GASTROENTEROLOGY. Official Publication of the American Gastroenterological Association

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1 GASTROENTEROLOGY Official Publication of the American Gastroenterological Association COPYRIGHT 1970 THE WILLIAMS & WILKINS CO. VOLUME 58 March 1970 NUMBER 3 GASTRIC MUCOSAL BLOOD FLOW FOLLOWING DAMAGE BY ETHANOL, ACETIC ACID, OR ASPIRIN NEWELL A. AUGUR, JR., M.D. Department of Physiology, University of Michigan, Ann Arbor, Michigan The gastric mucosal blood flow in vagally denervated oxyntic gland area (Heidenhain) pouches was estimated by aminopyrine clearance in four 30-min periods after irrigations with acid solutions of ethanol, acetic acid, and aspirin. Mucosal blood flow increased initially when either ethanol or acetic acid was used and subsequently declined with appearance of evidence of gastric mucosal barrier damage. In spite of similar evidence for gastric mucosal barrier damage the mucosal blood flow with the aspirin solutions was, by the fourth period, significantly greater than the mucosal blood flow with the acetic acid solutions. Studies with acetic acid and aspirin solutions were repeated in vagally innervated oxyntic gland area (Pavlov) pouches. The pattern of mucosal blood flow was similar and in those experiments with homogeneous base lines the mucosal blood flow of the aspirin studies in the fourth period was again significantly greater than that of the acetic acid study. Although a greater mucosal blood flow might be considered a beneficial and protective influence, invariable bleeding characterized the fourth period of the aspirin experiments while only inconstant and minimal bleeding was noted in the fourth period of the acetic acid experiments. The normal gastric mucosa forms a nearly impermeable barrier to the diffusion of sodium and hydrogen ions. This barrier, by preventing the diffusion of H + from Received June 20, Accepted September 16,1969. Address requests for reprints to: Dr. Newell A. Augur, Jr., Department of Physiology, University of Michigan, Ann Arbor, Michigan This work was supported in part by Grant AM from the National Institutes of Health, United States Public Health Service. Dr. Augur also received Training Grant from the National Institutes of Health. The author is indebted to Dr. Horace W. Davenport for his encouragement and advice. 311 the gastric lumen through the mucosa, maintains the concentration of secreted acid and protects the stomach from injurying itself by its own secretions. If this barrier is disrupted by noxious substances, back diffusion of H + and a large outpouring of Na+ occur. Bleeding frequently follows. In man as well as in dogs numerous chemicals and drugs have been proven or are strongly suspected to be causes of the induction of this acute gastric injury.1-6 Davenport has proposed7 that the increased permeability of the mucosal barrier after injury permits rapid diffusion of

2 312 AUGUR Vol. 58, No.8 acid into' the muco'sa. There the acid liberates histamine, which stimulates acid secretio'n and increases the permeability O'f muco'sal capillaries. The acid diffusing into' the muco'sa also' ruptures capillaries, pro' ducing interstitial hemo'rrhage and bleeding into' the lumen. The central features O'f this co'ncept are rapid back diffusio'n O'f acid and release O'f histamine. The co'mbined effect O'f these two' influences O'n the gastric muco'sal blo'o'd flo'w is unkno'wn. This paper repo'rts an attempt to' measure the respo'nse O'f the gastric muco'sal blo'o'd flo'w after damaging the gastric muco'sal barrier. The metho'd used to' measure gastric muco'sal blo'o'd flo'w was the gastric muco'sal clearance O'f amino'pyrine.8 The technique O'f this measurement avo'ids the difficulties O'f surgical trauma, sho'ck, and anesthesia and permits repeated study O'f the same animals. The gastric muco'sal clearance O'f amino'pyrine was measured after damaging the gastric muco'sal barrier with acid so'lutio'ns O'f ethano'l, acetic acid and aspirin. ' Materials and Methods Part I Three dogs with vagally denervated oxyntic gland area (Heidenhain) pouches were used. The animals were healthy and accustomed to the procedure and were fasted for at least 20 hr before each experiment. Hematocrit (Wintrobe tube) determinations were performed frequently. Initially the hematocrit of each dog was about 50%, but it subsequently fell and fluctuated from 38 to 48%. Neigher profound anemia nor manifestations of agranulocytosis developed. Experiments were carried out at intervals of no less than 1 week. The details of an experiment in which aminopyrine was infused have been described, as has the method of irrigating the pouches:' s. 7 Briefly, an experiment consisted of placing a dog in a supporting sling, filling the pouch with a solution of N aci and beginning an infusion of aminopyrine 'in saline through a hind leg vein. A loading dose of 20 mg per kg was given in 10 min and was followed by a constant maintenance infusion of 5 mg per kg per hr. Twenty minutes after the start of this maintenance dose the first of four 30-min periods was begun. The pouch was rinsed twice, gently aspirated, and filled with 40 ml of a solution of 100 mn HCI and 54 mn NaCI. Approximately 9.5 ml were removed, measured, and saved and the time was noted (zero time). At both the beginning and the end of a period, aspiration did not completely empty the pouch. The small residual or dead space volume has been estimated in each dog by a phenolsulfonphthalein dilution techniques.. and this volume is considered in the calculation (vide infra). At 15 min, 6 to 8 ml of venous blood were drawn from a front leg vein and centrifuged and the plasma was saved. At 30 min (the completion of the first or base line period), the fluid in the pouch was aspirated, measured, and saved. This procedure was repeated in identical manner for three subsequent periods with the exception that the pouch was filled with a test solution in the second period. The solution of 100 fin HCl and 54 mn N aci again was used in the third and fourth periods as in the first. The three test solutions were (1) 9% (w/v) ethanol in 100 mn HCl and 54 mn NaCl, (2) 100 mm acetic acid and 54 mn NaCl, and (3) 20 IIlM aspirin (acetylsalicylic acid) in 100 fin HCl and 54 fin N aci. Each solution was tested in each dog three or four times. To compare our technique with that of Jacobson et al.," at least one experiment was done on each dog in which, after the completion of the first period, histamine (50 J-tg per kg-hr) was infused with aminopyrine. This infusion was continued for three subsequent 30-min periods, during each of which the pouch was filled with the solution consisting of 100 mn HCI and 54 mn N aci. To establish whether the gastric mucosa damaged by the test solutions was permeable to aminopyrine, two controls were carried out with each of the test solutions on each dog. The solution with which the pouch was filled in all control periods contained aminopyrine (150 to 200 J-tg per ml), a concentration nearly equal to the greatest concentration found at the end of any 30-min period during an experiment. For the control animals, the test and two subsequent periods were preceded by two base line periods. Total acidity of the initial and final solutions from each period was measured by titration of a 1-ml sample with 0.1 N NaOH to the phenolphthalein end point. The net fluxes of H+ across the mucosa per 30-min period were determined by subtracting the initial from the final H+ mass. A positive net flux of H+ means a gain of H+ in the pouch lumen and, although evidence for active acid secretion, indicates only

3 March 1970 GASTRIC MUCOSAL BLOOD FLOW 313 the minimal amount that may have been secreted. A large negative net H+ flux means a loss of H+ from the pouch lumen and indicates that the gastric mucosal barrier is damaged. One cannot infer, however, from the negative flux that there was no H+ secreted. Acid secretion may have occurred and have been masked by the loss of H+ diffusing across the damaged gastric mucosal barrier. The hemoglobin concentration of the final solution from each period was estimated by comparison with samples of hemoglobin substrate powder of known concentration. These standards were made by adding hemoglobin substrate powder to 0.1 N HC!. The pouch aspirate was compared with these standards against a white background and the concentration was recorded to the nearest multiple of 5 mg per 100 m!. When the hemoglobin concentration was less than 40 mg per 100 ml, one easily distinguished concentration changes of less than 5 mg per 100 m!. The results are expressed in milligrams of hemoglobin per 100 milliliters. (If all of the red blood cells in a solution with a hematocrit of 1 % were lysed completely, the solution would then have a hemoglobin concentration of 333 mg per 100 m!.) The concentration of aminopyrine in each pouch and plasma sample was determined by the method of Brodie and Axelrod.' A I to 2-ml sample was alkalinized, extracted into ethylene dichloride, washed twice with sodium borate, re-extracted into an acid solution, and measured at 260 mp. on a Beckman double beam spectrophotometer. An aminopyrine solution standard with a concentration of 100 p.g per ml gave an absorbance reading of Neither aspirin, acetic acid, ethanol, nor hemoglobin interfered with this determination. The plasma aminopyrine clearance (PAC) was calculated for each period from the equation: Cl = (Gao X Vao) - (Go X Vo) P X 30 where Cl is the PAC in milliliters per minute, Go the intial concentration of aminopyrine in the pouch, Gao the concentration at the end of the period, Vo the initial volume of the gastric pouch (40 ml plus the residual volume and minus the amount aspirated, usually about 9.5 ml), Vao the final volume the amount aspirated plus the residual volume), and P the plasma aminopyrine concentration. The sample mean and sample standard deviation of the PAC's were calculated for each period. For each dog the mean PAC of each period of one test solution was compared with the mean PAC of the same period for the other two test solutions. Comparisons between periods were made by the Student's t-test. Part II Two dogs with vagally innervated oxyntic gland area (Pavlov) pouches were used. The intact vagal innervation was demonstrated by a 2-fold or greater increase in the acid secreted. per 15-min period following an intravenous injection of regular insulin (1.2 U per kg). Dog G's pouch was small and was filled with only 30 ml of the appropriate solution rather than the 40 ml as used with the other dogs. The details of the experimental procedure otherwise have been described in part 1. Before all experiments had been completed, dog G began refusing food and lost weight; she was not used again. The two test solutions were (1) 100 mm acetic acid and 54 mn NaCl and (2) 20 mm aspirin (acetylsalicylic acid) in 100 mn HCl and 54 mn NaC!. Two controls were carried out with each test solution on each dog. Aminopyrine, H+, and hemoglobin determinations were performed as indicated previously. In addition, Na+ concentration in the initial and final solutions from each period was measured by an internal standard flame photometer. The net Na+ fluxes per 30-min period were calculated by subtracting the initial from the final Na+ mass. The net flux of Na+ was almost invariably positive, indicating a net movement of Na+ into the pouch lumen. After introduction of the test solutions positive net fluxes of Na+ became larger, indicating that the gastric mucosal barrier had been broken.' Results Part I Comparison of technique. During maximal histamine stimulation, Jacobson et al.8 observed a minimal range of the ratio of gastric aminopyrine concentration to plasma aminopyrine concentration of 25 to 35. A maximal plasma aminopyrine clearance of 18 ml per min was recorded. In our dogs the minimal range of the ratio was 25 to 38, and the range of maximal plasma aminopyrine clearance was 21.2 to 30.3 ml per min. This comparison indicates satisfactory reproduction of their technique.

4 314 AUGUR Vol. 58, No. S TABLE 1. Percentage of aminopyrine recovered from oxyntic gland area pouches after five SO-min periods of irrigationa Vagally denervated pouches (3 dogs) Vagally innervated pouches (2 dogs) Test solution Nb Period Percentage aminopyrine Nb Period Percentage aminopyrine recovered recovered N aci and ± and ± 3.4 9% (w Iv) ethanol in 100 mn HCI and 54 mn N aci used in period ± 3.7 N aci ± 3.5 N aci ± mm acetic acid and 54 mn N aci used in period ± ± 7.6 N aci ± ± 1.9 N aci ± ± mm aspirin in 100 mn HCI and 54 mn N aci used in period ± ± 3.6 N aci ± ± 2.4 N aci ± ± 4.5 a Values are means ± SD of two experiments done with each test solution. The aminopyrine concentration was either 150 or 200 p.g per ml in all solutions. b N, number of observations. Control animals. The percentage of aminopyrine recovered in five consecutive 30-min periods is presented in table 1. Two experiments were done with each of the three test solutions in each of the 3 dogs. The data of base line periods from all tests were pooled and were expressed as one result. The figures show an over-all aminopyrine recovery of 96 to 101 % and were interpreted as indicating that the loss of aminopyrine through the damaged gastric mucosa of the vagally denervated oxyntic gland area pouch was negligible. Experiments. The mean net volume and H + changes, hemoglobin concentration, and plasma aminopyrine clearances for dogs C, F, and H are presented in table 2. The data of the base line periods for each dog were pooled and were expressed as one result. Second period (test solution). In each dog the highest mean PAC was found in the presence of the ethanol solution. In all 3 dogs this mean PAC was significantly greater than the mean PAC found with the aspirin solution, but not greater than the mean PAC of the acetic acid solution. The mean PAC of the acetic acid solution was significantly greater than the mean PAC of the aspirin solution in 2 of 3 dogs. The expected stimulation of acid secretion by ethanol with a net gain of H + in the pouch ll and a similarly expected net loss of H + with both acetic acid and aspirin solutions 2 were observed. Third period (follow-up 1). (To avoid awkwardness, those experiments in which ethanol was used henceforth are called the ethanol test or experiment, those experiments with acetic acid, the acetic acid tests, and so forth. The specific test solution, however, was used only in the second period.) The mean PAC of the ethanol tests was significantly greater than the mean PAC of the acetic acid tests in 2 of 3 dogs. There were no significant differences between the mean PAC of the aspirin and the mean PAC of the acetic acid tests. A positive net flux of H + in the ethanol tests and negative net fluxes of H + in the acetic acid and aspirin experiments again oc-

5 March 1970 GASTRIC MUCOSAL BLOOD FLOW 315 TABLE 2. Plasma aminopyrine clearance and net H+ and volume changes in vagauy denervated oxyntic gland area pouches of 3 dogs after four 30-min periods of irrigationa Solution Nb Period Net Net H+ Hbc Significant PACd volume differences DogC NaCI ± % (w/v) ethanol in NaCI ± 1.24 P < 0.05' NaCI ± 0.28 P < 0.05' P < O.05t NaCI ± 0.80 P < O.Olt 100 DlM acetic acid and 54 mn NaCI ±2.39 NaCl ± 1.58 NaCI ± mm aspirin in N aci ± 2.00 NaCI ± 1.50 NaCI ± 0.31 P < O.025t DogF NaCI ± % (w/v) ethanol in NaC! S ± 1.29 P < 0.05' NaCI ± 0.91 P < 0.025t NaCI O-trace ± 2.51 P < 0.10t 100 mm acetic acid and 54 mn NaCI ± 0.57 P < 0.05' NaCI O-trace 7.71 ± 1.17 NaCI ± mm aspirin in NaCI ± 2.25 NaCI ± NaC! ± 0.60 P < O.05t DogH NaCI ± % (w/v) ethanol in NaC! ± 1.64 P < 0.005' NaCI O-trace 8.08 ± 0.87 NaCI ± mm acetic acid and 54 mn NaC! ± 2.19 P < 0.05' NaC! ± 2.38 NaC! ± mm aspirin in NaC! ± 0.73 NaCI ± 2.08 NaC! ± 1.01 P < O.lOt a Mean net changes of volume in milliliters and of H+ in microequivalents per 30 min. A negative sign denotes 1088 of contents from the pouch. b Number of observations. C Range of hemoglobin concentration at end of 30-min period in milligrams per 100 milliliters. d PAC, plasma aminopyrine clearance in milliliters per minute, mean ± so. PAC of each period of a test solution compared with PAC of same period of other two test solutions. " mean PAC of this period is significantly greater than the mean PAC of same period of aspirin study. t, mean PAC of this period is significantly greater than mean PAC of same period of acetic acid study. curred. Some bleeding was noted in the dogs tested with aspirin. Fourth period (follow-up 2). The characteristic feature of this period was that the mean PAC of the aspirin experiments was greater than the mean PAC of the acetic acid test in all 3 dogs. This difference was significant at probability levels of P < (dog C), P < 0.05 (dog F), and P < 0.10 (dog H). Continued net loss of H + from the pouch occurred in the acetic acid and aspirin experiments. For the first time a negative net H + flux was observed in the ethanol experiments. This negative net flux and the bleeding noted in the pouch fluid of 2 of 3 dogs indicate that this 9% (w/v) ethanol solution can break the mucosal barrier in dogs. Although the amount of bleeding was variable in the dogs, each dog's response

6 "" ~ TABLE 3. Plasma aminopyrine clearances and net electrolyte changes in vagally innervated oxyntic gland area pouches of 2 dogs after four 30-min periods of irrigationa Solution Period Experiment 1 Experiment 2 Experiment 3 H+ Na+ PACb H+ Na+ PACb H+ Na+ PACb DogD 100 mn HCl and 54 mn NaCI mm acetic acid and 54 mn N aci mn HCl and 54 mn N aci mn HCl and 54 mn NaC! Experiment 4 Experiment 5 H+ Na+ PACb H+ Na+ PACb NaCI mm aspirin in 100 mn HCI and 54 mn NaC! mn HCl and 54 mn NaCI mn HCl and 54 mn N aci DogG 100 mn HCl and 54 mn N aci mm acetic acid and 54 mn N aci mn HCl and 54 mn N aci mn HCl and 54 mn N aci mn HCl and 54 mn N aci mm aspirin in 100 mn HCI and 54 mn NaCl N ac! mn HCl and 54 mn N aci a Electrolyte changes in microequivalents per 30 min. A negative sign denotes loss of contents from the pouch. b PAC, plasma aminopyrine clearance in milliliters per minute.

7 March 1970 GASTRIC MUCOSAL BLOOD FLOW 317 was remarkably consistent. Dog C bled in the fourth period in each of three aspirin experiments. She did not bleed in any period of the acetic acid or ethanol tests. By contrast, dog F's mucosa appeared more fragile as there was mild bleeding in thefouth period of two acetic acid experiments and severe bleeding in all three aspirin experiments. Bleeding was so severe in her pouch by the end of the third period in the first aspirin experiment that the procedure was discontinued. Dog H bled minimally in one ethanol test and regularly in the third and fourth periods of all the aspirin experiments. The characteristic features of these aspirin experi- TABLE 4. Plasma aminopyrine clearance and net electrolyte and volume changes of experiments with satisfactory base lines in vagally innervated oxyntic gland area pouches of 2 dogsa Solution Nb Period Net NetH+ Net Hbc PAC" Significant volume Na+ differences' DogD NaC! ± mm acetic acid and 54 mn NaC! ± 1.77 NaC! ± 1.59 NaC! ± mm aspirin in 100 mn HCI and 54 mn N aci trace ± 0.55 NaC! ± 0.31 NaC! ± 0.53 P < 0.025t DogG NaC! ± mm acetic acid and 54 mn NaC! ± 1.17 NaC! ± 0.77 NaC! ± mm aspirin in 100 mn HCI and 54 mn N aci NaC! trace NaC! a The experimental data from which this group are drawn has been presented in table 3. The selected experiments were for dog D, acetic acid tests 1, 2, and 4 and aspirin tests 1,4, and 5 and for dog G, acetic acid tests 1, 2, and 3 and aspirin test 1. Mean net changes of volume in milliliters and of electrolytes in microequivalents per 30 min. A negative sign denotes loss of contents from the pouch. b N, number of observations. c Range of hemoglobin concentration at end of 30-min period in milligrams per 100 milliliters. d PAC, plasma aminopyrine clearance in milliliters per minute, mean ± SD. 'PAC of each period of acetic acid study compared with PAC of same period of aspirin study. t, mean PAC of this period was significantly greater than mean PAC of same period in acetic acid study.

8 318 AUGUR Vul. 58, No.3 ments were that the dogs invariably bled, that this bleeding increased during the experiment, and that the bleeding was always greater than any that occurred with acetic acid or ethanol. Part II Controls. Table 1 shows an over-all aminopyrine recovery of 99.6 to 104.8%. These results were interpreted as indicating that the loss of aminopyrine through the damaged gastric mucosa of the vagally innervated oxyntic gland area pouch was negligible. Experiments. All experiments for the 2 dogs are presented in table 3. It was hoped that in the first or base line period of each experiment the dog's pouch would be in the resting state with a low PAC and minimal H + secretion. Marked variation in these base line measurements, however, was observed. Dog D, for instance, had in the first period a PAC range from 4.12 to 9.96 ml per min and a net H + fiux range from -36 to 319 p,eq per 30-min period. Comparisons of the results of these experiments with heterogeneous base lines were without significant differences. As the physiological state of the stomach determines in part its response to drugs, it seemed that more meaningful comparisons could be made between groups of experiments with more homogeneous base line periods as had been planned originally. Experiments where the dog's pouch was in the resting state (low PAC and minimal H+ secretion) during the first period were identified. Three experiments with satisfactory base lines for each test solution were obtained for dog D (acetic acid tests 1, 2, and 4 and aspirin tests 1, 4, and 5) and for the acetic acid solution for dog G (acetic acid tests 1, 2, and 3). Unfortunately only one acceptable experiment with the aspirin solution was obtained in dog G (aspirin test 1) before she stopped eating. Comparison of selected groups. The data from the selected experiments with satisfactory base lines are presented in table 4. For dog D the comparison indicates that the mean PAC of the fourth period of the aspirin test was significantly greater than the mean PAC of the fourth period of the acetic acid test. There were no other significant differences between mean PAC's. For dog G the acetic acid results are complete and mimic those of dog D. The one satisfactory aspirin test did show an increase in mean PAC from the first through the fourth period. As the data are incomplete for dog G, tests of significance were not applied. Bleeding invariably occurred in the aspirin tests and progressively increased through the fourth period. By contrast, bleeding was seen infrequently with the acetic acid tests, was minimal, and when present was not necessarily worst in the final period. The positive net N a + fiux and the negative net H + were both larger in the acetic acid tests than in the aspirin tests. Discussion The aminopyrine clearance technique is based on the work of Shore et aj.12 and Jacobson et al. 8 This clearance would reflect gastric mucosal blood flow if the following assumptions were made: (1) that aminopyrine is equally distributed between plasma and red blood cells, (2) that there is rapid equilibration of aminopyrine between plasma and red blood cells, (3) that aminopyrine is cleared completely from the blood by the gastric mucosa in one passage, and (4) that aminopyrine is not secreted actively by the stomach. With regard to the first assumption, plasma aminopyrine concentrations have been shown to equal whole blood concentrations and to be independent of drug binding to hemoglobin. 13 Determination of plasma aminopyrine concentration is more accurate than measurement of aminopyrine in whole blood and therefore is more representative of whole blood aminopyrine concentration. Assumptions 2, 3, and 4 have not been proven conclusively to be true, but there are sound reasons to accept them as valid Although conceivable, it seems unlikely that ethanol, acetic acid, or aspirin interferes with diffusion of aminopyrine from plasma to gastric pouch lumen. The results reported here as plasma

9 March 1970 GASTRIC MUCOSAL BLOOD FLOW 319 aminopyrine clearance are assumed for all practical purposes to reflect accurately the gastric mucosal blood flow. Alterations of gastric mucosal blood flow in response to injury induced by solutions of ethanol, acetic acid, and aspirin were determined in vagally denervated pouches. The three test solutions provoked large net negative H+ fluxes, large net positive Na+ fluxes, or bleeding, indicating that the gastric mucosal barrier had been damaged. These results are similar to extensive previous studies with ethanol, acetic acid, and aspirin The degree and direction of change of the mucosal blood flow varied for these solutions. In the ethanol experiments the second or test period was characterized by an increase in acid secretion and in mucosal blood flow. A change in the gastric acid secretory rate obligates a directionally corresponding change in mucosal blood flow. s As the ethanol had stimulated acid secretion in this test period, the observed increase in the mucosal blood flow is appropriate. A small additional contribution may have been the influence of absorbed ethanol. 11 The circulating ethanol, by depressing the central nervous system vasomotor center, might have vasodilated the mucosal vessels, reduced peripheral resistance, and thereby contributed to the augmented flow. The mucosal blood flow of the second period of the acetic acid study also increased, although in this period there was evidence of gastric mucosal barrier damage. As experiments in which secreted H + is trapped by glycine have shown only minimal acid secretion after irrigation with acetic acid,6 this increase in mucosal blood flow cannot be explained reasonably as a requirement of increased acid secretion as with ethanol. There is, however, a rapid release of histamine with peak concentrations in the gastric venous blood 5 min after exposure of the entire gastric mucosa to acetic acid. 15 Thus, histamine is present and could be exerting an influence that increases the gastric mucosal blood flow. As the ethanol and acetic acid experiments proceeded, evidence of gastric mucosal barrier damage. appeared and the mucosal blood flow declined. This decline was more precipitous in the acetic acid study where the net negative loss of H + from the lumen was larger than that in the ethanol experiments. This suggests that the decreasing mucosal blood flow is related causally to the back diffusion of H +. This is consistent with the observation that diffusion of H + into guinea pig mucosa induces vasoconstriction and stasis.16 One may envisage in the healthy stomach a gastric mucosal blood flow delivering adequate amounts of buffers to neutralize the few H + diffusing back across the gastric mucosal barrier. After damage to this barrier the marked increase in the back diffusion of H + overwhelms these buffers and the effects of the unbuffered H + on mucosal vessels decrease the mucosal blood flow. An unexpected discovery of this work was that in the fourth period of the aspirin studies the mucosal blood flow was significantly greater than that in the fourth period of the acetic acid experiments. As this discrepant response was unexplained and as a vagally stimulated cholinergic vasodilator mechanism has been recognized in the gastric vascular bed,17 the experiments with the acetic acid and aspirin solutions were repeated in dogs with vagally innervated pouches. In these studies (part II) the response of the mucosal blood flow with either solution was similar to the response for that same solution in dogs with vagally denervated pouches. Furthermore, in a group of experiments with homogeneous base lines the gastric mucosal blood flow in the fourth period of the aspirin study was again significantly greater than that of the acetic acid study. It is concluded that the changes of the mucosal blood flow after injury induced with acetic acid and aspirin are essentially independent of intact vagal innervation. Similar magnitudes of negative net H + flux occurred in both the acetic acid and aspirin experiments. If back diffusion of H+ after injury is related causally to decreasing mucosal blood flow as it seems to have been in the acetic acid and ethanol experiments, then some mechanism is pre-

10 320 AUGUR Vol. 58, No.3 venting the expected decrease of mucosal blood flow in the aspirin experiments. Part II of this work indicates that this effect is not mediated by the vagus nerve. After gastric mucosal injury with concentrations of acetic acid and salicylic acid identical with those used in these experiments, histamine release into an isolated gastric vein was similar in timing and peak con centrations.15 This suggests that the mechanism preventing the expected decrease of gastric mucosal blood flow in the aspirin experiment is not operating through a change in release of histamine. Other vasoactive substances may be liberated from the damaged gastric mucosa. In particular, the kinins are intriguing as they act in the pathogenesis of inflammation and aspirin may inhibit their formation or action. 18 Finally, one might expect greater mucosal blood flow to be a beneficial and protective influence by preventing hypoxia and by diluting, neutralizing, and rapidly removing noxious agents, e.g., H+ and aspirin, diffusing across the damaged gastric mucosal barrier. Although the final period of the aspirin studies had a significantly greater mucosal blood flow than the final period of the acetic acid study, the former was characterized, paradoxically, by invariable bleeding and the latter by inconstant and minimal bleeding. What this peculiar toxic property of aspirin is and whether it is also responsible for the response of the mucosal blood flow remain unknown. REFERENCES 1. Davenport, H. W., J. A. Warner, and C. F. Code Functional significance of the gastric mucosal barrier to sodium. Gastroenterology 47: '. Davenport, H. W Gastric mucosal injury by fatty and acetylsalicylic acids. Gastroenterology 46: Davenport, H. W Damage to the gastric mucosa: effects of salicylates and stimulation. Gastroenterology 49: Davenport, H. W Destruction of the gastric mucosal barrier by detergents and urea. Gastroenterology 54: Davenport, H. W Gastric mucosal hemorrhage in dogs. Effects of acid, aspirin, and alcohol. Gastroenterology 56: Schoen, A. M Acute gastritis, p In H. L. Bockus [ed.], Gastroenterology, Ed. 2, Vol. 1. W. B. Saunders Company, Philadelphia. 7. Davenport, H. W Fluid produced by the gastric mucosa during damage by acetic and salicylic acids. Gastroenterology 50: Jacobson, E. D., R. H. Linford, and M. 1. Grossman Gastric secretion in relation to mucosal flow studied by a clearance technic. J. Clin. Invest. 45: Hollander, F Determination of phenol red in gastric contents. Proc. Soc. Exp. Bioi. Med. 36: ro. Brodie, B. B., and J. Axelrod The fate of aminopyrine (Pyramidon) in man and methods for the estimation of aminopyrine and its metabolites in biological material. J. Pharmacol. Exp. Ther. 99: Davenport, H. W Ethanol damage to canine oxyntic glandular mucosa. Proc. Soc. Exp. Bioi. M ed. 126: Shore, P. A., B. B. Brodie, and C. A. M. Hogben The gastric secretion or drugs: a ph partition hypothesis. J. Pharmacol. Exp. Ther. 119: Harper, A. A., J. D. Reed, and J. R. Smy Gastric blood flow in anaesthetized cats. J. Physiol. (London) 194: Jacobson, E. D Progress in gastroenterology. Clearances of the gastric mucosa. Gastroenterology 54: Johnson, L. R., and B. F. Overholt Release of histamine into gastric venous blood following injury by acetic or salicylic acid. Gastroenterology 52: Watt, J The mechanism of histamine ulceration in the guinea pig. Gastroenterology 37: Naitove, A., and E. Colby, Evidence for a vagally stimulated cholinergic vasodilator mechanism in the gastric vascular bed. Gastroenterology 48: Kellermeyer, R. W., and R. C. Graham, Jr Kinins-possible physiologic and pathologic roles in man. New Eng. J. Med. 279: ,