Use of Pertechnetate Clearance in the Study of Gastric Physiology

Vo l. 63. No.1 (; ASTitO f: Nn :tW I.O(; Y

Print ed in U.S.A.

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USE OF PERTECHNETATE CLEARANCE IN THE STUDY OF GASTRIC PHYSIOLOGY ,JEHOME

G.

BICKEL, M.D. , THOMAS

A.

WITTEN, M . D ., AND MARY

K.

KILLIAN

Departm ent of M edicine, Division of Gastroenterology , Univ ersity of Colorado M edical Center, and Veterans Administration Hospital, Denver, Colorado

The gastric clearance of pertechnetate ( CTc) was compared with volume and acid output in 8 patients during basal, moderately stimulated (25 mg of betazole) and strongly stimulated (75 to 100 mg of betazole) states. Sodium pertechnetate solution was constantly perfused intravenously by means of a finger pump so that subjects received about 2 me in a 3-hr study period. Phenol red was constantly infused into the stomach as a marker substance and gastric contents were removed by continuous hand aspiration; volume corrections were made based on phenol red recovery; acid outputs were determined by titration. Gastric clearances of pertechnetate were computed from measured serum and gastric juice radioactivity. It was found that the factor of proportionality relating CT c to secretory output changes in relationship to the strength of stimulus employed. The same phenomenon was observed when the data of others relating aminopyrine clearance to secretory volume in dogs, and data relating iodide clearance to acid secretion in dogs, were subjected to similar computation. The results suggest that at a given level of stimulation, good proportionality exists between the clearances of pertechnetate, aminopyrine, and iodide and gastric secretory vigor. The active secretion of pertechnetate ion ( 99 mTc0 4 -) by gastric mucosa has interested several groups of investigators since the demonstration by Harper et al. 1 that

this ion is heavily concentrated during passage from blood to the gastric lumen and that its handling and distribution in the body are similar to those of iodide. These authors further pointed out the desirable characteristics of Technetium 9 9 m as an agent to be used in biologic studies in man. Its physical half-life is 6 hr, its biologic half-life 3 Vz to 4 hr, and its emission energy is an almost pure 'Y at 140 kev. Because it is distributed and handled in the body as if it were a halogen (in the form of pertechnetate) it is concentrated in the gastric juice, taken up by the thyroid, and rapidly excreted in saliva and urine. Harden et al. 2 employed pertechnetate to make scanning diagrams of the stomach. Irvine et al. 3 reported that the increase of gastric juice content of pertechnetate in the hour following a standard dose of histamine was proportional to the hydrogen

Received June 28, 1971. Accepted February 4, 1972. Address requests for reprints to : Dr. Thomas A. Witten, Veterans Administrat ion Hospital, 1055 Clermont Street, Denver, Colorado 80220. This investigation was supported in part by Veterans Administration Training Grant in Gastroenterology, TR 110. The authors gratefully acknowledge the assistance of Dr. Donald W. Brown, Chief of Nuclear Medicine Service, Veterans Administration Hospital, Denver, Colorado, who supplied the sodium pertechnetate used in this study ; Dr. Edward W. Moore, Gastroenterology Division, Medical College of Virginia, who offered valuable critical advice at an early phase of the study, and Dr. Philip G. Archer, Department of Biometrics, University of Colorado, for design of the statistical treatment of the data . 60

July 1972

PERTECHNETATE CLEARANCE IN GASTRIC PHYSIOLOGY

ion increase during that time. Meredith and Khan 4 measured the gastric clearance of pertechnetate in dogs using arteriovenous differences in the concentration of pertechnetate across isolated stomach vessels and compared the clearances so obtained with gastric blood flow rates measured with an electromagnetic flowmeter. They concluded that there was good agreement between the direct measurement of blood flow and the blood flow calculated from pertechnetate clearance. They did not measure secretory volume or hydrogen ion output. Jacobson et al. 5 measured gastric mucosal blood flow in dogs by means of aminopyrine clearance and related this value to secretory volume. They found good directional agreement between changes of volume secretion rate and blood flow so determined, but lack of proportionality between these variables. It is the purpose of this study to explore in man relationships between the gastric clearance of pertechnetate and hydrogen ion production by gastric mucosa. The clearance data of Meredith and Khan 4 suggest that pertechnetate clearance may provide an index of gastric mucosal blood flow and the observations of Jacobson et al. 5 suggest that an intimate, although incompletely understood, relationship pertains between mucosal blood flow and secretory activity. Patients Eight subjects (seven males and one female) were studied. One was a normal volunteer. The other 7 were patients of the Denver Veterans Administration Hospital or the Colorado General Hospital. Three had duodenal ulcer and 4 had complaints compatible with ulcer disease but without X-ray evidence of ulcer. Of these 4, 3 had normal gastric analyses and 1 had a high stimulated acid output.

Methods Subjects fasted at least 12 hr before the test. A double lumen nasogastric tube (constructed from a 16 French Levine type tube and a 7 French infant feeding tube with the tip of the smaller tube 15 em proximal to the nearest hole of the larger tube) was passed into the most dependent part of the stomach of the sitting sub-

61

ject. Its position was confirmed by fluoroscopy or by noting the completeness of return of instilled fluid. Phenol red solution (20 J.Lg per ml) was delivered to the stomach through the smaller of the two gastric lumens at about 1 ml per min. The precise rate of delivery of phenol red during each period was determined by reading a burette reservoir at the beginning and end of each period. A solution of Na 99 mTcO 4 in normal saline was infused intravenously through a constant rate pump which delivered 5 J.LC per min. At the beginning of the infusion, a loading dose of Na 99 mTc0 4 , 0.9 me, was injected intravenously. These doses were chosen after experience showed that they maintained a relatively steady blood level of pertechnetate activity. A waste period of 30 min following the introduction of pertechnetate and intubation was allowed for each subject. During this period, the stomach was emptied as completely as possible by hand aspiration with a syringe. Collection periods began at zero time. At least three base line collections of 10 to 15 min each were obtained by constant hand aspiration using a separate syringe for each period. After these periods, 25 mg of betazole were injected intramuscularly and collections were continued. The durations of the collection periods varied from 5 to 15 min, depending on the gastric juice flow rate . When gastric juice pH returned to base line values following this dose, an additional 75 to 100 mg of betazole were injected. Collections continued in .the same manner for an additional 60 min. The divided dose of betazole was designed to provide in each patient hydrogen ion output values as low, intermediate , and high secretory rates. During the procedure, patients were admonished against swallowing and reminded frequently to expectorate. Samples of venous blood were obtained every 20 min throughout the collection times for determination of radioactivity.

Measurements pH measurements were made immediately on each specimen of gastric aspirate usin g a Beckman expanded scale pH meter. Titratable acidity was determined by measuring the volume of 0.01 N sodium chloride required to raise the pH of a standard volume to 8.3. Sodium concentration was determined by standard flame photometry methods on an aliquot of each sample. Phenol red concentrations were

62

BICKEL ET AL.

measured on a Beckman D.B. spectrophotometer by the method of Hollander et al. 6 Serum was separated from each blood sample and counted in a scintillation well counter for radioactivity, the results being expressed as counts per minute per milliliter. One milliliter of each gastric aspirate was similarly assayed.

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99 mTc counts were expressed as minute out. puts. The nonparietal concentration of 99 mTc (TeN PI (NP, nonparietal) was assumed to be that of serum. The TeN P content was calculated and subtracted from the total 99 mTc output (Tcr) in each sample and the difference was called parietal pertechnetate output

(Tcp).

Computations Volume correction. Concentration phenolsulfonphthalein (PSP) in aspirate x volume of (1) aspirate = PSP, 11g per period, recovered Recovered PSP, 11g per period/ input PSP, 11g per period (2) = recovery fraction Recovered PSP, 11g per period/ concentration of input, 20 mg per ml (3) = contribution of PSP solution to volume of gastric aspirate recovered Volume of gastric aspirate recovered (4) - volume PSP solution recovered = volume gastric juice recovered Volume gastric juice recovered/ recovery fraction (5) = corrected volume gastric juice for period Parietal and nonparietal contributions. Using the assumptions of the two-component concept of gastric secretion as delineated by Makhlouf et al. 7 hydrogen ion concentrations were plotted against sodium concentrations of the gastric juice specimens and the intercepts of the regression line (H+I and (Na+l axes were noted. The (Na +l at (H+I = - 25 was taken as the sodium concentration of pure nonparietal secretion. The nonparietal sodium concentration was assumed to be constant. (The sodium ion concentration of each sample was divided by the derived nonparietal sodium concentration to give the fractional contribution of nonparietal secretion to the corrected volume of gastric juice.) The ••mTc activity (counts per minute per milliliter) of the various serum samples were plotted against time and the graphs constructed were used to determine the mean serum concentration of ••mTc during each collection period . The total gastric output of 99 mTc during each period was determined by applying the volume corrections described above. The corrected volumes of gastric juice and total

The total and parietal minute outputs of ••mTc divided by the mean serum concentrations give total and parietal pertechnetate clearance ( Grc and CTc ) in milliliters per minute. r P Specimens which showed obvious bile and those which yielded less than 80% or more than 120% recovery fraction were eliminated from the study. Seven specimens were rejected because of gross bile staining on the presumption that their titration values would be inordinately low; three were rejected because of poor fractional recoveries.

Results The absolute serum concentration of pertechnetate varied from patient to patient because the standard dosage was not adjusted to the individual body mass of the patient. The lowest mean serum level (2.12 x 10 4 counts per ml per min) occurred in a subject weighing 87 kg and the highest occurred (6.55 x 10 4 counts per ml per min) in a subject weighing 58 kg. For each experiment the maximum serum level deviation from the mean was not in excess of ±15%, but, more important, the differences between the means of adjacent serum level points and the values of the points which produce them did not exceed ±5%. It was shown that the plots of H + output against C Tc and eTc were insignificantly different: i.e., th~t their slopes, intercepts, and correlation coefficients were similar. The same was true of volume outputs ( Vr and Vp) plotted against CTc and CTcp respectively. The graphic repre: sentations are therefore of total minute outputs and total clearances only. Figure 1A shows the plot of total pertechnetate clearance ( CTc ) expressed as milliliters of serum per ~inute, against acid output expressed in milliequivalents

PERTECHNETATE CLEARANCE IN GASTRIC PHYSIOLOGY

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FIG. 1. A, H + output as a function of clearance of pertechnetate (CTch for the combined data (78 specimens). B, separate regressions computed for groups 1, 2, and 3. Progressively increasing slopes result as increasing levels of stimulation are applied. For definitions of groups, see text.

per minute. The correlation coefficient is 0.859. Only 50% of the points, however, lie between ± 15% of the hydrogen ion output predicted by this regression line; thus errors of estimates of acid output from CTc WOUld exceed these limits half Of the time. Attempts to demonstrate a pattern of curvilinearity by analysis of fragments of the regression line failed and the best fit to the plotted data points was considered to be a straight line. The increasing degree of divergence of

63

individual points from the over-all regression line at higher secretion rates in figure lA suggested that a changing, albeit still linear, relationship might obtain between H + and eTc under varying strengths of secretory stimulation. To explore the worth of this concept, the data was separated into three groups: Group 1 consisted of all specimens obtained before any secretory stimulus was given, to 1/z hr after a 25-mg dose of betazole. Group 2 consisted of all specimens obtained between Vz hr after the 25-mg dose of betazole and 1/z hr after the 75- to 100mg dose of betazole. Group 3 consisted of all specimens obtained more than 1/z hr after the 75- to 100mg dose of betazole. Figure lB shows the same plot of H+ output versus eTcr for these three groups. The group 1 (unstimulated) slope of 0.008 is half the value of the slope for moderately stimulated specimens (0.0164) and slightly more than one-third the value of the slope for strongly stimulated specimens (0.0234) . A Bartlett's test 8 on the three residual variances shows them to be significantly different (P < 0.005) and the slopes tested in pairs (i.e., 1 versus 2, and 2 versus 3) to be significantly different from one another (P < 0.005). Two-sided tolerance intervals 9 calculated separately for each slope for 90% of the population with a confidence coefficient of 95% yield the following relationships for estimating the hydrogen ion output from known pertechnetate clearance. Group 1

2

3

Formula H + = 0.00831 eTc ± 0.07 (95% confidence coefficient) H + = 0.01585 eTc ± 0.15 (95% confidence coefficient) H + = 0.02231 eTc ± 0.25 (95% confidence coefficient)

The relationships between H + output and pertechnetate clearance for the three groups are shown in figure lB. These ratios imply that: in the unstimulated State, a eTc Of 124 ml per min WOUld aSSO-

BICKEL ET AL.

64

ciate with an H + output of 1 mEq per min; in the moderately Stimulated State a eTc of 64 ml per min produces H + output of 1 mEq per min; in the heavily stimulated state Crc of only 48 ml per min is associated with H + output of 1 mEq per min. The relationship between minute volume and eTc UndergoeS a Change from a SlOpe of 0.0986 in the unstimulated group to about double that value for the two stimulated groups (table 1) . In the case of volume relationships, the linear regression line which relates VOlume OUtput to CTc for the combined data implies that an output of 1 ml per min is associated with CTc of 9.5 ml per min. Discussion These results show that the factors of proportionately relating H + and volume output to pertechnetate clearance undergo change when graded betazole stimulation is applied. The intercept at CTc = 7.5 ml per min of the regression line shown in figure 1A requires interpretation. We considered that this excess pertechnetate might be the product of salivary contamination of gastric contents. An alternative explanation might be that acid in an amount COrresponding tO CTc 7.5 ml per min (0.15 mEq per min) had indeed been secreted and had been reabsorbed by the mucosa. When, however, on separation of the data into groups representing the response to varying doses of betazole, it was found that all three regression lines intercepted 0, such considerations became less compelling. It remains important, nevertheless, to employ adequate means for excluding saliva from the gastric contents. 1. The ratios of volume output to total pertechnetate clearance under basal conditions (group 1) and at two different levels of betazole stimulation (for definitions of groups, see text)

TABLE

Vo lume: cT,(S lope)

All specimens .. Group 1 (unstimulated) ...... . Group 2 (moderate stimulation) . Group 3 (maximal stimulation) .

0.168 0.0986 0.195 0.183

0.920 0.84

0.90 0 .96

Vol . 63, No . I

The data of Jacobson et al., 5 relating volume output in the dog to the clearance of aminopyrine (cam)' show in their figure 3 a summary of observed relationships between volume output and C am. The least squares regression line that satisfies these data shows that for a 1.0 ml per min output a Cam of 31.6 ml per min would be re· quired. All of these data appear to have been obtained from animals receiving continuous stimulation with intravenous his· tamine (administration rates from 0.2 to 3.2 mg per hr) . The ratio between the aminopyrine and Tc clearance values associated with unit volume output per min is 31.6/9.5 = 3.32, a difference that could be accounted for by a difference in extraction ratios of the two indicator substances. Jacobson et al. 5 present a telling argument in support of the concept that the mucosal extraction of aminopyrine, under the conditions of their use, approximates 100%. The data of Meredith and Kahn 4 show that, in dogs, the gastric extraction ratio for pertechnetate is about 30%. If it were assumed that all of this extraction occurs at the mucosal level and that dogs resemble man in this extraction capability, the inference follows that in man the gastric mucosal blood flow would be estimated by multiplying the pertechnetate clearance by 3.3. The data depicted in figure 1B indicate that for a given CTc the output of acid varies with the intensity of stimulation. A changing ratio of secretory output to mu· cosal blood flow emerges from the data of Jacobson et al. when the volume output is used as a denominator and the Cam as a numerator at the various levels of histamine stimulation. The slopes in figure 2 are derived from measurements of Cam and volume output from figure 2 in Jacob· son et al. 5 using the mean of C am and of volume output for the second and third 15-min periods at each level of stimulation. Here, too, it is evident that for a given mucosal blood flow rate the secretory volume rate varies with the intensity of the stimulus. Figure 3, A and B, is constructed respectively from our pertechnetate clearance data and from the aminopyrine clear-

July 1972 .70

diluted (at higher secretion rates) by an increasing volume of iodine-poor secretion and that the two secretory processes were independent." Alternatively one might speculate that if iodide and pertechnetate clearance are proportional to mucosal blood flow, at a given rate of blood flow the H +-producing cell makes more or less use of this resource depending on the strength of the

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A ance data of Jacobson et al. In figure 3A the observed acid outputs shown in figure lB are related to betazole dosage in three separate curves representing pertechnetate clearances of 20, 40, and 60 ml per min. It is clear that under the stimulus of 25 or 100 mg of betazole, acid production is dependent on eTc. In figure 3B the data from Jacobson et al., shown in derived form in our figure 2, were treated similarly, using aminopyrine clearance levels of 5, 10, and 15 ml per min. It is evident that the two families of curves so generated are similar. They imply that for various rates of mucosal blood flow the effectiveness of the stimulant changes strikingly. Brown-Grant et al. 10 made similar observations of increasing ratios between H + output and gastric clearance of 131 I under increasing rates of administration of histamine. Their data can be arranged and plotted in a manner similar to the construction of figure 1B and figure 2 and such a plot again shows progressively steepening slopes with increasing doses of histamine. These authors offered as an explanation of this phenomenon the suggestion that "an iodide-rich secretion was being

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FIG. 3. A, the family of curves generated by plotting H + outputs as a function of betazole dosage at various levels of clearance of pertechnetate ( CTJ· The dotted curve for eTc 80 ml per min is hypothetical since clearance rates at this level were not observed. It indicates that in the system described, that given this eTc. the H + output would have been 0.58 mEq per min, 1.25 mEq per min, or 1.95 mEq per min, depending on whether the mucosa were responding to 0 Histalog, 25 mg of Histalog, or 100 mg of Histalog. B, the family of curves generated by plotting volume output as a function of histamine dosage at various levels of clearance of aminopyrine (C.ml (from data of Jacobson et al. ') .

66

BICKEL ET AL.

stimulation applied to it. Further consideration of this possibility must depend on experiments which would demonstrate directly the proportionality, between the clearances and measurements of mucosal blood flow. Whatever the mechanisms involved, pertechnetate clearance appears to provide a reasonably good estimate of H+ production when the level of stimulation is known, and to provide a much less precise estimate when that level is not known. Although we have not had an opportunity to examine a patient with a Zollinger-Ellison tumor by the method described, we might expect that such a patient, bearing an endogenous source of gastrin, would show a disproportionately low CTc in association with basal hypersecretion (i.e., would have points relating acid production rates during basal secretion to CTc somewhere along the group 2 and group 3 lines). REFERENCES 1. Harper PV, Andros G, Lathrop K: Semi-annual

report of the Argonne Cancer Research Hospital 18:76, 1962 2. Harden R MeG, Alexander WD, Kennedy I: Isotope uptake and scanning of the stomach in

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man with "mTc pertechnetate. Lancet 1:13051307, 1967 3. Irvine WJ, Stewart AG, McLoughlin GP, et al: Appraisal of the application of ••mTc in the as. sessment of gastric function . Lancet 2:648- 653, 1967 4. Meredith JH, Khan J: Gastric blood flow meas. urement of technetium clearance technic: an evaluation. Am Surg 33:969- 972, 1967 5. Jacobson ED, Linford RH, Grossman MI: Gas. tric secretion in relation to mucosal blood flow studied by a clearance technic. J Clin Invest 45: 1- 13, 1966 6. Hollander F, Penner A, Saltzman M: Determina· tion of phenol red in gastric contents. Proc Soc Exp Bioi Med 36:568-570, 1937 7. Makhlouf GM, McManus JPA, Card WI: A quan· titative statement of the two-component hy· pothesis of gastric secretion. Gastroenterology 51:149-171, 1966 8. Snedecor GW, Cochran WG: Statistical Meth· ods. Sixth edition. Ames, Iowa, The Iowa State University Press, 1967 9. Wallis AW: Tolerance limits for linear regression. Proceedings of the Second Berkeley Conference on Mathematical Statistics and Probability. Berkeley, California, University of California Press, 1951, p 43 10. Brown-Grant K, Cumming JD, Haigh AL, et al: The secretion of radioactive iodide by the stom· ach of the anesthetized dog in relation to total gastric blood flow and to acid secretion. J Physiol (Lond) 177:337-345, 1965