G. Liedberg

Effects of antrectomy or porta‐caval shunting on the histamine‐storing endocrine‐like cells in oxyntic mucosa of rat stomach. A fluorescence histochemical, electron microscopic and chemical study.

1. The argyrophil (enterochromaffin‐like) cells in the oxyntic gland area of the rat stomach contain histamine, which can be demonstrated fluorescence microscopically after exposure to gaseous OPT. After administration of L‐dopa (or L‐5‐hydroxytryptophan), these cells produce and temporarily store dopamine (or 5‐hydroxytryptamine), demonstrable by its characteristic formaldehyde‐induced fluorescence. Ultrastructurally, the enterochromaffin‐like cells, which have the appearance of polypeptide hormone‐secreting cells, comprise two main cell types, the most predominant one having vesicular type granules (EGL cells), the second most predominant one having smaller, uniformly electron dense granules (A‐like cells). 2. Rats were subjected to the following surgical treatments: antrectomy; porta‐caval shunting; antrectomy+porta‐caval shunting; or sham‐operation. Three to eight weeks after surgery the histamine‐storing cells (enterochromaffin‐like cells) of the oxyntic mucosa were analysed by fluorescence histochemistry, light and (quantitative) electron microscopy, and fluorometric determination of amines. 3. After antrectomy, fluorescence histochemistry and silver staining revealed a reduced number of enterochromaffin‐like cells. The histamine content in the oxyntic mucosa was reduced by about 50%. As in unoperated injection of pentagastrin seemed to mobilize histamine. Feeding or injection of insulin failed to do so in antrectomized as opposed to control rats. Ultrastructurally, the cytoplasmic granules of both endocrine‐like cell types were less numerous than in the unoperated rats. The reduction in cell number and granularity was particularly conspicuous with regard to the EGL cells. 4. After porta‐caval shunting the number of enterochromaffin‐like cells increased markedly. Chemical determination revealed a twofold increase in the histamine concentration of the oxyntic mucosa. Feeding or injection of insulin or pentagastrin lowered the histamine concentration. As judged by electron microscopy, the proliferation of endocrine‐like cells induced by porta‐caval shunting was restricted to the ECL cell type. Besides occurring in greater number, these cells were larger than those in unoperated controls, and their cytoplasm was densely packed with granules that were increased in size. 5. Following antrectomy of the porta‐caval shunted rats the number of enterochromaffin‐like cells and the oxyntic histamine concentration was reduced. 6. The results support the idea that gastrin exerts trophic as well as excitatory effects on oxyntic endocrine‐like cells.

Correlation between serum gastrin concentration and rat stomach histidine decarboxylase activity

After prolonged fasting the activity of histidine decarboxylase in the oxyntic mucosa of the rat stomach is low. Feeding or injection of gastrin or insulin rapidly raises the enzyme activity. It was earlier suggested that all enzyme‐activating agents act through release of gastrin. This view has found experimental support in studies which show that in antrectomized rats the enzyme is activated by gastrin but not by gastrin‐releasing stimuli like feeding or vagal excitation (insulin hypoglycemia). In the present investigation rats were subjected to a variety of treatments and serum gastrin concentrations and gastric histidine decarboxylase activities were measured. The main findings were as follows.

The role of endogenous gastrin in the activation of gastric histidine decarboxylase in the rat. Effect of antrectomy and vagal denervation

Abstract The activity of gastruc histidine decarboxylase in normal rats was markedly reduced after prolonged fasting. The reduction was reversed by feeding and by injection of gastrin, insulin (vagal stimulation) or carbachol. Feeding, vagal stimulation or treatment with parasympathomimetics released endogenous gastrin. The role of endogenous gastrin (believed to be located in the antrum) and of the vagus nerve in activating gastric histidine decarboxylase was studied in antrectomized and in vagally denervated rats. Antrectomy reduced the activation of gastric histidine decarboxylase induced by feeding and abolished that induced by the injection of insulin or carbachol (but not that induced by pentagastrin). Antral exclusion, on the other hand, caused a marked enzyme activation. These results indicate the existence of an antral agent (presumably gastrin) which mediated in the process of enzyme activation. The effect of antrectomy on gastric acid secretion was studied in rats fitted with chronic gastric fistulas. Antrectomy depressed basal acid secretion, but did not abolish the effect of insulin hypogylcemia on acid secretion. Vagal denervation (truncal vagotomy or selective vastric vagotomy) in combination with pyloroplasty caused a marked activation of the gastric histidine decarboxylase in fasted rats. This activation was totally abolished by combined vagotomy and antrectomy. Consequently, enzyme activation observed after vagotomy is induced by some agent from the antrum, possibly gastrin. The activation of gastric histidine decarboxylase seen upon truncal vagotomy was abolished not only by antrectomy but also by perfusion of the stomach with dilute hydrochloric acid. It is therefore suggested, that low antral pH inhibits the release of gastrin and that the elevated antral pH caused by vagotomy stimulates its release. In chronic fistula rats, vagotomy caused a profound decreased of acid secretion and inhibited the secretory response to insulin and pentagastrin. The following concept is advanced: The activity of gastric histidine decarboxylase is under the influence of endogenous gastrin. The influence of the vagus nerve on histidine decarboxylase is exerted via the gastrin-containing cell in the antrum and not by direct action on the cell containing the enzyme. The influence of the vagus on the gastrin cell has two facets, possibly constituting the components in a feed-back mechanism: a) vagal stimulation of the gastrin cell releases gastrin; b) vagal stimulation of the parietal cell causes acid hypersecretion, whicc inhibits gastrin release. Activation of gastrin histidine decarboxylase is a consequence of gastrin-induced stimulation of the enzyme-containing cell. Important factors regulating the gastrin release are the vagal activity and the antral pH.

Gastrin cell proliferation after chronic stimulation: effect of vagal denervation or gastric surgery in the rat.

Chronic stimulation of the antral gastrin cells by elevated antral pH was achieved by fundectomy, antrum exclusion, fundectomy plus antrum exclusion, antrocolic transposition, and vagal denervation plus pyloroplasty. For comparison we studied also the effects of pyloroplasty alone and of portacaval shunting. All operations that elevated the antral pH resulted in high gastrin concentrations in serum. Particularly high concentrations were observed in fundectomized rats. Vagal denervation of fundectomized or antrum excluded rats reduced the serum gastrin concentration slightly compared with the corresponding innervated animals. Portacaval shunting reduced the gastrin concentration in serum. The antral gastrin concentration was raised or unchanged following fundectomy and vagal denervation, and reduced following antrum exclusion, antrum exclusion plus vagotomy, fundectomy plus antrum exclusion, fundectomy plus vagotomy, antrocolic transposition and portacaval shunt. The gastrin cell density in the antral mucosa was raised following fundectomy, vagotomy, and fundectomy plus vagotomy, unchanged following fundectomy plus antrum exclusion and antrocolic transposition, and reduced following antrum exclusion and portacaval shunting. Ultrastructurally the gastrin (G) cells in the excluded antrum and in the antrum of fundectomized rats showed signs of secretory activity in that the granule volume density or the number of granules per unit cytoplasm was lowered. In the fundectomized rats moreover, the endoplasmic reticulum of the G cells was increased, the Golgi area enlarged and the proportion and volume density of electron dense granules greatly increased. The granule profile diameter was not affected by either antrum exclusion or fundectomy. The results on the excluded antrum indicate that elevated antral pH per se is not sufficient to produce gastrin cell proliferation. In the fundectomized rats, where the hyperlasia of antral gastrin cells was considerable, there is the added stimulus of ingested food. In fundectomized plus antrum excluded rats this stimulus is eliminated and no proliferation ensues. The passage of intestinal material, as in the rats subjected to antrocolic transposition, did not elicit gastrin cell proliferation which seems to suggest that the character of the luminal material is important. We propose therefore that gastrin cell proliferation is due to the combined stimulation of high antral pH and passage of food. Vagal innervation is not required.

Mechanisms of gastric acid secretion after pylorus and oesophagus ligation in the rat.

1. The effect of vagotomy on gastric acid secretion was studied in chronic gastric fistula rats at various times after denervation. In these rats basal and pentagastrin‐induced acid output was permanently reduced. Thus, the magnitude of the acid response to pentagastrin in the conscious fistula rat is dependent upon an intact vagus. 2. The acid response to pylorus ligation in vagally intact rats was unaffected by drainage of the stomach and therefore not caused by distension. Bilateral vagotomy, performed simultaneously with the ligation, completely abolished acid secretion, while unilateral vagotomy reduced the acid output by half. Hence, in innervated rats, an intact vagal impulse flow appears to be essential for the acid response to pylorus ligation. When the pylorus ligation was performed 2‐8 weeks after truncal vagotomy, the acid output showed a progressive return towards pre‐denervation values. In the denervated rats the acid response to pylorus ligation was blocked by drainage of the stomach and therefore probably caused by distension, a mechanism which is independent of the vagal impulse flow. 3. The response to pylorus ligation in innervated rats was blocked by atropine and chlorisondamine but not by metiamide. In the denervated rats, the response to pylorus ligation was blocked by all three drugs. 4. Following ligation of both the pylorus and the oesophagus the acid response was poor. With drainage of the oesophagus the acid response was much enhanced, suggesting that oesophageal distension inhibits acid secretion. In the vagotomized rat the poor acid response to oesophageal + pyloric ligation could not be overcome by drainage of the oesophagus. In the innervated rat gastric distension could overcome the inhibition induced by oesophageal ligation. Also in chronically, but not in acutely vagotomized rats, gastric distension brought about a good acid response. Conceivably, gastric reflex mechanisms can activate acid secretion through vagal and/or intramural pathways. Both in innervated and denervated rats the response to gastric distension was inhibited by atropine, chlorisondamine and metiamide. 5. The results suggest that in the innervated rat vago‐vagal reflexes are important for the gastric hypersecretion following ligation of the pylorus, and for the acid response to gastric distension following ligation of the pylorus and oesophagus. In the chronically vagotomized rat local intramural reflexes elicited by gastric distension are responsible for the acid response.

Effects of various gastrointestinal peptides on parietal cells and endocrine cells in the oxyntic mucosa of rat stomach.

1. The effects of secretin, glucagon, cholecystokinin‐pancreozymin (CCK‐PZ), gastric inhibitory peptide (GIP), vasoactive intestinal peptide (VIP), somatostatin, neurotensin and enkephalin on basal, pentagastrin‐ and histamine‐stimulated gastric acid secretion were investigated in the conscious fistula rat. 2. Glucagon and GIP were ineffective inhibitors of basal and pentagastrin‐stimulated secretion. CCK‐PZ stimulated acid secretion at a low dose level but at higher doses it inhibited both pentagastrin‐ and histamine‐induced secretions. VIP was ineffective at low doses and at high doses its action was complicated by reflux of stimulated pancreatic and intestinal secretions into the stomach. Met‐enkephalin inhibited histamine‐ but not pentagastrin‐stimulated secretion. Neurotensin inhibited the response to pentagastrin but had no effect on histamine‐stimulated secretion. Secretin and somatostatin were potent inhibitors of basal and pentagastrin‐stimulated acid secretion with little or no effect on the response to histamine. 3. At doses completely inhibitory to pentagastrin‐stimulated secretion secretin and somatostatin did not block the mobilization of gastric mucosal histamine by pentagastrin, although somatostatin caused partial competitive inhibition at lower doses of pentagastrin. Thus secretin and somatostatin inhibited pentagastrin‐induced secretion neither by blocking gastric mucosal histamine mobilization nor by abolishing the direct action of histamine on the parietal cell ‐‐ findings which are inconsistent with the proposed role of histamine as the mediator of the action of gastrin on the parietal cell.

Mechanism of activation of rat stomach histidine decarboxylase after vagal denervation

Abstract Vagal denervation causes a powerful activation of histidine decarboxylase in the oxyntic gland area of the rat stomach. It was decided to investigate the mechanism behind the activation following vagotomy and to identify the histamine-forming enzyme. A semi-purified preparation of gastric histidine decarboxylase from vagally denervated rats was found to be identical with the specific histidine decarboxylase previously characterized in extracts of gastric mucosa of normal rats and in tissues of the foetal rat. The pH optimum varied with the substrate concentration: 6.2 with 6 × 10−3 M histidine, 6.7 with 6 × 10−4 M histidine. The Michaelis-Menten constant increased with diminishing pH. Gastric histidine decarboxylase in the fasted rat was markedly activated after bilateral, but not after unilateral, vagal denervation. The enzyme activation normally seen in the rat stomach after feeding and after the injection of insulin persisted after unilateral denervation, both in the innervated and the denervated side of the stomach. These findings exclude a direct vagal regulation of the enzyme activity of the histamine-producing cell. Antrectomy abolished the enzyme activation following vagal denervation, suggesting that the enzyme activation is a consequence of enhanced release of some antral hormone, possibly gastrin. The release of gastrin is regulated by a variety of factors, one being the antral pH. Accordingly, perfusion of the stomach with hydrochloric acid markedly reduced the enzyme activity of vagotomized rats, whereas perfusion with phosphate buffer (pH 7.0) resulted in an increase in the enzyme activity of normal, fasted rats. Atropine and propantheline in doses sufficient to inhibit acid secretion activated the enzyme of normal, fasted rats. The enzyme-activating effect of atropine was abolished by antrectomy. Histamine, which stimulated acid secretion in normal rats but not in vagotomized rats, reduced the enzyme activity of normal, fasted rats, but not that of vagotomized rats. Histamine also inhibited the enzyme activation after feeding and after the injection of insulin but not that seen after the injection of pentagastrin. The following conclusion is drawn: All agents (except gastrin itself) and all experimental conditions which increase gastric acid secretion without directly affecting gastrin release, result in a reduction of the histidine decarboxylase activity through acid-induced inhibition of gastrin release. All agents and conditions which inhibit acid secretion but not gastrin release, cause an increase in the enzyme activity through enhanced gastrin release secondary to the elevation of antral pH.

Evidence for H2-receptor-mediated feed-back regulation of histamine release from endocrine cells in the rat stomach.

1. Pentagastrin reduces the concentration of gastric mucosal histamine in the rat. This effect is counteracted by large doses of histamine H2‐receptor agonists which are particularly effective if infused I.V. 2. After blockade of histamine H2‐receptors but not of H1‐receptors, the pentagastrin‐induced reduction of mucosal histamine is greatly augmented and H2‐receptor agonists no longer inhibit the response to pentagastrin. 3. It is suggested that in the rat stomach, H2‐receptors occur on the histamine storing endocrine cells as well as on parietal cells and that mobilized mucosal histamine inhibits further release of histamine by way of H2‐receptor‐mediated auto feed‐back.

Effect of pentagastrin and feeding on gastric histidine decarboxylase activity in the antrectomized rat

Abstract Feeding and pentagastrin injection evoked marked activation of gastric histidine decarboxylase in the fasted rat. After radical resection of the antrum together with the duodenal bulb, to achieve a total removal of endogenous gastrin stores, feeding failed to activate gastric histidine decarboxylase. Pentagastrin in maximal doses still activated the enzyme to the same extent in operated animals as in unoperated controls. Submaximal doses of pentagastrin were more effective in activating the enzyme in operated rats than in unoperated controls, indicating an increased sensitivity to gastrin after antrectomy. It may be concluded that the antrum proper as well as the adjacent portion of the oxyntic gland area together with the duodenal bulb has to be reserted in order to eliminate the endogenous gastrin stores, and that gastrin is the mediator of the enzyme activation induced by feeding.

Gastrin: Obligatory intermediate in the postprandial mobilization of gastric histamine in the rat

In unoperated fasted rats, feeding raised the serum gastrin concentration, reduced the gastric mucosal histamine content and activated the gastric histidine decarboxylase. The reduction of gastric histamine and activation of histidine decarboxylase was induced also by the injection of pentagastrin. In antrectomized rats, feeding failed to produce these effects. Injection of pentagastrin, however, still lowered gastric histamine and activated gastric histidine decarboxylase. Thus, antral gastrin seems to be an obligatory mediator of the postprandial activation of histidine decarboxylase and mobilization of histamine.