep (Ovis aries).

We examined the physiological role of nitrergic nerves in the regulation of omasal and abomasal motility in conscious healthy sheep and omasal muscle specimens. Nitric oxide (NO)-donor, S-nitroso-acethyl-dl-penicillamine (SNAP, 3-30 nmol/kg per min, i.v.) significantly inhibited omasal electromyographic (EMG) activity, whereas it did not alter EMG activity in the abomasal antrum. However, NO synthase inhibitor, Nomega-nitro-l-arginine-methyl ester (L-NAME, 0.3-3.0 micromol/kg per min, i.v.) did not alter EMG activity of the omasum and abomasum. In the in vitro experiments, SNAP application (6-200 micromol/l) significantly inhibited bethanechol (10 micromol/l)-induced contraction of longitudinal and circular muscles of the omasum. L-NAME application (0.03-3.0 mmol/l) enhanced electric field stimulation-induced contractions of the circular muscles. The results suggest that the omasal muscles are responsive to exogenous NO and that nitrergic nerves innervate the circular muscle layer of the omasum, however, nitrergic nerves are not or scarcely involved the physiological regulation of omasal and possibly abomasal motility in healthy sheep.

New aspects in the pathogenesis of abomasal displacement.

Impaired abomasal motility and an increased accumulation of gas are prerequisites for displacement of the abomasum in the cow. Predisposing factors are the breed (e.g. Holstein-Friesian, Simmental-Red-Holstein cross breeds and Guernsey), genetic background, twin pregnancy, first weeks of lactation, metabolic disorders (ketosis, increased lipomobilisation, insulin resistance), high-concentrate and low-fibre diets, as well as other concomitant diseases, such as endometritis, mastitis and claw disorders. There does not appear to be a strong correlation between increased milk yield or endotoxaemia and abomasal displacement. Recent studies have focused on possible functional disorders of the enteric nervous system within the abomasal wall, since cattle with abomasal displacement have an increased activity of neuronal nitric oxide synthase, as well as decreased acetylcholine sensitivity. In addition, there appear to be significant differences between breeds in the levels of the neurotransmitters substance P (SP) and vasoactive intestinal peptide (VIP) in the abomasal wall. For example, SP (stimulatory) was significantly less in German Holsteins in comparison to the German Fleckvieh, whereas VIP (inhibitory) was markedly increased. These risk factors may explain why Holstein cows are more susceptible to abomasal displacement than other breeds.

Cholinergic and noncholinergic innervation of the smooth muscle layers in the bovine abomasum.

The intrinsic innervation of muscle layers in the mammalian gastrointestinal tract has been mainly studied in nonruminants. The aim of this study was to identify intrinsic motor neurones in the bovine abomasum that innervate the circular and longitudinal muscles. Circular (CMN) and longitudinal muscle motor neurones (LMN) were selectively labeled by application of the retrograde tracer 1,1'-didodecyl-3,3,3',3'-tetramethyl indocarbocyanine perchlorate (DiI) onto the muscle layers. The transmitter phenotype was determined by immunohistochemical detection of choline acetyltransferase (ChAT), nitric oxide synthase (NOS), and neurone-specific enolase (NSE). On average, the myenteric ganglia contained 61 +/- 19 NSE-positive cell bodies, of which 89% were ChAT-positive and 10% were NOS-positive. Only 0.7% of NSE-positive neurones (41 of 5,777) contained both ChAT and NOS. Application of DiI onto the circular and longitudinal muscles revealed on average 60 +/- 27 (n = 4) and 68 +/- 36 (n = 4), respectively, labeled cell bodies in the myenteric plexus. For the circular and longitudinal muscles the proportions of ascending to descending neurones were 76 : 24% and 54 : 46%, respectively. While most ascending CMN were ChAT-positive (96%), 51% of the descending CMN were ChAT-negative. All ascending and 95% of descending LMN were ChAT-positive. It was concluded that cholinergic excitatory innervation is predominant in both muscle layers of the abomasum. Whereas the circular muscle receives cholinergic excitatory and nitrergic inhibitory innervation, the longitudinal muscle is only innervated by cholinergic pathways. This innervation pattern is different from that in gastric muscle layers in monogastric animals.

An immunohistochemical study of endocrine cells in the abomasum of vagotomized calf.

The effect of thoraco-vagotomy on the distribution and frequency of chromogranin-, serotonin-, somatostatin- and gastrin-immunoreactive cells in the abomasum of the calf were investigated by immunohistochemistry. Calves were vagotomized at 1 week old and sampled 2 and 4 weeks later. The endocrine cells generally decreased in number in vagotomized calves as compared to non-operated control calves. However, the detailed responses of endocrine cells to vagotomy varied depending on the endocrine cell type, region of gastric mucosa, and period after vagotomy. The present result suggests that the vagus nerve has an influence on the intrinsic regulatory system by endocrine cell control in the ruminant abomasum.

The effects of vagotomy on the abomasum in calves: radiography and protein gene product 9.5 immunohistochemistry.

Abomasal disorders of calves with total vagotomy, operated on at 1 week old, were investigated with radiography and protein gene product (PGP) 9.5 immunohistochemistry. Radiographic findings indicated abomasal atony with dilatation in all calves 2 weeks after vagotomy, while 4 weeks after vagotomy abomasal dilatation was detected in 2 calves and another 2 calves showed dilatation and impaction. The densities of PGP 9.5-immunoreactive nerves in the tunica muscularis decreased significantly in the corpus region of the greater curvature 2 weeks after vagotomy and in the pyloric region of the lesser curvature 4 weeks after vagotomy, and it was at its lowest 4 weeks after vagotomy in all regions examined. In conclusion, abomasal dilatation and/or impaction in vagotomized calves confirmed by radiography were related with a decreased frequency of nerves in the tunica muscularis of the abomasum.

Vagotomy suppresses cephalic phase insulin release in sheep.

The effect of selective vagotomy of the abomasum, pylorus, duodenum and liver on insulin release during the cephalic phase of digestion was investigated in wethers and lactating ewes. Electrical stimulation of the cervical vagus nerves was carried out to test the completeness of the vagotomies performed. In experiment 1, using wethers, the abomasal, pyloric and duodenal branches (ADV; n = 7) or the hepatic, abomasal, pyloric and duodenal branches (HADV; n = 10) of the ventral and/or dorsal vagus nerves were cut; a third group of wethers underwent sham-operation (SO; n = 8). In experiment 2, vagotomy (ADV; n = 5) or sham-operations (SO; n = 5) were carried out in lactating ewes. Jugular blood was drawn before and after presentation of food for glucose and insulin determination (experiments 1 and 2) or before, during and after the electrical stimulation of the peripheral ends of the cut cervical vagus nerves in randomly selected lactating ewes (experiment 3: ADV = 3, SO = 3) and wethers (experiment 4: ADV = 4, HADV = 4, SO = 4), for determination of insulin only. Presentation of food caused an immediate and significant (P < 0.05) rise in plasma insulin levels in SO animals compared with ADV or HADV wethers (experiment 1) or ADV ewes (experiment 2) without any significant change in blood glucose concentrations. In comparison with the SO group the baseline-corrected areas under the insulin response curve were significantly (P < 0.05) smaller for the respective vagotomized groups for periods 1-2, 2-4 and 4-6 min (experiment 1) and 1-2 and 2-4 min (experiment 2) after presentation of food. Total area under the response curve for 10 min was significantly (P < 0.05) lower (experiment 1) and tended (P < 0.10) to be lower (experiment 2) for the vagotomized groups compared with that of the control groups. Direct electrical stimulation of the cervical vagus nerves raised plasma insulin concentrations to significantly (P < 0.05) higher levels in the SO ewes but not in the ADV ewes (experiment 3). It was also evident that in experiment 1, HADV did not have any additive effect over that achieved by ADV alone. These results indicate that the vagal innervation of the gut mediates insulin release during the cephalic phase of feeding in sheep. It is concluded that insulin secretion from the pancreatic -cells in response to either food-related reflex activation of the vagal nuclei in the hypothalamus or direct cervical vagus nerve stimulation is mediated through the vagal efferent fibres carried in the abomasal, pyloric and duodenal branches of the vagus nerves in sheep.

Neuropeptides in the myenteric ganglia and nerve fibres of the forestomach and abomasum of grey, white and black Karakul lambs.

Previous studies indicated large, thin-walled, milk-filled rumens in lethal grey and white Karakul lambs. There was also a significant decrease in the number and size of the myenteric plexuses and the number of ganglion cells in these lambs. The purpose of this study was to determine whether the myenteric ganglia of the affected lambs are functional, by testing for the presence of vaso-active intestinal peptide, somatostatin, neurotensin, neuropeptide Y, met-enkephalin, calcitonin gene-related peptide and substance P in the myenteric ganglia and nerve fibres in the forestomach and abomasum of grey, white and black Karakul lambs. Four 1-cm2 samples were taken from analogous areas of the wall of the rumen, reticulum, omasum and abomasum of five grey, five white and five black newborn Karakul lambs. They were pinned to wax squares, fixed for 18 h in Zamboni's fixative, dehydrated and rehydrated through graded alcohols and stored in phosphate-buffered saline. The outer longitudinal muscle layer of each sample of the rumen, reticulum, omasum and abomasum was separated from the rest of the tissue layers, stained for each of the seven neuropeptides by employment of the immunofluorescence technique, and studied with a Leitz Orthoplan fluorescent microscope. All the material studied tested positive for all the neuropeptides. It is concluded that all the peptides tested for were present in all the lambs and that the myenteric ganglia are therefore functional in the lethal lambs.

Electrophysiological characteristics of tension receptors in the abomasal antrum of sheep.

Unitary impulse activity from tension receptors in the abomasal antrum in sheep anaesthetized with chloralose was studied with concurrent measurements of isometric tension in the antral wall, intraluminal pressures and electromyography (EMG). Rhythmical background activity had a close correlation with intraluminal pressure changes and usually a close correlation with local increases in EMG activity and isometric tension. Impulse frequencies increased, but were not sustained, following changes from isotonic to isometric conditions. Tension receptors were excited by transmural compression, by antral distension and by local intra-arterial injections of acetylcholine (2.5-25 micrograms), pentagastrin (10-20 micrograms), 5-hydroxytryptamine (1-20 micrograms), adrenaline (10 micrograms) cholecystokinin-8 (1-5 micrograms) or bradykinin (0.5-20 micrograms), and by systemic injections of bethanecol (5-1000 micrograms). Local intra-arterial perfusions of atropine (50 micrograms) and hexamethonium (100 micrograms) did not abolish spontaneous antral movements or affect the enhanced impulse activity evoked by acetylcholine or 5-hydroxytryptamine. Most antral tension receptors were excited by increases in antral tension or passive distension; they therefore functioned 'in series' with the muscularis externa. The discharges of three tension receptors were 'off-loaded' early during the contraction evoked by local injections of pentagastrin (10-20 micrograms).

Ultrastructure of the myenteric ganglia in the rumen, reticulum, omasum and abomasum of grey, white and black Karakul lambs.

Homozygous grey and white Karakul lambs die after they have reached weaning age. This is due to a lethal gene causing digestive disturbances. Previous studies revealed that grey and white lambs have large, milk-filled rumens; the phenomenon was attributed to a significant decrease in the number of myenteric ganglia and neurons in the rumen wall. This study was undertaken to determine whether any morphological differences exist in the ultrastructure of the myenteric ganglia in the forestomach and abomasum of grey, white and black Karakul lambs. Samples of the forestomach and abomasum of grey, white and black Karakul lambs were prepared routinely for electron microscopy and studied with a Phillips electron microscope. No morphological differences could be detected in the structure of the components of the myenteric ganglia in the forestomachs and abomasums of grey, white and black Karakul lambs. It was therefore concluded that the lethal gene in grey and white Karakul lambs results in a paucity of the myenteric ganglia, but does not affect the ultrastructure of these structures.

A comparative histological study of the number and size of the myenteric ganglia and neurones in the fore-stomach and abomasum of grey, white and black Karakul lambs.

Homozygous grey Karakul lambs are born with a lethal genetic factor responsible for death at weaning age. When put on a high roughage diet under field conditions they develop distended, thin-walled rumens and sand impacted abomasa. Homozygous white Karakul lambs have a similar factor but survive for a longer period. Black Karakul lambs are not affected. The present study was undertaken to compare by image analysis the number and size of the myenteric ganglia, and the number of myenteric neurones in the walls of the fore-stomach and abomasum of 24-h-old grey, white and black Karakul lambs. One square centimetre samples were taken from analogous areas of the rumen, reticulum, omasum and abomasum of 38 embalmed Karakul lambs. Haematoxylin and eosin stained histological sections of each sample were studied with a Vids 2 Image Analyzer. One way analysis of variance indicated a significant difference between the groups regarding the number and size of the myenteric ganglia and in the number of myenteric neurones in the reticulum, rumen and abomasum. The number and size of the ganglia and the number of neurones was greatest in the black lambs and decreased progressively in the white and grey lambs. The omasum was not affected. It is suggested that the paucity of myenteric ganglia and neurones in the regions examined is instrumental in causing the lethal condition described above.

[Emptying of the abomasum in adult ruminants]. / Evacuation de la caillette chez les ruminants adultes.

In the adult ruminant, abomasal emptying is a permanent phenomenon depending upon meal volume. Intradian rhythm involving the motor pattern of the duodenum and circadian rhythm of unknown origin modulate the transpyloric flow rate. The fundic tone, antro-duodenal coordination and pyloric resistance regulate gastric outflow. The break-like function of the pyloric resistance involves chyme viscosity. Transpyloric flow rate is controlled by a hierarchy of extrinsic and intrinsic mechanisms triggered at the duodenal level. The vagus permanently inhibits the motility of the abomasum. A similar relationship is observed between the pyloric sphincter and duodenal motility. Removal of the pyloric ring leads to an increased food intake.

Hexamethonium: a probe to assess autonomic nervous system involvement in upper gastrointestinal functions in conscious sheep.

Hexamethonium, which inhibits cholinergic transmission by preventing acetylcholine release, has been considered an ideal reference drug for the blockade of autonomic ganglia, Auerbach plexus and reflex gastrointestinal secretions. The degree of inhibition of ruminant gastrointestinal functions with this reference drug were as follows: cyclical contractions of the reticulo-rumen and abomasal motility greater than gastric acid secretion and duodenal migrating myoelectrical complexes. Although reduced at high dosages, the initiation of migrating myoelectric complexes was enhanced at clinically used dosages. The duration of the inhibition of reticular contractions was dose-related varying from 0.5 to 5 h for 1.25 to 20 mg/kg subcutaneously. Abomasal motility and acid secretion were similarly reduced but exhibited strong and long-lasting rebound effects. Inhibition of the reticulum by the blockade of muscarinic receptors by atropine was also dose-related lasting from 0.5 to 3 h for 0.5 to 2 mg/kg, whereas inhibition of the abomasal motor and secretory functions lasted from 1 to 6 h. These results suggest a higher degree of impingement of the parasympathetic pathways on abomasal acid secretion and motility than on the cyclical activity of the reticulum and only a modulatory role of the extrinsic neural activity on the cyclical motor events of the duodenum.

Nervous regulation of abomasum motor activity in sheep.

Pharmacological studies of abomasum motor activity were carried out in 3 sheep with chronic fistulae in the rumen and the fundus and pyloric part of the abomasum. The contractions of these parts of the stomach were recorded by the balloon method. The obtained results showed that in the abomasum stimulation of alpha-adrenergic receptors raises the degree of motor activity while stimulation of beta-adrenergic receptors inhibits it. It has been shown also that the sympathetic system has a greater influence through the beta-adrenergic receptors on the motor activity of the pyloric part than on that of the fundus of the abomasum. The analysis of the records obtained after bilateral vagotomy demonstrated that the automatism of the pyloric part was greater than that of the fundus of the abomasum.

[Distribution of the vagus nerve of the stomach and certain lymph nodes of the sheep in the prenatal period (author's transl)]. / Die Versorgung des Magens und einiger Lymphknoten durch den Nervus vagus beim Schaf in der pränatalen Periode.

The goal of this study was the investigation into the vagus nerve structure in the ruminant stomach, and a description of the participation of this nerve in the stimulation of the atrial lymph nodes of the rumen and dorsal abomasal lymph nodes. This survey was carried out on 30 sheep fetuses, whose ages are designated according to Zietzschmann and Krölling (1955). The fetuses came from the third to fifth months of pregnancy. Observations and general morphological nature may be summarized as follows: 1. to the wall of the rumen reach atrial branches coming from both vagus trunks plus a dorsal ruminal branch, and a right ruminal branch coming from the dorsal vagus trunk, 2. the reticulum receives reticular branches, coming from both vagal trunks, 3. the omasum is innervated by omasal branches of the dorsal vagus trunk plus an omasoabomasal branch coming from the ventral vagus trunk, 4. to the wall of the abomasum reach the reticulo-abomasal and pyloric branches coming from the ventral vagus trunk plus a branch to the major abomasal curvature and a visceral abomasal branch coming from the dorsal vagus trunk, 5. arterial lymph nodes of the rumen receive innervation from the reticulo-abomasal branch or the pyloric branch, both leaving from the ventral vagus trunk, 6. dorsal abdominal lymph nodes innervated by nerve branches coming from a parietal abomasal branch, coming also from the ventral vagus trunk.

A radiological study of gastric (abomasal) emptying in calves before and after vagotomy.

1. Gastric emptying has been studied in the conscious, standing calf by lateral radiography and fluoroscopy of radiopaque meals instilled into the abomasum before and after vagotomy. 2. Bilateral cervical vagotomy proved to be the only certain way of achieving total vagal transection. By instillation of milk into the abomasum through a cannula calves were maintained in normal, healthy condition for up to 36 days after vagotomy. 3. Motility of the antrum was not impaired by vagotomy so that some movement of gastric chyme to the duodenum occurred within minutes of instillation into the abomasum. Complete transference of the test meal was, however, delayed after vagotomy. 4. The greatest effect of vagotomy appeared to be on the abomasal body so that inadequate amounts of chyme were transferred to the antrum for pumping to the duodenum. 5. Delay in passage of contrast material through the intestine was related to delay in gastric emptying although vagotomy may have affected the intestine directly. 6. Following vagotomy the abomasum showed a resumption of normal motility and emptying after 7-29 days. This effect of vagotomy is similar to that seen in the simple stomach and is probably due to the establishment of intrinsic gastric control.

Pharmacological analysis of the abomasum-rumen reflex in sheep.

Contractions of the dorsal sac of the rumen and abomasum were recorded in the sheep with chronic fistulae. It was found that during reflex inhibition of rumen contractions, induced with stimulation of chemoreceptors in the abomasum, an increased sympathicotonia plays an essential role and the beta-adrenergic receptors mediate the reflex.

The effect of some molecules and ions on gastric function in the milk-fed calf.

1. The abomasum of the milk-fed calf has been examined using an adaptation of the Serial Test Meal method devised by Hunt & Spurrell (1951). The emptying process, acid secretion and pepsin secretion were studied.2. Using serial test meals of simple solutions instilled into the abomasum via a cannula, our investigation leaves no doubt that the osmolarity of the abomasal contents significantly modifies the rate of abomasal emptying.3. Hypotonic and isotonic solutions of sodium chloride and sodium bicarbonate increase abomasal emptying but bicarbonate is most effective.4. Increasing the concentration of solutes in the abomasal contents slows abomasal emptying. Sodium chloride, sodium bicarbonate, ammonium chloride and urea do not delay abomasal emptying until hypertonic concentrations are attained. Hypotonic solutions of potassium chloride, calcium chloride, glucose, lactose, hydrochloric acid and acetic acid delay abomasal emptying.5. The results obtained in the calf show that the abomasum is under restraint probably from duodenal receptors as is the simple stomach (Hunt & Knox, 1968) and that an osmoreceptor as postulated by Hunt (1956) is an important factor in this mechanism.6. Acid secretion is inhibited when hypertonic solutions are instilled into the abomasum.7. Pepsin secretion is not affected by simple solutions in the abomasum.8. Gastric function in the milk-fed calf appears to be controlled by mechanisms essentially similar to those already demonstrated in the simple stomach.