Effect of Alterations of Blood Glucose Lewels on Gastric Acid Secretion, Plasma Gastrin, and Plasma Osmolality in Man G. S T A C H E R MD, P. B A U E R PhD, D. S C H U L Z E , H. P O I N T N E R MD, and M. L A N D G R A F
In 16 experiments on 4 healthy subjects, the effect of procedures which alter blood glucose, ie, infusion of O.2 units/kg body wt/hr insulin and/or 0.66 g/kg body wt/hr glucose, on gastric acid secretion, plasma gastrin, and plasma osmolality was studied. Each subject underwent four different experimental procedures, each lasting 4 hr. All had in common one basal hour and the infusion of insulin in the second hour, but differed in the time of infusion of glucose or isotonic saline. To control for order effects, the four procedures were applied to the subjects in the form of a Latin square. Acid output was measured continuously by means of intragastric titration and a telemetering capsule; blood glucose, plasma gastrin, and plasma osmolality were determined in 15-min intervals. An inverse relationship between blood glucose and acid output was found: Low glucose levels were associated with high rates of acid secretion, high glucose levels with low acid secretion. No noticeable changes occurred in either plasmagastrin or plasma osmolality. These results reveal a determining influence of blood glucose levels on acid secretion. On the basis of earlier work in animals it is concluded that this influence is exerted via the reciprocal activities of the hypothalamic satiety and feeding centers.
The inhibitory effect of glucose administered orally into the stomach (1, 2), the d u o d e n u m (3-10), or the j e j u n u m (11, 12) on gastric acid secretion is well k n o w n and has been attributed to the attained intraluminal hyperosmolality (2, 7, 8, 10) or to the release of an enterogastrone and/or glucagon (1, 12). Parenterally administered glucose was found to inhibit basal acid secretion (6, 13-I8) as well as secretion stimulated by sham feeding or teasing with food in dogs (17, 19) and by histamine in rats (2). This effect was suggested to be due to an elevated plasma osmolality (13, 18, 19, 21), to an effect on the nervous phase of secretion (17), or to a reduction of p l a s m a gastrin (18). From the Psychophysiology Unit of the First Surgical and the Psychiatric Clinic, the Department of Medical Statistics and Documentation, and the First Medical Clinic, University of Vienna, Vienna, Austria. Address for reprint requests: Dr. G. Stacher, Psychophysiologisches Laboratorium, Psychiatrische Universit~tsklinik, Lazarettgasse 14, 1090 Wien, Austria. Digestive Diseases, Vol. 21, No. 7 (July 1976)
To elucidate the physiological significance of these possible m e c h a n i s m s , in this work gastric acid secretion, plasma gastrin, and plasma osmolality w e r e studied u n d e r the influence of p r o c e d u r e s which alter the blood glucose, ie, intravenous infusion of insulin and/or glucose. M A T E R I A L S AND M E T H O D S Subjects. Four healthy male students ranging in age from 21 to 25 years and in weight from 77 to 82 kg were chosen as subjects. None of them took any drugs at the time of the experiments. Measurement of Gastric Acid Secretion. Acid secretion was measured by means of the intragastric titration technique and a telemetering capsule (Heidelberg capsula) as an intragastric pH sensor (22). This technique has been shown to be a reliable method of measm-ing basal as well as histamine- (22-24) or insulin-stimulated (24) secretion. The capsules were calibrated in buffer solutions of pH 1 and 7 and thereafter swallowed by the subjects. The device was located in the fundic area and anchored to the subject's nose by a thin silk thread attached to it to assure
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EFFECTS OF ALTERING BLOOD GLUCOSE LEVELS a stable position. The signals emitted by the capsules were received by an antenna, amplified by a modified radio set, and recorded by a pen recorder. To avoid highfrequency interference the experiments were carried out in a screened room. The acidic gastric contents were neutralized by oral administration of 2.5 ml molar potassium bicarbonate, whenever the pH fell to a value below 1.5. The time between two such neutralizations was measured and assumed to correspond to the time during which 2.5 mEq hydrochloric acid were produced. Acid output was calculated as mEq per 15 min. Measurement of Blood Glucose Levels. True blood glucose was determined by the oxygen rate method, employing a Beckman oxygen sensor. Measurement of Plasma Gastrin Levels. Plasma gastrin was determined by means of a radioimmunoassay using antisera against synthetic human gastrin I (G 17) raised in rabbits. The mean level of plasma gastrin for fasting normals in our laboratory is 32.3 oz/ml -+ 8.1 SEM (N = 78). Plasma osmolality, 39.7 pg/ml -+ 1.3 SEM (N = 275), was determined electrometrically by means of a Knauer osmometer Type M. Procedure. The subjects came to the laboratory at 8:30 AM after an overnight fast. After swallowing the telemetering capsules, they lay on a bed in the screened room. A polyvinyl cannula was inserted into a vein of the left forearm and samples of blood for the estimation of blood glucose, plasma gastrin, and plasma osmolality were taken at 15-min intervals throughout the study. Glucose and osmolality were determined immediately, the samples for the determination of gastrin were stored after separation at -20 ~ C until the time of the assay. Each subject participated in four experiments with different procedures which had in common one basal hour. After this basal hour two separate cannulas were inserted into right forearm veins. In all experiments, 0.2 units/kg body wt glucagon-free insulin (Actrapid| Novo) diluted in 4.5 ml distilled water were infused in the following hour via one of the cannulas by means of a motor pump. The other cannula was used to infuse, also by motor pump, glucose or saline according to the four different experimental procedures (A, B, C, D) as indicated below. A. 2nd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 3rd hr: 225 ml isotonic (0.9%) saline 4th hr: nil B. 2nd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 3rd hr: as in 2nd hr 4th hr: nil C. 2nd hr: 225 ml isotonic saline 3rd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 4th hr: nil D. 2nd hr: 225 ml isotonic saline 3rd hr: as in 2nd hr 4th hr: 0.66 g/kg body wt glucose in 225 ml distilled water Design and Statistical Analysis of the Experiment. The four procedures were applied to the subjects in the form of a Latin-square design, each subject undergoing the four procedures in a different order. The interval between Digestive Diseases, Vol. 21, No. 7 (July 1976)
two experiments in any one subject was at least 4 days, but did not exceed 2 weeks. The sequence of the subjects entering the experiment was randomized. The analysis of the data of the four basal 15-rain periods was carried out according to a completely crossed threefactorial model (25) accounting for the factors "day" (lst-4th day), "periods" (lst--4th basal 15-rain period) and "subject" (subjects 1--4). The analysis of the treatment effects was performed on the differences between the values of the different variables in the four 15-min periods of the 2nd, 3rd, and 4th experimental hour, respectively, and the mean values over the basal hour. The analysis was carried out according to the Latin-square design with the four factors "treatment" (procedures A, B, C, D), "period" (the four 15rain periods in the 2nd, 3rd, and 4th experimental hours, respectively), "day" (lst-4th day), and "subject" (subjects 14). In order to account for an inhomogeneity of the initial values in that model, an analysis of covariance employing the initial values as covariables (26) was also performed. Since the analysis according to the Latin-square model did not reveal any systematic influence of the day factor, the differences between the 15-rain values of the 2nd, 3rd, and 4th hours and the mean values of the basal hour were also analyzed according to a completely crossed three-factorial model omitting the day factor.
RESULTS Basal H o u r . In this hour, none of the analyzed variables showed a significant (P < 0.05) influence of the day and the period factors, while there was an indication (P < 0.1) for an increase in acid production and blood glucose and for a decrease in plasma osmolality. These changes, however, were of no relevant magnitude; thus, the mean values over the four basal 15-rain periods were taken as initial values. Second hour. During simultaneous infusion of insulin and glucose (procedures A and B), there was a decrease of acid production with increasing glucose levels (Figure 1A and B). When saline was infused together with insulin (procedures C and D), blood glucose levels decreased to a minimum of 37 rag/100 ml (C) and 38 rag/100 ml (D) at the end of the 2nd hr, respectively, while acid secretion, which showed slightly falling (C) or rising (D) tendencies in the first 45 min, increased massively when the minimum of blood glucose was reached (Figure 1C and D). These changes were reflected by large F values for the treatment factor for both acid production and blood glucose (P "r 0.001) in the Latin square as well as in the factorial model. A comparison based on the factorial model (27) between the two pairs of procedures which were identical in the 2nd hr, ie, AB, and CD, revealed highly significant differences
565
STACHER ET AL (P "~0.001) for both acid output and blood glucose. The time course of acid secretion and blood glucose in procedures A and B vs C and D showed significantly different tendencies as indicated by the F values for the interaction treatment • time (P < 0.05). Blood glucose in procedures A and B reached a maximum 30 min after the start of the glucose infusion and then fell again; during infusion of saline (C and D) a more or less linear decrease of blood glucose occurred. The trends of the glucose levels in procedures A and B on the one hand and C and D on the other, differed significantly in the quadratic comparison over time (P ~0.001) as well as in the linear tendency (P < 0.001). In contrast to glucose, acid production in procedures A and B decreased gradually over the four periods, whereas in C and D there were increasing tendencies. The significant interaction treatment • time for acid secretion can be traced mainly to the difference between AB and CD in the linear comparison (P < 0.001). No significant differences between treatments were found for the variables plasma gastrin and plasma osmolality in either analysis. The height of the initial value was found to be of significant influence on acid output: The higher the initial value, the larger the decrease, and the smaller the increase, respectively (P < 0.01). Third hour. As in the 2nd hour, no significant differences between treatments were found for the variables plasma gastrin and osmolality. Massive treatment effects, by contrast, were observed for blood glucose (P < 0.001) and acid output. Glucose levels fell sharply after glucose infusion was stopped (A), but declined only slightly when glucose administration was continued (B). In procedure C the start of glucose infusion resulted in a sharp increase of blood glucose; in D, where saline was administered as in the previous hour, glucose levels rose only slightly. The overall effects differed between procedures A and B, A and C, B and D, and C and D [Tukey test (28), P < 0.01 ]. There were also significant differences between the procedures in the time course over the four 15-min periods, as indicated by the F values for the interaction treatment • time (Latin square, P < 0.05; factorial model, P < 0.001). Also, for acid secretion the treatment factor as well as the interaction treatment • time showed highly significant F values (P 0.001): In procedure A acid output rose sharply when the lowest glucose levels were reached. In procedure B acid secretion increased slightly at the end of the 3rd hour. In procedure C the hypo-
566
glycemia-induced increase of secretion was reversed with rising glucose levels. In procedure D the increasing tendency of acid output was maintained. Accordingly, there were significant differences in acid production averaged over the 3rd hour between procedures A and D, B and D, C and D (Tukey test, P < 0.01), and between B and C (P < 0.05). The differences between A and D and B and C are opposite in sign to the relative differences in glucose levels. While the acid output averaged over the whole hour did not differ between procedures A and C, the linear comparison over the four periods showed a highly significant difference .(P 0.001) due to the inverse trends of secretion as shown in Figure IA and D. The increasing tendency in procedure A was significantly larger than that in procedures B and D (P < 0.001), the increasing tendency in procedures B and D, on the other hand, differed significantly from the decreasing tendency in procedure C (P < 0.001 and P < 0.001, respectively). Because of the clearcut results, the statistical dependencies in some of the above comparisons can be ignored. The initial values were again of significant influence on the height of acid production (P < 0.01). Fourth Hour. In view of the results of the preceding two hours, the data of the last hour were analyzed mainly with respect to the relationship between glucose levels and gastric acid output. Once again, no influence of the treatment and time factors and their interaction was found for gastrin and osmolality. The interaction treatment • time, representing the different time course in the different procedures, showed large F values for blood glucose (in both analyses P < 0.001) and acid output (Latinsquare model, P < 0.1 ; factorial model, P < 0.001). As shown in Figure 1, blood glucose rose only slightly in procedure A but increased markedly under the infusion of glucose in procedure D. A linear comparison over the four periods based on the factorial model showed that this difference between A and D was significant (P < 0.001). In procedures B and C blood glucose levels showed falling tendencies, not distinguishable from each other by statistical means. The difference between the increasing tendency under A and D and the opposite tendency under B and C was highly significant (P < 0.001), as revealed by the corresponding linear comparison, orthogonal to the previous comparison of A vs D and B vs C. Acid output generally showed inverse tendencies as compared to the glucose levels, ie, decreasing in Digestive Diseases, Vot. 2I, No. 7(July 1976)
EFFECTS OF ALTERING BLOOD GLUCOSE LEVELS A and D and increasing in B and C; the increasing tendency in B was significantly larger than in C (P < 0.01). The difference between the falling tendency under A and D and the rising tendency under B and C was significant (P < 0.001). The obvious inverse relationship between blood glucose and acid output is clearly demonstrated by an inspection of the data of the 16 individual experiments on the 4 subjects. In 15 experiments, the linear tendencies of blood glucose and acid output were inverse. In 1 experiment of procedure D glucose levels increased so rapidly with the beginning of the glucose infusion that the blood glucose peak was reached in the first 15-min period while there was a slight decrease in the subsequent periods, so that the linear tendencies over the whole hour were in the same direction. DISCUSSION The results of the present study show that there is a definite relationship between blood glucose levels and gastric acid secretion. Rising glucose levels led to a diminution of gastric acid output, which was most accentuated in the periods with the highest glucose levels. By contrast, in the presence of low glucose levels an increase of gastric secretion took place. Both effects were reversed by an alteration of blood glucose, ie, by the start of an infusion of glucose or its discontinuation and the subsequent decrease of glucose levels as effectuated by exogenous and/or reactively liberated endogenous insulin. The explanation for the inhibitory effect of glucose on gastric acid secretion was, by some authors, suggested to be the release of a potent enterogastron and or glucagon (1, 12). While glucose instilled into the jejunum was shown to result in a significant rise in serum-immunoreactive glucagon (29), pharmacological doses of glucagon were found to inhibit gastric secretion much less than glucose instilled intrajejunally (12). Unfortunately, no determinations of the blood glucose levels were carried out in the latter authors' work, but it does not seem unreasonable to argue that hyperglycemia resulting from absorption of glucose might have accounted for the more potent inhibition of acid secretion as compared to the direct administration of glucagon. A number of authors suggested the inhibitory effect of glucose to be due to an alteration of intraluminal (2, 7, 8, 10) or plasma osmolality (13, 19, 21). However, the validity of such an exDigestive Diseases, Vol. 21, No. 7 (July 1976)
planation is challenged by the fact that intravenous glucose was found to have the same inhibitory effect in iso-, hypo-, and hypertonic solutions, if only the quantity was sufficient (14). Moreover, plasma hypertonicity induced by urea or saline was reported not to affect histamine-stimulated acid secretion in rats, whereas glucose-induced hyperosmolality caused inhibition (20). Plasma osmolality increased by means of saline, mannitol, or glucose was even reported to increase gastric hydrogen and chlorine concentrations but not to affect the output of gastric juice and of these ions (30). Moore (18) suggested that lowered plasma gastrin levels, as reported to occur after intravenous glucose loads (31, 32), might lead to the diminution of acid secretion after glucose administration. However, the results of this study indicate that neither changes in plasma osmolality nor in gastrin levels play any major role in the glucose-induced inhibition of gastric secretion, as neither of these two variables showed any systematic change in our experiment. Peripheral mechanisms based on alterations of the osmolality of the intestinal contents or on a release of an enterogastron or of glucagon induced by the contact of glucose with the intestinal mucosa could not have come into play in our study. However, they could play a role when glucose is present in the gut. While the above explanations of the inhibitory effect of glucose are not satisfactory, the work on the role of the central nerv.ous system in the regulation of food intake seems to offer the solution of the problem. There is good evidence that there are two opposing mechanisms in the hypothalamus one of which, situated in the lateral hypothalamus, initiates feeding while the other one, in the ventromedial hypothalamus, brings about satiation (33-35). In cats and dogs glucose infusions led to an increase of the frequency; of spikes recorded from satietycenter neurons, while the spike frequency of feeding-center neurons decreased (36). Saline infusions of the same osmolality did not produce such changes. Both feeding and satiety centers were found to have a regulatory influence on gastric motility (37) and acid secretion (38--40). Satiety-center damage caused by gold thioglucose led to an increased acid secretion in the rat and dog (40). Stimulation of the satiety center, by contrast, decreased and stimulation of the feeding center increased acid production (39). These findings strongly suggest that an analogous mechanism, based solely on the blood glucose levels as indicated by our study, can
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be held responsible for the inhibitory effect of glucose on gastric acid secretion in man also. ACKNOWLEDGMENTS F o r i n v a l u a b l e a s s i s t a n c e we are i n d e b t e d to Mrs. A l i z a Blau, Mrs. G i s e l h e i d S c h m i e r e r , a n d Mrs. Ilse L i n s .
REFERENCES 1. Friedman MHF: The influence of glucose administration on gastric secretion. Am J Physiol 126:495P, 1939 2. Shay H, Gershon-Cohen J, Fels SS, Siplet H: Concerning the influence of glucose on the response of the human stomach to test meals. Am J Dig Dis 9:363-367, 1942 3. Le Conte P: Fonctions gastro-intestinales. Etude Physiologique. Cellule 17:283-321, 1900 4. Kauders F, Porges O: Der Einfluss des Duodenalinhaltes auf die Magensekretion. Wien Klin Wochenschr 35:838-839, 1922 5. Gutzeit E: Uber die Beeinflussung der Magenreiz- und N0chternsekretion durch D0nndarmverweilsonden sowie durch duodenale und jejunale Nahrungszufuhr. Z Ges Exp Med 73:48-56, 1930 6. Matsuyama M: Influence of the introduction of glucose into the intestine upon the secretion of the gastric juice. Jpn J Gastroenterol 4:273-279, 1932 7. Shay H, Gershon-Cohen J, Fels SS: The role of the upper small intestine in the control of gastric secretion: The effect of neutral fat, fatty acid and soaps; the phase of gastric secretion influenced and the relative importance of the psychic and chemical phases. Ann Intern Med 13:294-307, 1939 8. Sircus W: Studies on tlae mechanisms in the duodenum inhibiting gastric secretion. Q J Exp Physio143:114-133, 1958 9. Konturek S, Grossman MI: Effect of perfusion of intestinal loops with acid, fat, or dextrose on gastric secretion. Gastroenterology 49:481-489, 1965 10. Ward AS, Wilkins RA, Cockel R, Windsor CWO: Duodenal inhibition of gastric secretion by osmotic agents in normal subjects and patients with duodenal ulcer. Gut 10:10201028, 1969 11. Kalk H, Meyer PF: Blutzuckerspiegel und Magensekretion. Z Klin Med 120:692-714, 1932 12. Christiansen J, Hendel L: Inhibition of pentagastrin-induced gastric acid secretion in man by intrajejunal glucose administration. Acta Chir Scand 140:246-248, 1974 13. Karmel J: [Yber den Einfluss intravenrs verabreichter hypertonischer Lrsungen auf die Magensekretion. Wien Klin Wochenschr 35:1007-1011, 1922 14. Okada S, Kuramochi K, Tsukahara T, Ooinoue T: Pancreatic function. IV. The humoroneural regulation of the gastric, pancreatic and biliary secretions. Arch Intern Med 43:446471, 1929 15. Babkin BP: Secretory Mechanism of the Digestive Glands. New York, Hoeber, 1950, p 651 16. Solomon SP, Spiro HM: The effects of glucagon and glucose on the human stomach. Am J Dig Dis 4:775-786, 1959 17. Dotevall G, Muren A: Effect of intravenous infusion of glucose on gastric secretory responses to feeding in Pavlov- and Heidenhain-pouch dogs. Acta Physiol Scand 52:234-241, 1961
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18. Moore JG: Gastric acid suppression by intravenous glucose solutions. Gastroenterology 64:1106-1110, 1973 19. Day JJ, Komraov SA: Glucose and gastric secretion. Am J Dig Dis 6:169-175, 1939 20. Adamkiewicz VW, Sacra PJ: Inhibition by sugars of histamine-induced gastric secretion. Arch Int Physiol Biochim 74:21-24, 1966 21. Thorsoe H: Reduction of gastric acid output in rats by glucose. Scand J Gastroenterol 6:319-321, 1971 22. Stavney LS, Hamilton T, Sircus W, Smith AN: Evaluation of the pH-sensitive telemetering capsule in the estimation of gastric secretory capacity. Am J Dig Dis 11:753-760, 1966 23. Connell AM, Waters TE: Assessment of gastric function by pH telemetering capsule. Lancet 2:227-230, 1964 24. Stacher G, Dinstl K: Measurement of basal, insulin and betazole stimulated acid secretion by intragastric titration and telemetry. Chir Gastroenterol 8:31-38, 1974 25. Scheff6 H: The Analysis of Variance. New York, John Wiley, 1959, pp 284-289 26. Stacher G, Bauer P, Brunner H, Grfinberger J: Gastric acid secretion, serum-gastrin levels and psychomotor function under the influence of placebo, insulin-hypoglycemia, and/or bromazepam. Int J Clin Pharmacol 13:1-10, 1976 27. Cochran WG, Cox GM: Experimental Designs, 2nd ed., New York, John Wiley, 1966 28. Scheff6 H: The Analysis of Variance. New York, John Wiley, 1959, pp 73-75 29. Dyck WP: Influence of intrajejunal glucose on pancreatic exocrine function in man. Gastroenterology 60:864-869, 1971 30. Powell DW, Hirschowitz BI: Effect of osmolar loading on stimulated gastric secretion in the dog. Am J Physiol 207:868-872, 1964 31. Young JD, Byrnes DJ, Chisholm DJ, Griffiths FB, Lazarus L: Radioimmunoassay of gastrin in human serum using antiserum against pentagastrin. J Nucl Med 10:746-748, 1969 32. Rehfeld JF, Stadil F: The effect of gastrin on basal- and glucose-stimulated insulin secretion in man. J Clin Invest 52:1415-1426, 1973 33. Anand BK, Brobeck JR: Localization of a "feeding center" in the hypothalamus of the rat. Proc Soc Exp Biol Med 77:323-324, 1951 34. Anand BK, Dua S, Shoenberg K: Hypothalamic control of food intake in cats and monkeys. J Physiol 127:143-152, 1955 35. Anand BK: Nervous regulation of food intake. Physiol Rev 41:677-708, 1961 36. Anand BK, Chhina GS, Sharma KN, Dua S, Baldev Singh: Activity of single neurons in the hypothalamic feeding centers: Effect of glucose. Am J Physiol 207:1146-1154, 1964 37. Sharma KN, Anand BK, Dua S, Baldev Singh: Role of stomach in regulation of activities of hypothalamic feeding centers. Am J Physiol 201:593-598, 1961 38. Ridley PT, Brooks FP: Alterations in gastric secretion following hypothalamic lesions producing hyperphagia. Am J Physiol 209:31%323, 1965 39. Misher A, Brooks FP: Electrical stimulation of hypothalamus and gastric secretion in the albino rat. Am J Physiol 211:403-406, 1966 40. Nagamachi Y: Effect of satiety center damage on food intake, blood glucose and gastric secretion in dogs. Am J Dig Dis 17:139-148, 1972
Digestive Diseases, Vol. 21, No. 7 (July 1976)
In 16 experiments on 4 healthy subjects, the effect of procedures which alter blood glucose, ie, infusion of O.2 units/kg body wt/hr insulin and/or 0.66 g/kg body wt/hr glucose, on gastric acid secretion, plasma gastrin, and plasma osmolality was studied. Each subject underwent four different experimental procedures, each lasting 4 hr. All had in common one basal hour and the infusion of insulin in the second hour, but differed in the time of infusion of glucose or isotonic saline. To control for order effects, the four procedures were applied to the subjects in the form of a Latin square. Acid output was measured continuously by means of intragastric titration and a telemetering capsule; blood glucose, plasma gastrin, and plasma osmolality were determined in 15-min intervals. An inverse relationship between blood glucose and acid output was found: Low glucose levels were associated with high rates of acid secretion, high glucose levels with low acid secretion. No noticeable changes occurred in either plasmagastrin or plasma osmolality. These results reveal a determining influence of blood glucose levels on acid secretion. On the basis of earlier work in animals it is concluded that this influence is exerted via the reciprocal activities of the hypothalamic satiety and feeding centers.
The inhibitory effect of glucose administered orally into the stomach (1, 2), the d u o d e n u m (3-10), or the j e j u n u m (11, 12) on gastric acid secretion is well k n o w n and has been attributed to the attained intraluminal hyperosmolality (2, 7, 8, 10) or to the release of an enterogastrone and/or glucagon (1, 12). Parenterally administered glucose was found to inhibit basal acid secretion (6, 13-I8) as well as secretion stimulated by sham feeding or teasing with food in dogs (17, 19) and by histamine in rats (2). This effect was suggested to be due to an elevated plasma osmolality (13, 18, 19, 21), to an effect on the nervous phase of secretion (17), or to a reduction of p l a s m a gastrin (18). From the Psychophysiology Unit of the First Surgical and the Psychiatric Clinic, the Department of Medical Statistics and Documentation, and the First Medical Clinic, University of Vienna, Vienna, Austria. Address for reprint requests: Dr. G. Stacher, Psychophysiologisches Laboratorium, Psychiatrische Universit~tsklinik, Lazarettgasse 14, 1090 Wien, Austria. Digestive Diseases, Vol. 21, No. 7 (July 1976)
To elucidate the physiological significance of these possible m e c h a n i s m s , in this work gastric acid secretion, plasma gastrin, and plasma osmolality w e r e studied u n d e r the influence of p r o c e d u r e s which alter the blood glucose, ie, intravenous infusion of insulin and/or glucose. M A T E R I A L S AND M E T H O D S Subjects. Four healthy male students ranging in age from 21 to 25 years and in weight from 77 to 82 kg were chosen as subjects. None of them took any drugs at the time of the experiments. Measurement of Gastric Acid Secretion. Acid secretion was measured by means of the intragastric titration technique and a telemetering capsule (Heidelberg capsula) as an intragastric pH sensor (22). This technique has been shown to be a reliable method of measm-ing basal as well as histamine- (22-24) or insulin-stimulated (24) secretion. The capsules were calibrated in buffer solutions of pH 1 and 7 and thereafter swallowed by the subjects. The device was located in the fundic area and anchored to the subject's nose by a thin silk thread attached to it to assure
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Digestive Diseases, Vol. 21, No. 7 (July 1976)
EFFECTS OF ALTERING BLOOD GLUCOSE LEVELS a stable position. The signals emitted by the capsules were received by an antenna, amplified by a modified radio set, and recorded by a pen recorder. To avoid highfrequency interference the experiments were carried out in a screened room. The acidic gastric contents were neutralized by oral administration of 2.5 ml molar potassium bicarbonate, whenever the pH fell to a value below 1.5. The time between two such neutralizations was measured and assumed to correspond to the time during which 2.5 mEq hydrochloric acid were produced. Acid output was calculated as mEq per 15 min. Measurement of Blood Glucose Levels. True blood glucose was determined by the oxygen rate method, employing a Beckman oxygen sensor. Measurement of Plasma Gastrin Levels. Plasma gastrin was determined by means of a radioimmunoassay using antisera against synthetic human gastrin I (G 17) raised in rabbits. The mean level of plasma gastrin for fasting normals in our laboratory is 32.3 oz/ml -+ 8.1 SEM (N = 78). Plasma osmolality, 39.7 pg/ml -+ 1.3 SEM (N = 275), was determined electrometrically by means of a Knauer osmometer Type M. Procedure. The subjects came to the laboratory at 8:30 AM after an overnight fast. After swallowing the telemetering capsules, they lay on a bed in the screened room. A polyvinyl cannula was inserted into a vein of the left forearm and samples of blood for the estimation of blood glucose, plasma gastrin, and plasma osmolality were taken at 15-min intervals throughout the study. Glucose and osmolality were determined immediately, the samples for the determination of gastrin were stored after separation at -20 ~ C until the time of the assay. Each subject participated in four experiments with different procedures which had in common one basal hour. After this basal hour two separate cannulas were inserted into right forearm veins. In all experiments, 0.2 units/kg body wt glucagon-free insulin (Actrapid| Novo) diluted in 4.5 ml distilled water were infused in the following hour via one of the cannulas by means of a motor pump. The other cannula was used to infuse, also by motor pump, glucose or saline according to the four different experimental procedures (A, B, C, D) as indicated below. A. 2nd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 3rd hr: 225 ml isotonic (0.9%) saline 4th hr: nil B. 2nd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 3rd hr: as in 2nd hr 4th hr: nil C. 2nd hr: 225 ml isotonic saline 3rd hr: 0.66 g/kg body wt glucose in 225 ml distilled water 4th hr: nil D. 2nd hr: 225 ml isotonic saline 3rd hr: as in 2nd hr 4th hr: 0.66 g/kg body wt glucose in 225 ml distilled water Design and Statistical Analysis of the Experiment. The four procedures were applied to the subjects in the form of a Latin-square design, each subject undergoing the four procedures in a different order. The interval between Digestive Diseases, Vol. 21, No. 7 (July 1976)
two experiments in any one subject was at least 4 days, but did not exceed 2 weeks. The sequence of the subjects entering the experiment was randomized. The analysis of the data of the four basal 15-rain periods was carried out according to a completely crossed threefactorial model (25) accounting for the factors "day" (lst-4th day), "periods" (lst--4th basal 15-rain period) and "subject" (subjects 1--4). The analysis of the treatment effects was performed on the differences between the values of the different variables in the four 15-min periods of the 2nd, 3rd, and 4th experimental hour, respectively, and the mean values over the basal hour. The analysis was carried out according to the Latin-square design with the four factors "treatment" (procedures A, B, C, D), "period" (the four 15rain periods in the 2nd, 3rd, and 4th experimental hours, respectively), "day" (lst-4th day), and "subject" (subjects 14). In order to account for an inhomogeneity of the initial values in that model, an analysis of covariance employing the initial values as covariables (26) was also performed. Since the analysis according to the Latin-square model did not reveal any systematic influence of the day factor, the differences between the 15-rain values of the 2nd, 3rd, and 4th hours and the mean values of the basal hour were also analyzed according to a completely crossed three-factorial model omitting the day factor.
RESULTS Basal H o u r . In this hour, none of the analyzed variables showed a significant (P < 0.05) influence of the day and the period factors, while there was an indication (P < 0.1) for an increase in acid production and blood glucose and for a decrease in plasma osmolality. These changes, however, were of no relevant magnitude; thus, the mean values over the four basal 15-rain periods were taken as initial values. Second hour. During simultaneous infusion of insulin and glucose (procedures A and B), there was a decrease of acid production with increasing glucose levels (Figure 1A and B). When saline was infused together with insulin (procedures C and D), blood glucose levels decreased to a minimum of 37 rag/100 ml (C) and 38 rag/100 ml (D) at the end of the 2nd hr, respectively, while acid secretion, which showed slightly falling (C) or rising (D) tendencies in the first 45 min, increased massively when the minimum of blood glucose was reached (Figure 1C and D). These changes were reflected by large F values for the treatment factor for both acid production and blood glucose (P "r 0.001) in the Latin square as well as in the factorial model. A comparison based on the factorial model (27) between the two pairs of procedures which were identical in the 2nd hr, ie, AB, and CD, revealed highly significant differences
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STACHER ET AL (P "~0.001) for both acid output and blood glucose. The time course of acid secretion and blood glucose in procedures A and B vs C and D showed significantly different tendencies as indicated by the F values for the interaction treatment • time (P < 0.05). Blood glucose in procedures A and B reached a maximum 30 min after the start of the glucose infusion and then fell again; during infusion of saline (C and D) a more or less linear decrease of blood glucose occurred. The trends of the glucose levels in procedures A and B on the one hand and C and D on the other, differed significantly in the quadratic comparison over time (P ~0.001) as well as in the linear tendency (P < 0.001). In contrast to glucose, acid production in procedures A and B decreased gradually over the four periods, whereas in C and D there were increasing tendencies. The significant interaction treatment • time for acid secretion can be traced mainly to the difference between AB and CD in the linear comparison (P < 0.001). No significant differences between treatments were found for the variables plasma gastrin and plasma osmolality in either analysis. The height of the initial value was found to be of significant influence on acid output: The higher the initial value, the larger the decrease, and the smaller the increase, respectively (P < 0.01). Third hour. As in the 2nd hour, no significant differences between treatments were found for the variables plasma gastrin and osmolality. Massive treatment effects, by contrast, were observed for blood glucose (P < 0.001) and acid output. Glucose levels fell sharply after glucose infusion was stopped (A), but declined only slightly when glucose administration was continued (B). In procedure C the start of glucose infusion resulted in a sharp increase of blood glucose; in D, where saline was administered as in the previous hour, glucose levels rose only slightly. The overall effects differed between procedures A and B, A and C, B and D, and C and D [Tukey test (28), P < 0.01 ]. There were also significant differences between the procedures in the time course over the four 15-min periods, as indicated by the F values for the interaction treatment • time (Latin square, P < 0.05; factorial model, P < 0.001). Also, for acid secretion the treatment factor as well as the interaction treatment • time showed highly significant F values (P 0.001): In procedure A acid output rose sharply when the lowest glucose levels were reached. In procedure B acid secretion increased slightly at the end of the 3rd hour. In procedure C the hypo-
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glycemia-induced increase of secretion was reversed with rising glucose levels. In procedure D the increasing tendency of acid output was maintained. Accordingly, there were significant differences in acid production averaged over the 3rd hour between procedures A and D, B and D, C and D (Tukey test, P < 0.01), and between B and C (P < 0.05). The differences between A and D and B and C are opposite in sign to the relative differences in glucose levels. While the acid output averaged over the whole hour did not differ between procedures A and C, the linear comparison over the four periods showed a highly significant difference .(P 0.001) due to the inverse trends of secretion as shown in Figure IA and D. The increasing tendency in procedure A was significantly larger than that in procedures B and D (P < 0.001), the increasing tendency in procedures B and D, on the other hand, differed significantly from the decreasing tendency in procedure C (P < 0.001 and P < 0.001, respectively). Because of the clearcut results, the statistical dependencies in some of the above comparisons can be ignored. The initial values were again of significant influence on the height of acid production (P < 0.01). Fourth Hour. In view of the results of the preceding two hours, the data of the last hour were analyzed mainly with respect to the relationship between glucose levels and gastric acid output. Once again, no influence of the treatment and time factors and their interaction was found for gastrin and osmolality. The interaction treatment • time, representing the different time course in the different procedures, showed large F values for blood glucose (in both analyses P < 0.001) and acid output (Latinsquare model, P < 0.1 ; factorial model, P < 0.001). As shown in Figure 1, blood glucose rose only slightly in procedure A but increased markedly under the infusion of glucose in procedure D. A linear comparison over the four periods based on the factorial model showed that this difference between A and D was significant (P < 0.001). In procedures B and C blood glucose levels showed falling tendencies, not distinguishable from each other by statistical means. The difference between the increasing tendency under A and D and the opposite tendency under B and C was highly significant (P < 0.001), as revealed by the corresponding linear comparison, orthogonal to the previous comparison of A vs D and B vs C. Acid output generally showed inverse tendencies as compared to the glucose levels, ie, decreasing in Digestive Diseases, Vot. 2I, No. 7(July 1976)
EFFECTS OF ALTERING BLOOD GLUCOSE LEVELS A and D and increasing in B and C; the increasing tendency in B was significantly larger than in C (P < 0.01). The difference between the falling tendency under A and D and the rising tendency under B and C was significant (P < 0.001). The obvious inverse relationship between blood glucose and acid output is clearly demonstrated by an inspection of the data of the 16 individual experiments on the 4 subjects. In 15 experiments, the linear tendencies of blood glucose and acid output were inverse. In 1 experiment of procedure D glucose levels increased so rapidly with the beginning of the glucose infusion that the blood glucose peak was reached in the first 15-min period while there was a slight decrease in the subsequent periods, so that the linear tendencies over the whole hour were in the same direction. DISCUSSION The results of the present study show that there is a definite relationship between blood glucose levels and gastric acid secretion. Rising glucose levels led to a diminution of gastric acid output, which was most accentuated in the periods with the highest glucose levels. By contrast, in the presence of low glucose levels an increase of gastric secretion took place. Both effects were reversed by an alteration of blood glucose, ie, by the start of an infusion of glucose or its discontinuation and the subsequent decrease of glucose levels as effectuated by exogenous and/or reactively liberated endogenous insulin. The explanation for the inhibitory effect of glucose on gastric acid secretion was, by some authors, suggested to be the release of a potent enterogastron and or glucagon (1, 12). While glucose instilled into the jejunum was shown to result in a significant rise in serum-immunoreactive glucagon (29), pharmacological doses of glucagon were found to inhibit gastric secretion much less than glucose instilled intrajejunally (12). Unfortunately, no determinations of the blood glucose levels were carried out in the latter authors' work, but it does not seem unreasonable to argue that hyperglycemia resulting from absorption of glucose might have accounted for the more potent inhibition of acid secretion as compared to the direct administration of glucagon. A number of authors suggested the inhibitory effect of glucose to be due to an alteration of intraluminal (2, 7, 8, 10) or plasma osmolality (13, 19, 21). However, the validity of such an exDigestive Diseases, Vol. 21, No. 7 (July 1976)
planation is challenged by the fact that intravenous glucose was found to have the same inhibitory effect in iso-, hypo-, and hypertonic solutions, if only the quantity was sufficient (14). Moreover, plasma hypertonicity induced by urea or saline was reported not to affect histamine-stimulated acid secretion in rats, whereas glucose-induced hyperosmolality caused inhibition (20). Plasma osmolality increased by means of saline, mannitol, or glucose was even reported to increase gastric hydrogen and chlorine concentrations but not to affect the output of gastric juice and of these ions (30). Moore (18) suggested that lowered plasma gastrin levels, as reported to occur after intravenous glucose loads (31, 32), might lead to the diminution of acid secretion after glucose administration. However, the results of this study indicate that neither changes in plasma osmolality nor in gastrin levels play any major role in the glucose-induced inhibition of gastric secretion, as neither of these two variables showed any systematic change in our experiment. Peripheral mechanisms based on alterations of the osmolality of the intestinal contents or on a release of an enterogastron or of glucagon induced by the contact of glucose with the intestinal mucosa could not have come into play in our study. However, they could play a role when glucose is present in the gut. While the above explanations of the inhibitory effect of glucose are not satisfactory, the work on the role of the central nerv.ous system in the regulation of food intake seems to offer the solution of the problem. There is good evidence that there are two opposing mechanisms in the hypothalamus one of which, situated in the lateral hypothalamus, initiates feeding while the other one, in the ventromedial hypothalamus, brings about satiation (33-35). In cats and dogs glucose infusions led to an increase of the frequency; of spikes recorded from satietycenter neurons, while the spike frequency of feeding-center neurons decreased (36). Saline infusions of the same osmolality did not produce such changes. Both feeding and satiety centers were found to have a regulatory influence on gastric motility (37) and acid secretion (38--40). Satiety-center damage caused by gold thioglucose led to an increased acid secretion in the rat and dog (40). Stimulation of the satiety center, by contrast, decreased and stimulation of the feeding center increased acid production (39). These findings strongly suggest that an analogous mechanism, based solely on the blood glucose levels as indicated by our study, can
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be held responsible for the inhibitory effect of glucose on gastric acid secretion in man also. ACKNOWLEDGMENTS F o r i n v a l u a b l e a s s i s t a n c e we are i n d e b t e d to Mrs. A l i z a Blau, Mrs. G i s e l h e i d S c h m i e r e r , a n d Mrs. Ilse L i n s .
REFERENCES 1. Friedman MHF: The influence of glucose administration on gastric secretion. Am J Physiol 126:495P, 1939 2. Shay H, Gershon-Cohen J, Fels SS, Siplet H: Concerning the influence of glucose on the response of the human stomach to test meals. Am J Dig Dis 9:363-367, 1942 3. Le Conte P: Fonctions gastro-intestinales. Etude Physiologique. Cellule 17:283-321, 1900 4. Kauders F, Porges O: Der Einfluss des Duodenalinhaltes auf die Magensekretion. Wien Klin Wochenschr 35:838-839, 1922 5. Gutzeit E: Uber die Beeinflussung der Magenreiz- und N0chternsekretion durch D0nndarmverweilsonden sowie durch duodenale und jejunale Nahrungszufuhr. Z Ges Exp Med 73:48-56, 1930 6. Matsuyama M: Influence of the introduction of glucose into the intestine upon the secretion of the gastric juice. Jpn J Gastroenterol 4:273-279, 1932 7. Shay H, Gershon-Cohen J, Fels SS: The role of the upper small intestine in the control of gastric secretion: The effect of neutral fat, fatty acid and soaps; the phase of gastric secretion influenced and the relative importance of the psychic and chemical phases. Ann Intern Med 13:294-307, 1939 8. Sircus W: Studies on tlae mechanisms in the duodenum inhibiting gastric secretion. Q J Exp Physio143:114-133, 1958 9. Konturek S, Grossman MI: Effect of perfusion of intestinal loops with acid, fat, or dextrose on gastric secretion. Gastroenterology 49:481-489, 1965 10. Ward AS, Wilkins RA, Cockel R, Windsor CWO: Duodenal inhibition of gastric secretion by osmotic agents in normal subjects and patients with duodenal ulcer. Gut 10:10201028, 1969 11. Kalk H, Meyer PF: Blutzuckerspiegel und Magensekretion. Z Klin Med 120:692-714, 1932 12. Christiansen J, Hendel L: Inhibition of pentagastrin-induced gastric acid secretion in man by intrajejunal glucose administration. Acta Chir Scand 140:246-248, 1974 13. Karmel J: [Yber den Einfluss intravenrs verabreichter hypertonischer Lrsungen auf die Magensekretion. Wien Klin Wochenschr 35:1007-1011, 1922 14. Okada S, Kuramochi K, Tsukahara T, Ooinoue T: Pancreatic function. IV. The humoroneural regulation of the gastric, pancreatic and biliary secretions. Arch Intern Med 43:446471, 1929 15. Babkin BP: Secretory Mechanism of the Digestive Glands. New York, Hoeber, 1950, p 651 16. Solomon SP, Spiro HM: The effects of glucagon and glucose on the human stomach. Am J Dig Dis 4:775-786, 1959 17. Dotevall G, Muren A: Effect of intravenous infusion of glucose on gastric secretory responses to feeding in Pavlov- and Heidenhain-pouch dogs. Acta Physiol Scand 52:234-241, 1961
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18. Moore JG: Gastric acid suppression by intravenous glucose solutions. Gastroenterology 64:1106-1110, 1973 19. Day JJ, Komraov SA: Glucose and gastric secretion. Am J Dig Dis 6:169-175, 1939 20. Adamkiewicz VW, Sacra PJ: Inhibition by sugars of histamine-induced gastric secretion. Arch Int Physiol Biochim 74:21-24, 1966 21. Thorsoe H: Reduction of gastric acid output in rats by glucose. Scand J Gastroenterol 6:319-321, 1971 22. Stavney LS, Hamilton T, Sircus W, Smith AN: Evaluation of the pH-sensitive telemetering capsule in the estimation of gastric secretory capacity. Am J Dig Dis 11:753-760, 1966 23. Connell AM, Waters TE: Assessment of gastric function by pH telemetering capsule. Lancet 2:227-230, 1964 24. Stacher G, Dinstl K: Measurement of basal, insulin and betazole stimulated acid secretion by intragastric titration and telemetry. Chir Gastroenterol 8:31-38, 1974 25. Scheff6 H: The Analysis of Variance. New York, John Wiley, 1959, pp 284-289 26. Stacher G, Bauer P, Brunner H, Grfinberger J: Gastric acid secretion, serum-gastrin levels and psychomotor function under the influence of placebo, insulin-hypoglycemia, and/or bromazepam. Int J Clin Pharmacol 13:1-10, 1976 27. Cochran WG, Cox GM: Experimental Designs, 2nd ed., New York, John Wiley, 1966 28. Scheff6 H: The Analysis of Variance. New York, John Wiley, 1959, pp 73-75 29. Dyck WP: Influence of intrajejunal glucose on pancreatic exocrine function in man. Gastroenterology 60:864-869, 1971 30. Powell DW, Hirschowitz BI: Effect of osmolar loading on stimulated gastric secretion in the dog. Am J Physiol 207:868-872, 1964 31. Young JD, Byrnes DJ, Chisholm DJ, Griffiths FB, Lazarus L: Radioimmunoassay of gastrin in human serum using antiserum against pentagastrin. J Nucl Med 10:746-748, 1969 32. Rehfeld JF, Stadil F: The effect of gastrin on basal- and glucose-stimulated insulin secretion in man. J Clin Invest 52:1415-1426, 1973 33. Anand BK, Brobeck JR: Localization of a "feeding center" in the hypothalamus of the rat. Proc Soc Exp Biol Med 77:323-324, 1951 34. Anand BK, Dua S, Shoenberg K: Hypothalamic control of food intake in cats and monkeys. J Physiol 127:143-152, 1955 35. Anand BK: Nervous regulation of food intake. Physiol Rev 41:677-708, 1961 36. Anand BK, Chhina GS, Sharma KN, Dua S, Baldev Singh: Activity of single neurons in the hypothalamic feeding centers: Effect of glucose. Am J Physiol 207:1146-1154, 1964 37. Sharma KN, Anand BK, Dua S, Baldev Singh: Role of stomach in regulation of activities of hypothalamic feeding centers. Am J Physiol 201:593-598, 1961 38. Ridley PT, Brooks FP: Alterations in gastric secretion following hypothalamic lesions producing hyperphagia. Am J Physiol 209:31%323, 1965 39. Misher A, Brooks FP: Electrical stimulation of hypothalamus and gastric secretion in the albino rat. Am J Physiol 211:403-406, 1966 40. Nagamachi Y: Effect of satiety center damage on food intake, blood glucose and gastric secretion in dogs. Am J Dig Dis 17:139-148, 1972
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