The Trophic Action of Gastrointestinal Hormones

Vol. 70, No.2 Printed in U.S.A.

70:278-288, 1976 Copyright © 1976 by The Williams & Wilkins Co.

GASTROENTEROLOGY

PROGRESS IN GASTROENTEROLOGY THE TROPHIC ACTION OF GASTROINTESTINAL HORMONES LEONARD

R.

JOHNSON, PH.D.

Department of Physiology, University of Texas Medical School, Houston, Texas

The discoveries of secretin in 1902 and gastrin in 1905 resulted in a new concept with important and far reaching physiological implications. Regulation of gastrointestinal function via the nervous system had been the subject of study in physiological laboratories for a number of years, and digestive processes were thought to be totally regulated by neural activity. Bayliss and Starling regarded secretin as one of a group of chemical messengers which would join the nervous system in coordinating and regulating the activities of the entire body. In 1905 Starling introduced the word hormone as a label for this group of substances. Recent definitions of the word, hormone, usually include the phrase, "-blood-borne regulator of metabolism and/or secretion." The obvious actions of the gastrointestinal hormones, the actions which led to their discoveries, and the actions for which they are named are their dramatic effects on secretion and motility of the digestive tract. The other hormones are primarily regarded as metabolic regulators, although some, such as thyroid-stimulating hormone and adrenocorticotrophic hormone, also regulate the secretion of other hormones. This dichotomy of actions resulted in a schism, which for the most part, has placed the gastrointestinal hormones in the area of physiology known as digestion and excluded them from that area known as endocrinology. I know of no endocrinology textbook which gives the gastrointestinal hormones more than a superficial treatment. A number of recent findings, however, indicate that this division may be completely spurious. Gastrointestinal hormones are now known to influence and cause the release of other hormones. In turn other hormones have been shown to affect the release of gastrointestinal hormones. It is now clearly established that gastrointestinal hormones regulate the metabolism and growth of a number of digestive tract tissues. 1 These effects on growth appear to be one of the few "physiological" actions of this group of peptides. The purpose of this article is 2-fold: first, to review the evidence leading to the conclusion that gastrointestinal hormones are trophic hormones, and second, to try to place this trophic effect in perspective with the physiology of digestive control and with endocrinology in general. Received May 27, 1975. Accepted July 10, 1975. This work was supported by National Institutes of Health Grant Am 16505 and Research Career Development Award Am 28972.

Trophic Responses to Exogenous Gastrin The first presumptive evidence that gastrointestinal hormones influence growth is found in a number of studies describing the long-term effects of antrectomy on the remaining oxyntic gland mucosa. Lees and Grandjean 2 biopsied the gastric mucosal remnant in 33 healthy post-antrectomy patients. One of these was considered normal and 22 exhibited moderate to complete atrophy. In another study 56 patients, 81.5% of whom had normal preoperative gastric mucosal biopsies, underwent partial gastrectomy for duodenal ulcer.3 Twelve months later 70.4% had varying degrees of atrophic gastritis, and the thickness of the parietal cell layer had decreased from a mean of 0.71 mm preoperatively to 0.51 mm. These results cannot be explained on the basis of disuse hypotrophy, for vagotomy and antrectomy both decrease acid by about 60% yet mucosal atrophy does not occur after vagotomy. 4 The decrease in acid secretion after antrectomy in man can be partially prevented by infusing pentagastrin continuously during the 1st week after antrectomy. 5 This finding indicates that exogenous gastrin prevents mucosal atrophy in man following antrectomy. The opposite picture, mucosal hyperplasia, occurs in patients having hypergastrinemia due to the so-called Zollinger-Ellison syndrome. 6 Gastric mucosal hyperplasia with an increased parietal cell count are characteristic of this disease. 7 Clinically, therefore, the overproduction of gastrin is associated with gastrointestinal mucosal growth and the lack of the hormone with mucosal atrophy. These observations could be explained if gastrin were a trophic hormone and led us to examine the effect of pentagastrin on protein synthesis. B Rats were divided into three groups and injected with either saline, pentagastrin, or histamine . Doses of pentagastrin ranged from threshold to supermaximal for gastric acid secretion. The animals were killed 90 min after a single injection and homogenates of various tissues incubated with [l4C ]leucine. A dose of pentagastrin, sub maximal for acid secretion, caused a 60 to 100% stimulation of protein synthesis in oxyntic glandular mucosa and a 300% stimulation in duodenal mucosa. B There was no stimulation of protein synthesis in either liver or skeletal muscle. Histamine had no effect on protein synthesis in any of the tissues examined. The stimulation of leucine incorporation was related to the dose of pentagastrin in a

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typical sigmoid manner. From this study we concluded that: (1) gastrin stimulates protein synthesis, (2) this effect is specific to certain tissues of the digestive tract, and (3) the effect is independent of secretory phenomena. In addition we hypothesized that gastrin was a trophic hormone and regulated the growth of gastrointestinal tract mucosa. 8 The above conclusions have been supported by numerous studies and gastrin has been shown to stimulate most of the metabolic responses associated with growth. Spectrum of growth-related responses affected by gastrin. Chronic in vivo administration of pharmacological amounts of pentagastrin produces oxyntic gland hyperplasia,9 and enlargement of the exocrine pancreas 10 and duodenum. 10 Crean et al. 11 found that duodenal obstruction in the rat led to a marked hyperplasia of the gastric mucosa. Surmising that this effect could be due to increased stimulation from large amounts of circulating gastrin released by antral distention, they injected rats with 4 mg of pentagastrin per day for 21 days. They found large increases in the weights of the whole stomach and the oxyntic gland area. 9 The antrum was not affected. In addition mucosal height and volume were increased and the parietal cell mass had undergone hyperplasia. The peptic cell population increased about 20%, but this was not statistically significant. Although histamine injections did not result in parietal cell hyperplasia, they attributed the effects of pentagastrin to either increased acid secretion or a direct trophic effect of the hormone. Neuburger et al. 12 examined the resected stomachs from 4 patients with Zollinger-Ellison syndrome and found hyperplasia of both the parietal and chief cell populations. In addition they commented on thetconsiderable number of mucin-secreting cells found in the specimens. The rest of their findings, increased fundic mucosa height and volume, also correlated well with the changes found in animals 9 after chronic gastrin administration. The administration of large doses of pentagastrin (2 mg per 100 g of body wt per day) over a 2-week period resulted in pancreatic acinar cell hypertrophy. 10 Hypertrophy was accompanied by a decrease in the specific activities of pancreatic enzymes, which was a result of increased protein and RNA content of the tissue. There was no increase in pancreatic DNA; thus, the RNA:DNA ratio increased significantly. Chronic histamine administration had no similar effects on the pancreas. Mayston and Barrowman 10 concluded that gastrin was a trophic hormone for the pancreas and that this tissue should be added to the others shown to be under its influence. Actually the results of this study10 do not suggest a growth-stimulating effect on the pancreas, for DNA did not increase and acinar cell hyperplasia did not occur, only hypertrophy. In a more recent study, however, the same authors have demonstrated both pancreatic hypertrophy and hyperplasia in hypophysectomized animals treated with pentagastrin. 13 The deleterious results of hypophysectomy were prevented not only in the pancreas but also the stomach and duodenum. Pentagastrin did not stimulate growth of the liver, kid-

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neys, or adrenal glands of the hypophysectomized rats. 13 Why chronic pentagastrin did not stimulate pancreatic hyperplasia in intact animals 10 is not known. Trophic hormones regulate a number of growthrelated processes in their target tissues. These include stimulation of protein synthesis, RNA synthesis, DNA synthesis, increased amino acid uptake, and decreased protein catabolism. Collectively this reaction to a growth hormone is termed the pleiotypic response. Stimulation of RNA, protein and DNA synthesis have all been shown to occur in response to pentagastrin. 1 Three injections of 250 J.lg per kg of pentagastrin resulted in a significant stimulation of RNA synthesis in both duodenal and gastric mucosa. u RNA synthesis was measured by following the incorporation of [UC ]orotic acid into RNA. Gastrin did not stimulate RNA synthesis in the liver. Pentagastrin prevents the decrease in RNA and DNA content of fundic and duodenal mucosa in antrectomized animals.14 Chronic administration of pentagastrin increases the total amount of pancreatic RNA in both normal 10 and hypophysectomized rats. 13 This effect appears to be restricted to the acinar cells. Duodenal and gastric RNA synthesis appears to peak 2 to 3 hr after a single injection of pentagastrin. 15 As mentioned earlier, pentagastrin stimulates the in vitro incorporation of leucine into protein of gastric and duodenal mucosa. 8 Synthetic human gastrin caused a significant stimulation of in vivo protein synthesis in both duodenal and gastric mucosa 16 Since the physiologicallyactive circulating forms of gastrin are primarily the 17 amino acid form (referred to as G 17 or "little" gastrin) and the 34 amino acid form (G 34 or "big" gastrin), it will be interesting and necessary to compare the trophic effects of these compounds with each other and with those of pentagastrin. The study16 noted above, utilizing synthetic human G 17, is the only one, to my knowledge, involving a pure circulating form of gastrin. Protein synthesized under the stimulation of pentagastrin appears to be confined to the gastric mucosal cells rather than secreted. Enochs and Johnson 17 injected rats with pentagastrin or saline and incubated pieces of oxyntic gland mucosa in tissue culture medium containing labeled amino acid. Over a period of time labeled protein appeared in both mucosa itself and the medium. However, the amounts appearing in the medium were the same for both groups of rats whereas there was considerable stimulation of the synthesis of protein in the tissue. 1, 17 Sutton and Donaldson 18 supported this finding by demonstrating that the in vitro addition of pentagastrin to isolated gastric mucosa maintained by organ culture increased the over-all incorporation of [UC ]leucine into gastric mucosal protein. Acetylcholine, cholecystokinin, secretin, and pentagastrin all stimulated the secretion of macromolecular protein, but only pentagastrin increased synthesis of tissue protein. 18 Sutton and Donaldson interpreted their findings as confirming the trophic effect of pentagastrin. Tissue cultures of gastric mucosal epithelial cells exposed to pentagastrin accumulated protein much more rapidly than their counterparts exposed to saline or histamine. 19 Five days after innoculating culture flasks

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with equal amounts of identical cells, those treated with pentagastrin contained twice as much protein as the saline or histamine controls. In order to state that a particular agent has a trophic effect one must demonstrate an increased number of cells after exposure to the agent. Increased DNA synthesis or the total amount of DNA are biochemical indications of comparable significance. Whereas protein and RNA synthesis are stimulated by trophic hormones and total RNA and protein increase during growth, they also increase when cells enlarge or hypertrophy. Increased DNA and its synthesis, however, are indicative of cell division and hyperplasia. Willems et al. 20 found stimulation of the uptake of [3HJthymidine into canine gastric mucosa after a 4-hr infusion of porcine gastrin. Thymidine uptake was significantly stimulated at 12 hr after gastrin infusion and peaked at 16 hr. Also using autoradiography we have demonstrated that pentagastrin stimulates the uptake of [3HJthymidine into nuclei of cultured duodenal cells. 21 In neither of the above studies was it possible to distinguish between bound thymidine and thymidine actually incorporated into DNA. There is little doubt, however, that incorporation occurred because thymidine uptake was followed by increased cell division in both studies. 20 . 21 Assessment of DNA synthesis by measuring the incorporation of [3HJthymidine into DNA isolated from gastrointestinal mucosa has proven that gastrin stimulates the formation of DNA and has provided a rapid and convenient method of investigating the trophic effects of the gastrointestinal hormones. 22 , 23 In a recent study rats were injected with 250 J.l.g per kg of pentagastrin or an equivalent volume of saline and killed at various time after injection. 24 Small pieces from the oxyntic gland area, duodenum, ileum, and liver were removed and incubated for 30 min in tissue culture medium containing [3HJthymidine. The DNA was extracted and the incorporation of thymidine determined. Pentagastrin had no effect on DNA synthesis in the liver. In all other tissues maximal stimulation occurred 16 hr after injection of hormone. In this particular study incorporation of thymidine in pentagastrin-injected animals was 275% of control for the stomach, 300% for the duodenum, and 480% for the ileum. In another series of studies histamine (20 mg per kg) had no effect on DNA synthesis. If pentagastrin is administered over a 48-hr period as six equally spaced injections, one is able to detect significant increases in total DNA and RNA as well as in DNA synthesis.23 These effects of gastrin on parameters related to growth are essentially identical to those which have been described for growth hormone, androgens, and estrogens.24, 25, 26 Tissue and species specificity of the trophic effect of gastrin. The trophic effects of gastrin on the oxyntic gland area, duodenum, and pancreas are documented in the previous section and in two review articles. 1, 15 In addition a single injection of pentagastrin resulted in a several-fold increase in ileal DNA synthesis. 21 Six injections of pentagastrin over a 48-hr period significantly stimulated DNA synthesis and increased the total

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amounts of DNA and RNA present in the large intestine (Johnson, unpublished results). While some of these studies have not been completed, it appears that gastrin may be a trophic hormone for the entire small and large bowels. The most notable exceptions to the trophic effects of gastrin on gastrointestinal tissue are the esophagus and antrum. Six injections of pentagastrin over a 48-hr period failed to increase DNA synthesis or the total amounts of DNA and RNA in the rat esophagus (Johnson, unpublished results). The area of the esophagus used in these studies did not include the lower esophageal sphincter. If gastrin has trophic effects on the sphincter, it would resolve some of the controversy over the physiological role of the hormone in this tissue.27, 28, 29 Gastrin does not increase the in vivo incorporation of [14C Jleucine into antral mucosa,16 nor does it stimulate DNA synthesis in this tissue. 15 Chronic administration of pentagastrin significantly increased the weight of the fundus of hypophysectomized rats without having any effect on antral weight. 13 In a study of parietal and chief cell populations in four cases of Zollinger-Ellison syndrome Neuburger et al. 12 found a diminution in antral size in spite of hyperplasia of parietal and chief cells. They suggested, in fact, that fundic mucosa was enlarged at the expense of the antrum. It is not surprising that gastrin stimulates growth of the two tissues, oxyntic gland and duodenum, on either side of the antrum without affecting the antrum itself. Regulation of antral growth by gastrin would be in opposition to the general concepts of endocrine physiology, for this tissue is the origin of most physiologically released gasJ.rin. Thyroxine, cortisol, androgens, and estrogens regulate metabolism and growth in a number of tissues, but not in their glands or origin. The growth of the thyroid, adrenals, and sex glands is regulated by pituitary peptide hormones. Endocrine cells of the antrum proliferate during periods of chronic stimulation for the release of gastrin. Antrocolic transposition resulted in significant increases in enterochromaffin cells 30 , 31 and gastrin cells. 31 Lichtenberger et al. 32 have demonstrated that fasting reduces both antral and serum gastrin content. Both were increased by feeding. In general stimulation of gastrin release results in higher levels of antral gastrin. Trophic effects of gastrin are apparently restricted to the mucosa, at least in the stomach and duodenum. 16 Gastrin did not increase [14C Jleucine incorporation into muscle of the oxyntic gland area. 16 Nor did it stimulate DNA synthesis in the smooth muscle layers of the stomach or duodenum even though DNA synthesis in the mucosa scraped from the same tissues was doubled by pentagastrin. 15 There have been no trophic effects described for gastrin in any tissue outside the gastrointestinal tract. Those which have been examined include liver,8, 10, 22. 33 skeletal muscle,8 kidneys, 10, 34 spleen, 10, 34 and testes. 34 As in the case of most fields of study involving acute biochemical experiments, the standard animal employed for the investigation of the trophic effects of

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gastrin has been the rat. Almost all of the experiments and subsequent subcultures. Control cultures showed described in the preceeding discussion have utilized this rapid growth of fibroblasts and ultimate loss of epithelial animal. A noteworthy exception is the demonstration cells . Fibroblastic outgrowth was inhibited and epithethat gastrin stimulates thymidine uptake and increases lial cell proliferation stimulated in the cultures receiving the mitotic index in dog. 20 Evidence of the trophic action pentagastrin. At confluency the mitotic activity of the of gastrin in man has been indirect, stemming in a large pentagastrin-treated subcultures was more than twice part from observations of mucosal hyperplasia in pa- that of the controls. Cell lines first exposed to pentagastients with hypergastrinemia due to Zollinger-Ellison trin and then to saline in the subsequent cultures syndrome. 7 • 12 These studies plus mucosal atrophy in developed a control type growth pattern and low mitotic patients having had subtotal gastric resection furnish index which was unaltered by gastrin. Gastrin-treated excellent presumptive evidence that growth of the subcultures contained twice the protein present in sahuman gastrointestinal tract is affected by gastrin in the line-treated control subcultures which had been started same manner as the rat's. At this point, it would be with an identical inoculum. Individual cell types were worthwhile to investigate directly the trophic activity of not identified, but electron microscopic examination gastrin on the human gut. Perhaps the best means of showed a primitive epithelial populace in the pentagasdoing this would be to study in vitro DNA synthesis in trin-treated cultures presenting as a composite of the mucosal biopsy specimens from patients with altered anatomical features seen in the differentiated gastric endogenous gastrin levels (e.g., fasting versus fed) and mucosal cell flora . Numerous junctional complexes refrom volunteers treated with exogenous gastrin . sembling desmosomes and typical of epithelial cells were found between cells in the gastrin-treated flasks, but Mechanism of the Trophic Action of Gastrin never in the saline controls. 19 The ability of gastrin to There is a number of basic questions which one can stimulate growth of gastric mucosa in vitro is supported ask about the mechanism of action of any hormone. by the finding that pentagastrin but not other seFirst, does the hormone act directly, or does it trigger the cretagogues stimulates the synthesis of cell protein in release or production of another hormone which in turn organ cultures from the oxyntic gland area . 18 Lichtenberger et al. 21 examined the effects of penproduces the effect under question? Second, are the observed metabolic effects the result of another action of tagastrin on duodenal cells growing in tissue culture. the hormone, such as, in the case of gastrin, the After establishing cultures of adult rat duodenal cells, stimulation of secretion or motility? Third, what is the the contents of half the flasks were exposed to pentagasbiochemical mechanism primarily triggered by the hor- trin once daily and half to saline. After 3 months the pentagastrin-treated cultures contained approximately mone? Trophic effects of gastrin in vitro. The best evidence 90 % epithelial cells and 10 % fibroblasts , whereas the that a hormone is acting directly on its target cells is to control cultures constituted an epithelial cell-fibroblast be able to demonstrate its effects by exposing thsue to admixture of about 50% each. Epithelial cells appeared the hormone in vitro. Such a demonstration, of course, quite similar in structure to crypt cells, although a does not eliminate the possibility that a second messen- positive identification was not made. However, because ger, such as one of the cyclic nucleotides, is involved in gastrin-stimulated cultures of gastric mucosal cells also stimulating processes within the cell . It does, however, contained poorly differentiated cells, this evidence sugmean that the hormone is not causing the synthesis gests that the trophic effect of gastrin is on generative and/or release of a second agent which is transported by cells in the mitotic focus and not on secretory cells. the blood to react with receptors in the target cells . Pentagastrin-treated cultures had a faster doubling Growth hormone and somatomedin offer the best exam- time, 19.5 hr, compared with 31.5 hr for saline treated ple of this second type process. 35. 36 Stimulation of controls. This was attributed in part to the greater sulfate uptake and protein synthesis in cartilage are percentage of cells in the proliferative population in the widely recognized effects of growth hormone . These hormone treated cultures, 73 %, in comparison to the effects cannot be produced by adding growth hormone controls, 36%. This latter measurement was arrived at directly to cartilage in vitro. Plasma from normal rats by autoradiographic determination of [3HJthymidine added to cartilage from hypophysectomized rats , how- incorporation into DNA. The most important conclusion from these studies ever, stimulates sulfate and amino acid incorporation. 35 Recently this "sulfation factor" contained in normal utilizing cell and organ culture techniques is that the plasma has been identified as a peptide whose synthesis trophic effects of gastrin shown in vivo can be accounted in the liver is dependent on growth hormone . This for by direct action of the hormone and are not due to the substance has been named somatomedin and is responsi- production and/or release of another hormone or factor. Independence of the trophic and secretory actions of ble for many, and perhaps most, of the effects of growth gastrin. The in vitro studies mentioned in the previous hormone. a6 To date there are three studies which demonstrate section also provide some evidence on this point. Bethat pentagastrin can stimulate trophic activity in vitro. cause the systems used were completely buffered, it is Miller et al. 19 used pentagastrin to maintain tissue obvious that exposure of cells to an acid environment cultures of rat and human oxyntic gland mucosa. Either cannot account for the trophic effects of gastrin. In pentagastrin, to a concentration of 0.5 Ilg per ml, or an addition, it is unlikely that acid secretion was stimuequal volume of saline was added daily to the cultures lated in the cell cultures. Although this cannot be

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proven, the trophic effect of gastrin occurred in the probable absence of the secretory effect. Many experiments involving the trophic action of gastrin have included a group of histamine-injected animals to control for acid secretion. The results of these studies have been unanimous in that metabolic actions of gastrin were not duplicated by histamine. These experiments have demonstrated gastrin stimulation of parietal cell hyperplasia,9 pancreatic hyperplasia,lO gastrin and duodenal protein synthesis,8 gastric and duodenal RNA synthesis,33 DNA synthesis and growth of cultured duodenal cells,21 and gastric, duodenal, and ileal DNA synthesis. 22, 23 The studies listed above were done on rats, and histamine is an extremely poor secretagogue when compared with gastrin in the rat. 37 Therefore, it can be argued that these experiments are not really effective controls for the effects of acid secretion. For this reason the study by Willems et al. 20 using the dog is especially significant. They infused either gastrin or histamine over a period of 4 hr in doses producing nearly identical acid outputs. Thymidine uptake into gastric mucosal cells and cell division were significantly stimulated in the animals receiving gastrin. There was no stimulation in any histamine-infused dog. Further evidence that the trophic actions of gastrin are independent of secretory effects is provided by a recent study in which DNA synthesis in response to pentagastrin was studied in the presence of inhibitors of gastric acid secretion. 23 Metiamide, a histamine H 2-receptor antagonist which also inhibits gastrin stimulated acid secretion, was administered in combination with pentagastrin in a dose shown to block acid secretion almost completely. In this experiment pentagastrin alone caused a 40% stimulation of DNA synthesis in the oxyntic gland mucosa and an 80% increase in duodenal mucosa. These values were not significantly altered when metiamide was administered with pentagastrin. Biochemical mechanism of the trophic action of gastrin. The increases in cell division and the peak stimulation of DNA synthesis in target tissues in response to growth hormone, thyroid-stimulating hormone, adrenocorticotrophic hormone, and other growth-promoting hormones are preceded by increases in both RNA and protein synthesis. While the initiating event in response to gastrin has not been elucidated, considerable effort has been expended along lines with regard to other trophic substances. Korner 38 has shown that hypophysectomized animals have a deficiency in the formation of polyribosomes which can be corrected by the addition of polyuridine, indicating a decrease formation of messenger RNA. The addition of growth hormone resulted in an initial stimulation of messenger RNA followed by increases in all types of RNA and the stimulation of protein synthesis. 39 Subsequent experiments indicated that growth hormone facilitated the formation of a DNA-RNA polymerase complex and hence messenger RNA synthesis!O Data involving the time course of synthesis of various macromolecules in response to gastrin have been consist-

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ent and present a picture which indicates that stimulation of RNA production may be the initial event.' Thymidine uptake into canine gastric mucosa peaks 16 hr after the infusion of gastrin and is followed at 20 and 24 hr by a 5-fold increase in the mitotic index. 20 In the rat ileal, duodenal, and oxyntic gland area DNA synthesis is maximally elevated 16 hr after a single injection of pentagastrin, although it is significantly elevated as early as 8 hr in the duodenum. 22 The final degree of stimulation appears to depend on the epithelial cell turnover time of the tissue involved. Protein synthesis in the rat gastric and duodenal mucosa increases approximately 4 hr after pentagastrin injection and peaks 2 or 3 hr later.17 Protein synthesis in organ cultures of gastric mucosa was stimulated 7 hr after exposure to pentagastrin.18 RNA synthesis increases more rapidly and attains maximal rates 2 or 3 hr after pentagastrin injection. 15 Stimulation of messenger RNA occurs early, at 1 hr, and eventually the snythesis of all three species of RNA is stimulated (Enochs and Johnson, unpublished results). Whether the increases in protein and DNA synthesis and eventually cell division are dependent on the transcription of RNA has not been completely investigated. Nevertheless, the picture of the time course of biochemical events after gastrin administration is remarkably consistent with those shown for other trophic hormones. Whether gastrin increases protein and RNA synthesis in cells incapable of DNA synthesis and division is not known.

Trophic Responses to Endogenous Gastrin: Physiological Significance Although there is some disagreement on the criteria which must be satisfied before a particular response to a gastrointestinal hormone can be considered physiological, one criterion which must certainly be met is that the effect should occur in response to levels of the hormone normally occurring in the blood. The concept of normal levels needs further clarification. First, normal level may mean the increment in the serum hormone concentration due to the physiological stimuli for its release; in the case of gastrin the stimuli would be those arising from the ingestion and presence of a meal within the gastrointestinal tract. Second, the effective level may depend on the normal continuing presence of the hormone. A constant background presence of hormone might be important in the case of a permissive effect or an effect which requires repeated or continuing exposure to the hormone. It goes without saying that the effect should occur whether the serum levels are increased by exogenous or endogenous hormone. There is general concensus that the trophic action of gastrin is physiological,41 and in fact, it may be the most important physiological effect of the hormone. Antrectomy. Atrophic gastritis occurs in man after partial gastrectomy and gastroenteroanastomosis. 3, 42, 43 General gastric mucosal atrophy has also been produced by antrectomy in rats,44 Some investigators have suggested that gastric mucosal atrophy after antrectomy is due to reflux of duodenal contents into the stomach. 45 , 46

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However, Sander et al. 47 found no histological abnormalities in rat gastric mucosa after 90 days of exposure to duodenal contents. Martin et al. H hypothesized that removal of endogenous gastrin was responsible for the atrophy of oxyntic gland mucosa after antrectomy in the rat. The classical method for proving that a tissue is dependent on a particular hormone for its growth and integrity is to remove the source of the hormone and observe atrophy or cessation of growth in the target tissues, and then to supply the hormone exogenously and reverse these changes brought on by extirpation of the endocrine cells. In the rat, antrectomy resulted in approximately 40% decreases in gastric and duodenal mucosal RNA and DNA during the month after surgery. U A series of pentagastrin injections given to half the antrectomized animals almost completely prevented the decreases in nucleic acid content. 14 Thus, negative effects on growth of gastrointestinal tissues are caused by removing endogenous gastrin and reversed by supplying the hormone exogenously. This study is supported by one mentioned earlier demonstrating that the decrease in human acid secretory capacity after antrectomy fails to develop if the patients are given pentagastrin after surgery.5 These experiments are strong evidence that endogenous gastrin has a physiological function as a trophic hormone.

Association between serum and antral gastrin levels and growth of gastrointestinal mucosa. Perhaps the best evidence that the trophic actions of gastrin are physiologically significant comes from examining growth of the gastrointestinal tract after natural alteration of serum and antral gastrin levels. There is a number of times when gastrin levels change dramatically due to deVelopmental and physiological events. It is possible to exaggerate and prolong these changes without resorting to surgery. The rat small intestine undergoes major developmental changes during the 3rd week of postnatal life. These include increases in relative intestinal wet weight, villus and crypt height, mitotic activity, villus migration rate, and nucleic content. 48, 49 Between days 14 and 21 intestinal enzymatic activity also changes to approach adult patterns with decreases in lactase and a kaline phosphatase 50 and increases in maltase, sucrase, and trehalase. 51 Using a relatively insensitive bioassay, Zelenkova and Gregor 52 first found measurable levels of an antral gastrin during the 3rd week of life. With radioimmunoassay, antral levels of gastrin were 1 to 3 JLg per g during days 1 to 18 and increased to adult levels of 15 to 20 JLg per g on day 21. 53 In order to test whether the appearance of adult levels of gastrin might be responsible for some of the coincident changes in gut structure and function, one group of 14-day-old rats was prevented from weaning while a second group was allowed to wean. 53 Ten days later the rats were killed, and the ratios of gut wet weight to body weight, RNA to body weight, and protein to body weight were significantly lower in the non weaned rats. The antral gastrin concentration also remained at low levels in the non-

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weaned animals. In the next series of animals pentagastrin was injected into 14-day-old rats prevented from weaning. Ten days of pentagastrin treatment significantly increased the gut weight to body weight, RNA to body weight, and proteih to body weight ratios over those found in saline-injected control subjects. These increases were comparable to those seen after normal weaning. Lichtenberger and Johnson 53 concluded that some aspects of gut development are dependent on weaning, and that the mediator between these two events may be gastrin. Starvation is known to cause dramatic changes in small intestinal structure and function. Villus and crypt height in the rat are diminished after 3 to 6 days of starvation, 54, 55 and mucosal RNA, DNA, and protein are markedly decreased after 3 to 6 days starvation. 32 , 56 These decreases are significantly greater than the loss of body weight. During 3 days of starvation rat antral gastrin levels dropped from 32 JLg per g of wet weight to 5 JLg per g of wet weight, and serum gastrin concentration decreased from 330 pg per ml to 70 pg per ml. 32 In these same animals intestinal DNA, RNA, and protein content decreased significantly compared with body weight. Specific lactase and maltase activity increased. Animals injected with pentagastrin during the period of starvation showed significantly smaller changes than the starved saline-injected control rats. 32 Ingestion of a meal is followed by increases in DNA synthesis and in the mitotic index of canine fundic mucosa. 57 This pattern can be reproduced by a 4-hr infusion of gastrin,20 suggesting that gastrin might be one of the factors responsbile for postprandial cell renewal. Although the studies mentioned above implicate decreased gastrin levels as a cause of the profound deleterious effects which short periods of starvation have on the gut, interpretation of these studies is made difficult by the inability to separate the metabolic changes associated with starvation from those caused purely by the absence of food from the gut. In a recent series of studies our laboratory has used the intravenously alimented rat as a model to study gut structure and function in the well nourished animal whose gastrointestinal tract has gone unexposed to food and the stimuli arising from its ingestion and presence. Several findings from these studies are of special significance. First, the parenterally fed animals often gained weight and always remained in positive nitrogen balance. 58, 59 Second, tissue to body weight ratios for oxyntic gland area, small intestine, and pancreas were significantly decreased in the parenterally fed animals, whereas the weights of other organs were unaffected. 34 , 59, 60 Third, specific and total activities of the different disaccharidase enzymes were only a fraction of those found in the orally fed controls. 58, 61 Fourth, the parenterally fed animals were nearly depleted of antral gastrin. 34 , 60 Fifth, these results could not be completely explained on the basis of food intake, dietary constitutents, enzyme inducion, or the absence of luminally derived nutrition in the parenterally fed animals.

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In the latest of these studies one group of parenterally nourished animals received a continuous infusion of 6.0 J.Lg per kg per hr pentagastrin, a dose considerably less than the D50 for acid secretion in this species. 34 The animals were killed approximately 2 weeks later and compared with parenterally fed rats which had received either histamine or nothing in addition to the intravenous diet. Serum as well as antral gastrin concentrations decreased significantly in all groups of parenterally fed animals. Weights of the oxyntic gland area, small intestine, and pancreas decreased significantly in all parenterally fed rats except those receiving gastrin. Gastrin completely prevented the decrease in disaccharidase activity normally associated with total parenteral nutrition. These data were interpreted to indicate that the oral ingestion of food and its presence in the gastrointestinal tract are necessary to maintain endogenous gastrin levels, and that the trophic action of endogenous gastrin is essential for the day to day maintenance of the structural and functional integrity of the gut. 3' Viewed as one body of evidence, the parallelism between serum and antral gastrin levels and growth of gastrointestinal tract tissue is striking. In each instance decreased endogenous gastrin levels were associated with decreased growth, and the addition of exogenous gastrin was able significantly to increase growth towards normal levels. In the study involving parenterally fed animals the dose of gastrin infused was well below the limit considered to be physiological. Since the effects of exogenous gastrin in this study were dramatic,34 it may indicate that the continuing presence of a low level of the hormone is more effective in stimulating growth than periodic extreme fluctuations in hormone concentration . In summary, the evidence in favor of the trophic action of gastrin being a physiological action is strong, perhaps as strong as for any other action of a gastrointestinal hormone.

Cholecystokinin (CCK) CCK is structurally and functionally related to gastrin. The active C-terminal tetrapeptide amide of gastrin is duplicated in CCK. The major structural difference which dictates whether a peptide of the CCK-gastrin family has a gastrin-like or CCK-like pattern of activity is the position of the tyrosyl residue and whether or not it is sulfated. Qualitatively the actions of gastrin and CCK are identical, but gastrin has high affinity for receptors stimulating acid secretion and low affinity for those involved in gallbladder contraction and pancreatic enzyme secretion. The opposite pattern prevails for peptides more closely related to CCK. Owing to the overlapping activity patterns and the tendency for CCK to have a higher affinity for tissues located more distally in the digestive tract, it seemed likely that cck would have trophic influences on the pancreas and small intestine. Rothman and Wells 62 found that CCK stimulated the synthesis of pancreatic enzymes in the rat and caused a significant increase in pancreatic weight. Secretin was without effect. The authors concluded that CCK aug-

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mented the synthesis of exportable protein. The study was not designed to examine whether or not CCK was a' trophic hormone for the pancreas. Mainz et al. 63 found evidence that CCK regulates growth of the pancreas as well as secretion. Chronic administration of CCK was associated with increased RNA, protein, and DNA content and with the stimulation of [14C ]thymidine incorporation into DNA. Chronic administration of bethanechol chloride, an analogue of acetylcholine, led to increased RNA and protein content but did not increase DNA content or stimulate DNA synthesis. Thus, although both CCK and acetylcholine stimulated pancreatic protein secretion and synthesis, only CCK appeared to be capable of stimulating growth Jf the pancrease. Barrowman and Mayston 64 conducted a study similar to the one mentioned above by injecting rats with 12.5 units of CCK per 100 g of body weight per day for 9 days . They found a slight but significant increase in total pancreatic DNA and interpreted their results as indicating that CCK is a trophic hormone for the pancreas. We have recently examined CCK for trophic effects on the oxyntic gland area of the stomach and the duodenal mucosa as well as the pancreas. 65 Low doses of CCK octapeptide (CCK-OP) causing a small but significant increase in pancreatic DNA synthesis had no stimulatory effect on mucosa of the oxyntic gland area or duodenum of the same animals. DNA content of the pancreas was also increased, indicating that the increase in synthesis was not matched by an increase in turnover. At higher doses of CCK-OP there was a slight increase in duodenal DNA synthesis and content which was of borderline statistical significance. Further increasing the dose of CCK-OP inhibited the trophic effect of pentagastrin in both the stomach and duodenum. 65 We concluded that although CCK is probably a physiologically important regulator of growth of the exocrine pancreas, it is unlikely that it exerts a trophic influence on either the stomach or duodenum.

Secretin Although there have been relatively few studies involving the trophic or possible trophic action of CCK , to my knowledge there are only two reports on secretin. In one study rats having gastric cannulas were injected three times daily for 2 weeks with pentagastrin, secretin, pentagastrin plus secretin, or saline. 66 Basal and maximal acid outputs were measured before, during and after the injection period. Parietal cell mass was determined at the end of the study. Pentagastrin injection led to a 90% increase in maximal acid output. This increase failed to occur in the rats receiving secretin in addition to pentagastrin. The parietal cell population increased by 70% in the gastrin-injected rats. This too was prevented by secretin. The animals receiving only secretin had slightly lower secretory capacities and parietal cell counts than the saline-injected controls.66 The effects of secretin on gastric acid secretion are well known in a variety of species. In the rat the dose of secretin used in the study outlined above causes nearly complete inhibition of acid secretion stimulated by a

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maximal dose of pentagastrin. 67 Therefore, even though the trophic action of gastrin is independent of acid secretion, Stanley et al. 68 were unable to conclude whether the decreased secretory capacity and parietal cell mass was due to an antitrophic effect of secretin or to the inhibition of gastric acid secretion. Secretin, however, has recently been shown to inhibit the gastrin stimulation of DNA synthesis and accumulation in both the oxyntic gland region of the stomach and the duodenum. 23 This observation provides a biochemical basis for the effects of chronically administered secretin on secretory capacity and parietal cell mass. In the same study metiamide, a potent inhibitor of gastrinstimulated acid secretion, had no significant effect on the trophic response to pentagasrin.23 The remaining question is whether secretin has antitrophic activity of its own or whether it acts solely by inhibiting the growth-promoting effects of gastrin. Chronic administration of secretin by itself caused a decrease in parietal cells and acid secretion when compared with saline-injected control subjects. 68 Considering the profound inhibition of gastrin trophic activity caused by secretin, it is entirely possible that these antitrophic effects were due to the inhibition of endogenous gastrin. Because it is almost impossible to remove all sources of gastrin surgically, this point will best be settled by studying the metabolic effects of secretin on in vitro systems involving cell or organ culture.

Significance of the Trophic Action of Gastrointestinal Hormones In a previous section of this article I concluded that the trophic action of gastrin is physiologically significant. This final section will focus on the over-all ~ignifi­ cance of the trophic actions of gastrointestinal hormones and try to place this action in perspective with endocrinology, digestive physiology, and certain clinical conditions. Although the gut is the largest endocrine organ in the body, gastrointestinal hormones have traditionally been considered in the realm of digestive rather than endocrine physiology. The fact that they have now been shown to stimulate growth of gastrointestinal tract tissues as well as their secretion and motility makes it imperative that gastrointestinal hormones be included in any serious treatment of the regulation of metabolism by endocrine substances. It is also becoming obvious that the gastrointestinal hormones physiologically influence and are influenced by the other endocrines. Glucagon inhibits the release of endogenous gastrin. All three gastrointestinal hormones stimulate insulin release, and gastricinhibitory peptide (GIP) may prove to be the "incretin" responsible for the rapid clearing of glucose seen during oral glucose tolerance tests. CCK has been shown to stimulate glucagon release, and both CCK and gastrin stimulate calcitonin release. 68 Clinically, hyperplasia of antral gastrin cells (G cells) and increased antral gastrin content have been demonstrated in patients with primary hyperparathyroidism and acromegaly. 69 Creutzfeldt et al. 69 interpreted these findings as indicating that serum calcium

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and growth hcrmone may have a trophic action on the G cells. However, a recent study by the same laboratory was unable to confirm their previous findings.70 The possibility that growth hormone has a trophic action on the antrum is especially intriguing, because the secretion and/or synthesis of other trophic hormones as well as the growth of the glands forming them is dependent on th~ pituitary. Chronic injections of pentagastrin have recently been shown to prevent atrophy of the pancreas, duodenum, and oxyntic gland region of stomach after hypophysectomy. 13 Gastrin treatment had no effects on the antrum, liver, kidney, or adrenal glands in hypophysectomized animals. The body weights of the saline- and pentagastrin-injected hypophysectomized rats were identical. The oxyntic gland and duodenal weights of gastrin-treated hypophysectomized rats were 100 mg heavier than their saline-treated counterparts and not significantly different from those of normal animals. This was especially noteworthy because the unoperated controls were not pair fed with the hypophysectomized group and were significantly larger. Mayston and Barrowman 13 found their results compatible with the hypothesis that at least some of the effects of the pituitary on the gastrointestinal tract may be mediated by influences upon the endocrine-secreting cells of the digestive tract. The above studies, indicating a relationship between growth hormone and gastrin, suggested that gastrin production and/or secretion could be regulated by growth hormone, and that the effects of growth hormone on the gastrointestinal tract are mediated by gastrin. In order to test this idea we examined serum and antral gastrin levels in hypophysectomized, hypophsectomized plus growth hormone, and sham-operated control rats.71 All groups of rats were pair fed to keep body weights nearly the same and to eliminate the dramatic effect of feeding on gastrin levels. Serum gastrin concentrations of 48-hr fasted hypophysectomized rats were 100 pg per ml compared with 220 pg per ml for controls. Fed, normal rats had serum gastrins of 300 pg per ml compared with 120 pg per ml for fed hypophysectomized rats. Treatment with growth hormone (500 Ilg per 100 g per day for 10 days) maintained serum gastrins at normal levels in both fed and 48-hr fasted hypophysectomized rats. The antral gastrin concentration from hypophysectomized rats was only one-fourth of control levels, and growth hormone increased this significantly to one-half of control levels. 71 Single injections of growth hormone had little or no effect on serum gastrin in hypophysectomized or normal rats. Based on these observations Enochs and Johnson 71 concluded that growth hormone is necessary for the normal maintenance of gastrin levels. These studies support the hypothesis that gastrin production may be regulated by growth hormone in the same manner in which cortisol and thyroxin are dependent on ACTH and TSH respectively. For the past 70 years, in spite of having no factual evidence, we have assumed that gastrointestinal hormones had no interactions with or functions similar to those of the other endocrines. At this time it would be much more produc-

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tive to treat and think of the gastrointestinal hormones as an integral part of endocrinology in general, at least until some evidence to the contrary is obtained. Numerous clinical implications of the trophic action of gastrointestinal hormones and gastrin in particular have already been mentioned. These include effects caused by altered gastrin levels after antrectomy or during Zollinger-Ellison syndrome, and perhaps acromegaly and hyperparathyroidism. Whether increased cell division stimulated by gastrin can be used to protect the gastric mucosa from ulceration is largely univestigated. In a similar vein the antitrophic action of secretin could be useful in duodenal ulcer therapy, because secretin would theoretically decrease the mass of acid -secreting cells as well as inhibit acid secretion and gastrin release. Gastrin and secretin are also potentially useful where the primary defect is the under- or overproduction of cells dependent of these hormones for growth. The effects of these trophic hormones on tumors of the gastrointestinal tract should be investigated. An interesting and apparent anomaly in this field involves patients with pernicious anemia who have abnormally high circulating levels of gastrin and atrophy of the oxyntic gland mucosa. There are causes of endocrine disease in addition to over- or underproduction of a hormone. Two of these are end organ receptor insensitivity and the production of inactive hormone. These could prove to be fruitful areas of investigation in pernicious anemia, especially because these patients have an increased risk of developing carcinoma of the stomach. In fact, although it has not been confirmed in a full publication, Hansky et al. 72 have already reported that the gastrin circulating in patients with pernicious anemia is biologically inactive. As I have already pointed out, the trophic action of gastrin may be important in gastrointestinal development and in cases of malnutrition. The severe effects on the gastrointestinal tract caused by total parenteral nutrition which are prevented by gastrin 34 should be examined carefully, because this is a widely used technique clinically. Although these clinical implications must be labeled speculative, the evidence to date warrants their investigation. If the trophic effects of gastrointestinal hormones on the digestive tract prove to be as important as those of the other hormones on the rest of the body, these studies would prove extremely worthwhile. Teleologically, it is revealing to speculate on the existence of gastrointestinal hormones . Gastrointestinal motility is largely under neural regulation. Some aspects of secretion are also regulated by extrinsic nerves and intramural reflexes. Absorption is largely unregulated. A recent report indicates that the canine Heidenhain pouch will secrete near maximal amounts of acid in response to liver extract placed directly in the pouch.73 Gastrin is not involved in this response. 73 The integrity of the cells of the gastrointestinal tract is, however, necessary for the production of digestive enzymes, intrinsic factor , and for the absorption of nutrients. This raises the consideration that the trophic action of gastrointestinal hormones may be their most important,

and although not the first to be discovered, primary action. Regardless of the final answers to the questions raised here, it is obvious that gastrointestinal hormones regulate much more than the muscular contractions of the gut and the flow of juices from the various faucets along the digestive tract. REFERENCES l. Johnson LR: Gut hormones on growth of gastrointestinal mucosa. In Endocrinology of the Gut. Edited by WY Chey and FP Brooks. Thorofare, N.J. Chas. B. Slack, Inc , 1974, p 163- 177 2. Lees F, Grandjean LC: The gastric and jejunal mucosae in healthy patients with partial gastrectomy. Arch Intern Med 101 :9437- 9451, 1968 3. Gjurldsen ST, Myren J, Fretheim B: Alterations of gastric mucosa following a graded partial gastrectomy. Scand J Gastroenterol 3:465- 470. 1968 4. Melrose AG , Russell RI . Dick A: Gastric mucosal structure a nd function after vagotomy. yut 5:546-549, 1964 5. Olbe L: Differences between human and animal gastric acid secretion. Syllabus for AGA Postgraduate Course on Peptic Ulcer Disease, San Francisco, 1974 6. Gregory RA, Grossman MI, Tracy HJ , et al : Nature of the gastric secretagogue in Zollinger-Ellison tumors. Lancet 2:543-544, 1967 7. Ellison EH, Wilson SO: Further observations on factors influencing the symptomatology manifest by patients with Zollinger-Ellison syndrome . In Gastric Secretion. Edited by TK Shnitka. JAL Gilbert, RC Harrison. New York, Pergamon, 1967. p 363-369 8. Johnson LR, Aures D. Yuen L: Pentagastrin induced stimulation of the in vitro incorporation of ["C )Ieucine into protein of the gastrointestinal tract. Am J Physiol 217:251-254. 1969 9. Crean GP, Marshall MW. Rumsey ROE: Parietal cell hyperplasia induced by the administration of pentagastrin (lCI 50, 123) to rats. Gastroenterology 57:147-156, 1969 10. Mayston PO, Barrowman JA: The influence of chronic administration of pentagastrin on the rat pancreas. Q J Exp Physiol 56: 113-122, 1971 1l. Crean GP , Rumsey ROE . Hogg DH: Hyperplasia of the gastric mucosa produced by duodenal obstruction. Gastroenterology 56: 193-199, 1969 12. Neuburger PH, Lewin M, de Recherche C, et al: Parietal and chief cell populations in four cases of the Zollinger-Elliso n syndrome. Gastroenterology 63:937-942, 1972 13. Mayston PO, Barrowman JA: Inf1uence of chronic administration of pentagastrin on the pancreas in hypophysectomized rats. Gastroenterology 64:391- 399, 1973 14. Johnson LR, Chandler AM : RNA and DNA of gastric and duodenal mucosa in antrectomized and gastrin-treated rats. Am J Physiol 224:937-940, 1973 15. Johnson LR: Trophic action of gastrointestinal hormones. In International Symposium on Gastrointestinal Hormones . Edited by JC Thompson. Austin, Texas, University Texas Press, (in press) , 1975 16. Johnson LR, Aures D , HAkanson R: Effect of gastrin on the in vivo incorporation of ["C)leucine into protein of the digestive tract. Proc Soc Exp Bioi Med 132:996-998, 1969 17. Enochs MR, Johnson LR: Pentagastrin stimulates tissue growth in stomach and duodenal tissues by stimulating protein and nucleic acid synthesis. Fed Proc 33:309. 1974 18. Sutton D, Donaldson RM: In vitro biosynthesis and secretion of pepsinogen by rabbit gastric mucosa. Gastroenterology 66:786. 1974 19. Miller LR, Jacobson ED, Johnson LR: Effect of pentagastrin on gastric mucosal cells grown in tissue culture. Gastroenterology 64:254-267, 1973

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20. Willems G, Van Steenkiste Y, Limbosch JM: Stimulating effect of 45. Lawson HH: Effect of duodenal contents on the gastric mucosa gastrin on cell proliferation kinetics in canine fundic mucosa. under experimental conditions. Lancet 1:469-472, 1964 Gastroenterology 62:385-389, 1972 46. Menguy R, Martin HM: Influence of bile on the canine gastricantral mucosa. Am J Surg 119: 177 -182, 1970 21. Lichtenberger L, Miller LR, Erwin DN, et al: The effect of pentagastrin on adult rat duodenal cells in culture. Gastroenterol- 47. Sander S, Myren J, Helsingen N: The effect of bile reflux on the ogy 65:242-251, 1973 gastric mucosa. An experimental study in rats. Gastroenterologia 22. Johnson LR, Guthrie PD: Mucosal DNA synthesis: A short term 101:3-12, 1964 index of the trophic action of gastrin. Gastroenterology 67:453-459, 48. Herbst JJ, Sunshine P: Postnatal development of the small 1974 intestine of the rat. Pediatr Res 3:251-254, 1969 23. Johnson LR, Guthrie PD: Secretin inhibition of gastrin-stimulated 49. Kodolvsky 0, Herbst JJ, Burke J, et al: RNA and DNA in intestinal mucosal during development of normal and cortisone deoxyribonucleic acid synthesis. Gastroenterology 67:601-606, treated rats. Growth 34:359-367, 1970 1974 24. Tomkins GM, Gelehrter TD: The present status of genetic 50. Moog F, Thomas ER: The influence of various gonadal steroids on regulation by hormones. In Biochemical Actions of Hormones, vol the accumulation of alkaline phosphatase in the duodenum of the suckling mouse. Endocrinology 56:187-196, 1955 II. Edited by G Litwack. New York, Academic Press, 1972, p 1-20 25. Williams-Ashman HG, Reddi AH: Androgenic regulation of tissue 51. Rubino A, Zimblatti F, Auricchio S: Intestinal disaccharidase activities in adult and suckling rats. Biochem Biophys Acta growth and function. In Biochemical Actions of Hormones, vol II. 92:305-311, 1971 Edited by G Litwack. New York, Academic Press, 1972, p 257-294 26. Jensen EV, DeSombre ER: Estrogens and progestins. In Biochemi- 52. Zelenkova J, Gregor 0: Development of gastrin activity. Scand J Gastroenterol 6:653-656, 1971 cal Actions of Hormones, vol II. Edited by G Litwack. New York, 53. Lichtenberger L, Johnson LR: Gastrin in the ontogenic developAcademic Press, 1972, p 215-256 ment of the small intestine. Am J Physiol 227:390-395, 1974 27. Castell DO, Harris LD: Hormonal control of gastroesophageal 54. McNeil LK, Hamilton JR: The effect of fasting on disaccharidase sphincter strength. N Engl J Med 282:866-889, 1970 activity in the rat small intestine. Pediatrics 47:65-72, 1971 28. Cohen S, Lipshutz W: Hormonal regulation of human lower esophageal sphincter competence-interaction of gastrin and 55. Altmann GG: Influence of starvation and refeeding on mucosal size secretin. J Clin Invest 50:449-454, 1971 and epithelial renewal in the rat small intestine. Am J Anat 29. Grossman MI: What is physiological? 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Johnson LR, Castro GA, Lichtenberger LM, et al: The significance rat. Am J Dig Dis (in press), 1975 33. Chandler AM, Johnson LR: Pentagastrin-stimulated incorporaof the trophic action of gastrin. Gastroenterology 66:718, 1974 tion of [UC]orotic acid into RNA of gastric and duodenal mucosa. 60. Johnson LR, Copeland EM, Dudrick SJ, et al: Structural and Proc Soc Exp Bioi Med 141:110-113, 1972 hormonal alterations in the gastrointestinal tract of parenterally 34. Johnson LR, Lichtenberger LM, Copeland, EM, et al: Action of fed rats. Gastroenterology 68:1177-1183,1975 gastron on gastrointestinal structure and function. Gastroenterol- 61. Castro GA, Johnson LR, Copeland EM, et al: Decreased intestinal ogy 68:1184-1192, 1975 disaccharidase and peroxidase activity in hyperalimented rats. 35. Daughaday WH, Garland JT: The sulfation factor hypothesis: Fed Proc 33:692, 1974 recent observations. In Growth and Growth Hormone. Edited by A 62. 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Korner A: Regulation of the rate of synthesis of messenger secretin and pentagastrin on acid secretion and parietal cell ribonucleic acid by growth hormone. Biochem J 92:449-456, 1964 number in rats. Gastroenterology 63:264-269, 1972 40. Pegg AE, Korner A: Growth hormone action on rat liver RNA 67. Tumpson DB, Johnson LR: Effect of secretin and cholecystokinin polymerase. Nature 205:904-905, 1965 on the response of the gastric fistula rat to pentagastrin. Proc Soc 41. Walsh JH, Grossman MI: Gastrin. N Eng J Med (in press), 1975 Exp Bioi Med 131:186-188, 1969 42. Johnston DH: A biopsy study of the gastric mucosa in postopera- 68. Grossman MI: Gastrointestinal hormones: Spectrum of actions tive patients with and without marginal ulcer. Am J Gastroenterol and structure-activity relations. In Endocrinology of the Gut. 46: 103-118, 1966 Edited by WY Chey and FP Brooks. Thorofare, N.J., Chas. B. 43. 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72. Hansky J, Royle JP, Soveny C, et al: Relationship of immunoreactivity to biological activity of gastrin. Gastroenterology 64:739; 1973 73. Debas HT, Grossman MI: Chemical bathing oxyntic gland area stimulate acid secretion. Gastroenterology 66:836, 1974