GASTROENTEROLOGY. Official Publication of the American Gastroenterological Association. COPTRIGHT 1970 THE WILLIA148 & WILI<IN8 Co.

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1 GASTROENTEROLOGY Official Publication of the American Gastroenterological Association COPTRGHT 1970 THE WLLA148 & WL<N8 Co. VOLUME 58 April 1970 NUMBER 4 PROFLE OF GASTRC POTENTAL DFFERENCE N MAN Effects of aspirin, alcohol, bile, and endogenous acid MCHAEL G. GEALL, M.B., SDNEY F. PHLLPS, M.D., AND W. H. J. SUMMERSKLL, D.M. Gastroenterology Unit, Mayo Clinic and Mayo Foundation, Rochester, Minnesota Measurement of transmural potential difference (PD) of the human stomach using a reference electrode placed in a peripheral vein showed that PD was higher in the body of the stomach than in the antrum. Stimulation of gastric acid secretion by betazole caused only a trivial and temporary decrease in PD below the basal value, the decrease occurring shortly after the stimulus was injected and prior to the onset of peak acid output. The administration of acetylsalicylic acid, alcohol, or duodenal contents rich in bile and pancreatic enzymes all caused rapid and profound reductions in gastric PD. Maintenance of gastric PD may be a sensitive indicator of mucosal integrity in man, since changes in PD occur with agents shown to cause damage by other methods in other species. Since changes in transmural electric potential difference (PD) in the stomach coincide with and may reflect certain types of injury to the gastric mucosa in animals,! we have investigated the effects on gastric PD in man of agents shown to cause histological or permeability changes in the stomach by other methods or in other species. 2-5 As a preliminary, PD at various sites in the stomach and Received August 28, Accepted October 24, Address requests for reprints to: Dr. W. H. J. Summerskill, Mayo Clinic, Rochester, Minnesota This investigation was supported in part by Research Grant AM-6oo8 from the National nstitutes of Health, United States Public Health Service. 437 adjacent organs was delineated. Earlier measurements of PD, using skin as the site of the reference electrode, have yielded variable results. 6 Moreover, changes in PD occur between the stomach and duodenum or esophagus 7-11 and regional differences in gastric PD in animals 12 are likely to apply in man. 7 Therefore, we used the technique of Andersson and Grossman,1 which employs peripheral venous blood for the reference electrode and gives reproducible results,t3 to define the profile of PD throughout the stomach and at the junctional zones of the organ in man. The precise interrelationships between PD and physiological or pathological factors affecting function or structure in the stomach are unknown. Transmural PD S located in the mucosa and has

2 438 GEALL ET AL. Vol. 58, No.4 o...l...j -L_.-----L-_ L - -._ J L...L L L J..._ J.... L 2 em pulls at each nterval FG. 1. PD profile of human upper gastrointestinal tract. Mucosa negative to serosa (mean ± SE, n = 9). been considered essential to or influenced by acid secretion Endogenous acid secretion enhances gastric mucosal injury due to aspirin and alcohol.19 Mucosal injury reduces PD in animals l 20 and an intact gastric mucosa is believed to be essential for maintenance of the ionic gradients with which gastric mucosal PD is associated intimately.20 Aspirin,2 alcoho,3 and bile 4 impair the gastric mucosal barrier to ionic diffusion in animals and various studies have incriminated each of t hese agents as causing morphological changes in t he human gastric mucosa. Therefore, the effects of these compounds on PD in the healthy stomach were investigated. n addition, PD was determined before and during stimulation of gastric secretion with betazole. Material and Methods Studies were made in the fasting state on 9 healthy male volunteers, whose ages ranged from 23 to 38 years, and prior to gastric surgery on 2 patients with the Zollinger-Ellison syndrome. The method of measuring gastric PD 13 is similar to that used in animals by Davenport and colleagues' and was applied originally to man by Andersson and Grossman. 7 The e lectrolyte bridges consisted of polyethylene tubes containing a saturated solution of KC in 3% agar. After gas sterilization, the reference electrolyte bridge (inner diameter, 0.08 cm) was placed in a peripheral vein through a 15-gauge needle. The gastrointestinal electrolyte bridge was a polyethylene tube (inner diameter, 0.17 cm) filled with a saturated solution of KC in 3% agar. Placement of this electrode and subsequent verification of its position were made under fluoroscopic control. The electrolyte bridges led to two beakers containing saturated KC solution, in which two balanced Calomel half-cells (Beckman fiber junction type) were placed. These were connected to a ph meter (Beckman Expandomatic as a high impedance direct current millivolt meter and PD was recorded continuously by a visual recorder (Honeywell Visicorder model 1508) calibrated at the beginning and end of each study. For observations relating to acid secretion, aspirin, bile, and alcohol, PD was measured at 3-min intervals. The anatomical profile of PD in the stomach, duodenum, and esophagus was constructed from 9 healthy volunteers by placement of the electrolyte bridge initially in the postbulbar duodenum and t hereafter by withdrawal of the bridge by 2 -cm pulls under fluoroscopic control. Since the bridge passed unpredictably up either the lesser or the greater curvature of the stomach, the length of the PD profile necessarily differed and therefore is plotted with an interrupted line between the gastroduodenal and gastroesophageal junctions (fig. 1) to indicate the apparent variations in stomach length. To record the effect of aspirin, alcohol, or bile (duodenal contents) on gastric PD, studies were made in healthy volunteers who lay on their left side to encourage passage of test material down the greater curvature toward the region of the body of the stomach, where PD was measured. As a preliminary, gastric PD was monitored continuously until stable. Fifty milliliters of normal saline then were given by mouth as a control; thereafter, solutions of aspirin or alcohol (by mouth) or of bile (by Levin tube) w ere given. All solutions previously were warmed t o 37 C. Aspirin was given in one of two doses (650 and 1300 mg) as finely powdered acetylsalicylic acid suspended in 50 ml of normal saline (ph 2.35 to 2.50). Alcohol was given either as a mixture containing 25 ml of 80-proof bourbon whiskey and 25 ml of normal saline or as 50 ml of undiluted bourbon whiskey. For studies employing bile (duodenal contents rich in biliary and pancreatic secretion), duodenal juice was obtained from fasting healthy individuals after pancreozymin-cholecystokinin secretion had been stimulated maximally by lyophilized cholecystokinin (0.25 to 0.35 Crick Harper-Raper units per kg per min, Gastro-

3 April 1970 GASTRC POTENTAL DFFERENCE 439 ntestinal Hormone Research Unit, Karolinska nstitutet, Stockholm, Sweden) given by vein. Twenty-five milliliters of bile then were delivered to a site 4 cm proximal to that of PD measurement. The effect of maximal betazole stimulation on gastric PD was studied in 8 healthy volunteers and PD during basal acid secretion in 2 patients with the Zollinger-Ellison syndrome. The latter diagnosis was supported by gastric secretory studies, laparotomy, histological study, and bioassay of gastrin-containing tumor tissue. The electrolyte bridge was positioned in the body of the stomach and a Levin tube was sited in the most dependent part of the antrum. After aspiration of fasting juice, a 1-hr basal collection was made. Then betazole (1.5 mg per kg) was given by intramuscular injection and secretions were collected for seven consecutive 15- min periods by continuous mechanical and intermittent hand suction. Results Anatomical profile of PD in the upper gastrointestinal tract (fig. 1). Duodenal PD was always less than 10 mv (mucosa negative) and, at the estimated (radiologically) duodenopyloric junction, an immediate and highly significant (P < 0.001) increase in negative mucosal PD was observed. A further significant (P < 0.001) increase was found in the region of the junction of the antrum and body of the stomach. At the estimated gastroesophageal junction, the PD decreased rapidly to 12 mv (P < 0.001) as the electrode passed orad. Mean values for PD (±SE) were 6.2 ± 0.5 mv in the duodenum, 24.9 ± 2.5 mv in the antrum, 35.5 ± 0.5 mv in the body of the stomach, and 10.1 ± 1.3 mv in the esophagus. Effects of acetylsalicylic acid (fig. 2). n the smaller dose (650 mg), ingestion of aspirin was followed by a significant (P < 0.01) decrease in gastric PD within 6 min; some reduction in PD (P < 0.05) persisted for 1 hr. The depression of PD was significantly greater (P < 0.01) with the larger dose of aspirin (1300 mg) than with the smaller. The onset was evident more immediately, occurring at 3 min (P < 0.001), and some effect persisted until the test was completed 1 hr later. Normal saline caused no change in PD l, ",, 1,,, krt1-+i-hl" A, 1 e : ' \ 1 Aspirin (650 mg), "t ("00 \ ' "1 " m,1 i ",. Aspirin Time in 3 minute intervals FG. 2. Effect of aspirin and saline control on PD of human gastric mucosa. Mucosa negative to peripheral blood (mean ± SE, n = 10).

4 440 GEALL ET AL. Vol. 58, No. 4 f.' r : :,0 \1 i (50 mil \ 28 r. 11 ". -'l26:'" / 1St Time in 3 minute intervals FG. 3. Effect of alcohol on PD of human gastric mucosa. Mucosa negative to peripheral blood (mean ± SE, n = 7). Effects of alcohol on gastric PD (fig. S). Alcohol in both concentrations caused reductions in PD within 3 min of ingestion, that with the larger dose (P < 0.02) being greater than that with the smaller (P < 0.05). Maximal depression occurred in 12 min and PD remained significantly different (P < 0.01) from that measured in the control period for 20 min with the diluted whiskey and for 30 min with the undiluted whiskey. Effects of bile-stained duodenal contents on gastric PD (fig. 4). The administration of bile caused reduction in gastric PD within 3 min (P < 0.02). The decrease was greatest at 15 min (P < 0.01) and persisted until the study ended at 30 min (P < 0.05). Gastric PD during acid secretion (fig. 5). Stimulation of gastric acid secretion by betazole resulted in a small but significant (P < 0.01) decrease in PD shortly after the stimulus was inj ected. PD returned to preinjection levels within 15 min and prior to the onset of peak acid output. During maximal rates of gastric secretion, PD did not differ significantly from that measured during the control period, despite a 6-fold increase in acid output. Thus, no significant correlation between gastric PD and either basal or peak acid output was observed. n fact, PD was normal in the 2 patients with the Zollinger-Ellison syndrome under basal circumstances (39 and 43 mv), although acid output at these times was 30 and 63 meq per hr, respectively. Discussion The values for duodenal, esophageal, and gastric PD's which we found are similar to some reported from 5 p ersons studied with a similar technique by Andersson and Grossman n addition, the finding that PD in the antrum is lower than in the body of the stomach 7 was shown to be significant. The origin of electric potentials across the gastrointestinal mucosa is uncertain. Active transport of ions, the permeability of mucosal and serosal surfaces of epithelial cells to ions, and the differences between intracellular and extracellular concentrations of ions all must be considered. 21 f gastric PD is determined largely by continuous secretion of chloride into the lumen,22 is to some degree sodium-dependent,23 and is greatest in parietal cells,16 the dif " 34.> '" 32 :l\ <:: ml "bile" given Time in 3 minute intervols FG. 4. Effect of bile on PD of human gastric mucosa. Mucosa negative to peripheral blood (mean ± SE, n = 6).

5 April 1970 GASTRC POTENTAL DFFERENCE r , >:- t:: -.!.... '" 40 Histalog t:: <::: X... 6 r j 5 4 i '<t 25 0 Time in 3 minute intervals FG. 5. PD of human gastric mucosa (mucosa negative to peripheral blood) during basal acid secretion and after maximal betazole stimulation (mean ± SE, n = 8). Output of HCl (mean ± SE) shown for each 5-min period. '". '" ference in PD between antrum and corpus is to a degree predictable. Mechanisms of ionic transport across duodenal and esophageal mucosae, and hence the origin of the PD in these regions, are unknown. Our studies further confirm the usefulness of PD in locating the gastroduodenal and gastroesophageal junctions, where the site of maximal change corresponds closely with the junctional zones The apparent failure of PD to correlate with gastric acid output accords with some previous observations. B, 24, 25 The small decrease in PD observed when gastric acid secretion is stimulated is believed to reflect a junction potential between the acid secreted and the sodium chloride already in the stomach. 25 The striking reductions in gastric PD, found after administration of aspirin, alcohol, or bile in quantities judged to be therapeutic, customary, or physiological, complement work (using other methods) describing physicochemical or morphological changes occasioned by these substances. These findings imply that maintenance of gastric PD is a sensitive indicator of mucosal integrity. PD in the canine stomach disappears after cyanide injection 26 and physical or chemical agents that damage the gastric mucosa cause a reduction in PD.20 Exfoliation of the canine gastric mucosal cells with eugenol results in simultaneous impairment of the gastric mucosal barrier to sodium and reduction in gastric PD, consistent with the belief that PD depends upon the maintenance of ionic gradients across the mucosa.! Davenport 2-4 also showed that aspirin, alcohol, and bile disrupted the gastric mucosal barrier to ionic diffusion in dogs, but measurements of PD were not made simultaneously. Adverse gross or histological effects of aspirin on the human gastric mucosa have been reported from gastroscopy, gastrectomy specimens, or mucosal biopsy27-32 and morphological changes in the gastric mucosa of animals receiving aspirin have been described. 33 Also, aspirin breaks the gastric mucosal barrier to sodium in man. 34 The mechanism by which aspirin produces morphological, chemical, and electrical damage to the gastric mucosa may be mediated by absorption of unionized acetylsalicylic acid in an acid medium. 2, 19 nhibition of mucus production by aspirin35 or exfoliation of gastric epithelial cells exceeding their replacement rate and thus leading to gastric erosions 36

6 442 GEALL ET AL. Vol. 58, No.4- is an explanation less consistent with the immediate decrease in PD that we found. Changes in morphology due to alcohol have been documented for more than a century,37 but the mechanism of alcohol damage to mucosal function, structure, and potential difference is uncertain. n animals, alcohol influences electrical activity in several ways, including depolarizing nerve membranes, lowering muscle membrane potentials, and inhibiting transport of cations and amino acids Contents of duodenal juice, particularly bile and pancreatic enzymes, also may produce histological changes in the gastric mucosa of animals and, in man, bile reflux into the stomach is abnormally great in patients with gastric ulcer The mechanism of damage is uncertain, but bile acids have been incriminated most directly Our results show that duodenal contents also cause striking reductions of potential difference in the healthy stomach. REFERENCES 1. Davenport, H. W., H. A. Warner, and C. F. Code Functional significance of gastric mucosal barrier to sodium. Gastroenterology 47: Davenport, H. W Gastric mucosal injury by fatty and acetylsalicylic acids. Gastroenterology 46: Davenport, H. W Ethanol damage to canine oxyntic glandular mucosa. Proc. Soc. Exp. Biol. M ed. 126: Davenport, H. W Destruction of the gastric mucosal barrier by detergents and urea. Gastroenterology 54: Code, C. F., J. A. Higgins, J. C. Moll, A. L. Orvis, and J. F. Scholer The influence of acid on the gastric absorption of water, sodium and potassium. J. Physiol. (London) 166: Ravin, 1. S., J. J. Kneisel, Jr., R. Taylor, H. M. Lemon, E. M. Thompson, and R. H. Smithwick Measurement of electropotentials of the stomach. Proceedings of the forum sessions of the thirty-sixth clinical congress of the American College of Surgeons, Boston, Massachusetts. Surg. Forum, 6!f-73 (October). 7. Andersson, S., and M.. Grossman Profile of ph, pressure, and potential difference at gastroduodenal junction in man. Gastroenterology 49: Rovelstad, R. A., C. A. Owen, Jr., and T. B. Magath Factors influencing the continuous recording of in situ ph of gastric and duodenal contents. Gastroenterology 20: 60!f Helm, W. J., J. F. Schlegel, C. F. Code, and W. H. J. Summerskill dentification of the gastroesophageal mucosal junction by transmucosal potential in healthy subjects and patients with hiatal hernia. Gastroenterology 4B: Meckeler, K. J. H., and F. J. ngelfinger Correlation of electric surface potentials, intraluminal pressures, and nature of tissue in the gastroesophageal junction of man. Gastroenterology 52: H ernandez, N. A., and 1. T. Beck Gastroesophageal transmural potential difference measured by a new constant infusion method. Amer. J. Dig. Dis. 14: Dennis, W. H., C. Canosa, and W. S. Rehm Potential difference across the pyloric antrum. Amer. J. Physiol. 197: Geall, M. G., D. C. Mclrath, S. F. Phillips, C. F. Code, and W. H. J. Summerskill Measurement of the transmucosal potential difference of stomach in unanesthetized man (abstr.). Gastroenterology 54: Rehm, W. S Evidence that major portion of gastric potential originates between submucosa and mucosa. Amer. J. Physiol. 147: 6!f Sawyer, P. N., J. E. Rhoads, and R. Panzer An evaluation of electrogastrography in the diagnosis of gastric cancer. Surgery 26: Villegas, L Cellular location of the electrical potential difference in frog gastric mucosa. Biochim. Biophys. Acta 64: Rehm, W. S A theory of the formation of HC by the stomach. Gastroenterology 14: Andersson, S., and M.. Grossman Effects of histalog and secretin on gastroduodenal profile of ph, potential difference, and pressure in man. Gastroenterology 51: Davenport, H. W Gastric mucosal hemorrhage in dogs. Effects of acid, aspirin, and alcohol. Gastroenterology 56: Rehm, W. S Positive injury potentials

7 April 1970 GASTRC POTENTAL DFFERENCE 443 of the stomach. Amer. J. Physiol. 140 : Schultz, S. G., and P. F. Curran ntestinal absorption of sodium chloride and water, p n C. F. Code and W. Heidel [eds.], Handbook of physiology. Sect. 6: Alimentary canal. Vol. : A critical, comprehensive presentation of physiological knowledge and concepts. American Physiological Society, Washington, D. C. 22. Hogben, C. A. M Active transport of chloride by isolated frog gastric epithelium: origin of the gastric mucosal potential. Amer. J. Physiol. 180: {i Cummins, J. T., and B. E. Vaughan onic relationships of the bioelectrogenic mechanism in isolated rat stomach. Biochim. Biophys. Acta 94: Crane, E. E., R. E. Davies, and N. M. Longmuir Relations between hydrochloric acid secretion and electrical phenomena in frog gastric mucosa. Biochem. J. 43: Rehm, W. S The effect of histamine and HC on gastric secretion and potential. Amer. J. Physiol. 141: Quigley, J. P., J. Barcroft, G. S. Adair, and E. N. Goodman The difference in potential across gastric membranes and certain factors modifying the potential. Amer. J. Physiol.119: Douthwaite, A. H., and G. A. M. Lintott Gastroscopic observation of effect of aspirin and certain other substances on stomach. Lancet 2: Paul, W. D Effect of acetylsalicylic acid (aspirin) on gastric mucosa: gastroscopic study. J. owa Med. Soc. 33: Alvarez, A. S., and W. H. J. Summerskill Gastrointestinal haemorrhage and salicylates. Lancet 2: Vickers, F. N., and M. M. Stanley Aspirin gastritis: gastroduodenoscopic observations. Four case reports. Gastroenterology 44: Muir, A., and 1. A. Cossar Aspirin and ulcer. Brit. M ed. J. 2: Spiro, H. M Stomach damage from aspirin, steroids, and antimetabolites. Amer. J. Dig. Dis. 7: Roth, W. L., A. Valdes-Dapena, P. Pieses, and E. Buchman Topical action of salicylates in gastrointestinal erosion and hemorrhage. Gastroenterology 44: Overholt, B. F., and H. M. Pollard Acetylsalicylic acid and ionic fluxes across the gastric mucosa of man. Gastroenterology 54: Hurley, J. W., and L. A. Crandall The effect of salicylates upon the stomachs of dogs. Gastroenterology 46: Croft, D. N Exfoliative cytology of the stomach after the administration of salicylates, p n A. St. J. Dixon, B. K. Martin, M. J. H. Smith, and P. H. N. Wood [eds.], Salicylates: an international symposium. Little, Brown & Company, Boston. 37. Beaumont, W Experiments and observations on the gastric juice and the physiology of digestion, p Dover Publications, nc., New York. 38. Gallego, A On the effect of ethylalcohol upon frog nerve. J. Cell. Compo Physiol. 31: Knutsson, E Effects of ethanol on the membrane potential and membrane resistance of frog muscle fibers. Acta Physiol. Scand. 52: Kalant, H., W. Mons, and M. A. Mahon Acute effects of ethanol on tissue electrolytes in the rat. Canad. J. Physiol. Pharmacol. 44: Chang, T., J. Lewis, and A. J. Glazko Effect of ethanol and other alcohols on the transport of amino acids and glucose by everted sacs of rat small intestine. Biochim. Biophys. Acta 135: Grant, R., M. 1. Grossman, K. J. Wang, and A. C. vy The cytolytic action of some gastrointestinal secretions and enzymes on.epithelial cells of the gastric and duodenal mucosa. J. Cell. Compo Physiol. 37: Lawson, H. H Effect of duodenal contents on the gastric mucosa under experimental conditions. Lancet 1: Capper, W. M Factors in the pathogenesis of gastric ulcer. Ann. Roy. Coll. Surg. Eng. 40 : Rhodes, J., D. E. Barnardo, S. F. Phillips, R. A. Rovelstad, and A. F. Hofmann ncreased reflux of bile into the stomach in patients with gastric ulcer. Gastroenterology 57: