- Email: [email protected]
Somatostatin and the interdigestive migrating motor complex in man
Regulatory Peptides, 5 (1983) 209-217
209
Elsevier BiomedicalPress
Somatostatin and the interdigestive migrating motor complex in man T.L. Peeters, J. Janssens and G.R. Vantrappen Division of Gastroenterology, Department of Medical Research, Catholic University of Leuven, Gasthuisberg, B- 3000 Leuven, Belgium
(Received27 September1982; accepted for publication 3 December 1982) Summary The relationship between somatostatin and the interdigestive migrating motility complex (MMC) was determined in human volunteers. Motor activity was monitored manometrically by means of seven perfused catheters: four in the stomach, one in the duodenum, two in the jejunum. Blood samples were drawn every 10 min and radioimmunoassayed for motilin, pancreatic polypeptide and somatostatin. In four volunteers two activity fronts (AF) were recorded and somatostatin levels correlated to the manometric data. The start of an A F in the upper duodenum was accompanied by somatostatin peaks. Peak values, taken as the mean of the levels in the sample obtained after the start of an AF, the preceding sample and the next one, averaged 32 + 4 pM compared to 12 + 2 pM in the remaining period. In four volunteers somatostatin was infused in doses of 1.2, 2.4 and 4.8 p M / k g per min over three consecutive periods of 90 min, causing dose-dependent increments in plasma somatostatin levels of 7, 32 and 76 pM. In all volunteers and for all doses all gastric activity was completely inhibited. In the intestine phase 2 was abolished but phase 3 stimulated: during somatostatin infusion phase 3 occurred with an interval of 39 _ 6 min. Motilin and PP levels were decreased. As the two lowest infusion doses caused increases in somatostatin levels that might be considered as physiological, somatostatin seems to have a physiological role in the regulation of the migrating motor complex. We propose that it facilitates the progressing of the activity front into the small intestine. intestinal motility; gastric motility; interdigestive motility; motilin; pancreatic polypeptide
Introduction
The interdigestive migrating motor complex (MMC) is a typical motility pattern of the small intestine in the fasting state, consisting essentially of a rest period 0167-0115/83/0000-0000/$03.00 © 1983 ElsevierBiomedicalPress
210 (phase 1), a period of irregular contractions (phase 2), a period of rhythmic contractions occurring at maximal frequency (phase 3) and a period of rapid decline in activity (phase 4). All phases progress aborally, and occur cyclically with a period of about 92 min in man [1]. Several gastrointestinal hormones have been shown to be related to the MMC. From a recent review on this subject, it can be concluded that there is evidence for the involvement of motilin in the initiation of phase 3 in the stomach. Infusion of cholecystokinin, gastrin, glucagon, insulin and secretin disrupts the interdigestive pattern and produces a fed pattern [2]. Somatostatin, originally discovered as the growth hormone release inhibiting factor [3;4], has also been found to influence gastrointestinal function. It is generally accepted that it has a powerful inhibitory effect on the release of gastrointestinal hormones and on gastric and pancreatic secretion [5]. However, its influence on gastrointestinal motility is controversial: depending upon species, organ, dose and various other factors, different and sometimes conflicting results have been reported. In anesthetized dogs, Boden et al. [6] did not observe an effect on gastric antral motility, but Tansy et al. [7] noted a brief inhibitory effect. In man, inhibitory effects [8] on gastric motor function and an initial increase followed by a decrease of the gastric emptying rate [9] were observed. In monkeys, somatostatin had no effect on antral motility [10]. In the small intestine induction [11] as well as inhibition [12-14] of the activity front in the interdigestive period have been reported in dogs. In man, Lux et al. [15], using pharmacological doses of somatostatin, found a decrease in the period of the activity front, and we confirmed this observation [16]. The aim of the present study was to determine whether somatostatin has a physiological role in the regulation of the interdigestive motor activity in man.
Methods
Experimental protocol The study, approved by the ethical committee of the University Hospital, was performed in 8 normal human volunteers, 5 men and 3 women, aged between 22 and 34 years. Upper intestinal motor activity was monitored by means of seven perfused manometric catheters. The four proximal catheters had side-openings 3 cm apart. The distance between the three distal catheters was 25 cm, and catheters four and five were separated by 17 cm. The catheter assembly was positioned under fluoroscopic control so that the four proximal openings were located in the gastric antrum. In this way the 5th opening was in the duodenum and the two distal openings in the upper jejunum, 25 and 50 cm more distally. Details of the recording technique have been published elsewhere [1]. Measurements were started after an overnight fast and blood samples were drawn every 10 min throughout the study. In four volunteers (group I) blood sampling was done until two activity fronts had been recorded. In four other volunteers (group II) somatostatin (Serono, Freiburg, Germany) was infused in doses of 1.2, 2.4 and 4.8
211 p m o l / k g per min over three consecutive periods of 90 min. The infusion was started immediately after the passage of an activity front in the upper small bowel.
Radioimmunoassay Venous blood (5 ml) was collected into plastic tubes containing 0.5 ml of a solution of trisodium citrate (31.3 g/l) and aprotinin (5.109 units/l). Plasma was immediately separated and stored at - 2 0 ° C until radioimmunoassayed for motilin, PP and somatostatin by methods described previously [17-19]. Briefly, motilin was determined by a radioimmunoassay developed to porcine motilin with antibody (antiserum G-71) obtained from Dr. J.C. Brown (Vancouver, Canada). For concentrations from 0 to 150 pM, our assay has a precision of 9 pM (precision is defined as two standard errors). Pancreatic polypeptide was determined using reagents (bovine PP for iodination, antiserum to bovine PP, human PP as standard) obtained from Dr. R.E. Chance, Eli Lilly, Indianapolis, U.S.A., using the procedures described by Track et al. [20]. In a range of 0-120 pM our assay has a precision of 4 pM. Somatostatin was radioimmunoassayed after acid/ethanol extraction using a minor modification of our previously described procedure [18]. Indeed, it was found that after evaporation of the acid/ethanol supernatant, traces of acetic acid remained in the residue which influenced the p H during the incubation for radioimmunoassay. Therefore, a constant amount of N a O H was added to all tubes after evaporation. This amount was determined by titrating five control sera extracted and evaporated together with test samples to pH 7.0. The somatostatin radioimmunoassay has a precision of 5 pM in the 0-150 pM range.
Analysis of pressure records Manometric recordings were visually analysed during the experiment for the presence of intestinal activity fronts and this visual analysis was used to determine the end of the experimental period (group I) or the start of the infusion (group II). For further analysis, the number of contractions per min was counted on the manometric tracings for all recording sites and transformed into graphs such as the one shown in Figure 2. The criteria for the identification of an activity front were similar to those reported previously [1]. The gastric component of the activity front was defined as a burst of contractions, occurring at a rhythm of 3 per min, and followed by a period of quiescence.
Results
For seven of the eight activity fronts recorded in group I subjects, a peak of plasma somatostatin was found to coincide with the start of the front in the upper small bowel. The peak value was defined as the mean of 3 consecutive values, the first one being the plasma level obtained before the 10-min period in which the activity front was detected in the most proximal catheter. The mean peak value
212
( + S.E.) was 32 + 4 pM compared to 12 __.2 pM in the remaining period (P < 0.01). In Figure 1 somatostatin fluctuations are summarized using our previously described transformation technique [17]. In this technique data are obtained for ten equal subdivisions of the period between two activity fronts by interpolation, and expressed as normal variates. For these experiments, the transformation into normal variates, which we used on our motilin and PP data, was dropped. The purpose of the transformation into normal variates is to compensate for the wide interindividual fluctuations in motilin and PP levels, but as somatostatin levels in our volunteers were rather similar, this was not required. Dropping the transformation into normal variates has the advantage that the data are directly expressed in concentrations and more easily visualized. It is again seen that somatostatin shows a peak around the time of the start of the MMC in the gastroduodenal area. The intravenous infusion of somatostatin at doses of 1.2, 2.4 and 4.8 pmol/kg per min over three consecutive periods of 90 min caused dose dependent increments in plasma somatostatin of 7, 32 and 76 pM. Plasma motilin and plasma PP levels were already decreased by the lowest infusion dose to 80 and 74% of their pre-infusion level respectively. Further increases in the somatostatin infusion rate had little effect (Table I). In all subjects and for all doses, intestinal phase 2 was completely abolished and intestinal phase 3 occurred at increased frequency. This is clearly demonstrated by the example in Figure 2. The mean interval between two intestinal activity fronts was 39 + 6 rain compared to a normal period length of 92 min (P < 0.001). The characteristics of somatostatin-induced activity fronts were not different from those reported in earlier control experiments, as is shown in Table II, except for their point of origin.
50
P~
4o
Z
~- 30 0
~ z
20
I
I
I
I
l
I
I
i
50 °/o OF PERIOD LENGTH
I
100
Fig. 1. Analysis of the relationship between somatostatin levels (mean + S.D., n = 8) and the occurrence of activity fronts in the human small intestine. Using the start of phase 3 in the duodenum as a reference point (0.100%), the time interval between two intervals was divided into ten units, and somatostatin values were pooled accordingly. For further explanation see text.
213 TABLE I Hormone levels during somatostatin infusion (1.2, 2.4 and 4.8 p M / k g per rain) Volunteer
Somatostatin (A pM)
Motilin (%)
Pancreatic polypeptide (%)
1.2
2.4
4.8
1.2
2.4
4.8
1.2
2.4
4.8
P.Z. M.C. L.T. W.C.
13 1 8 7
44 25 23 38
98 55 72 78
75 70 83 90
82 70 73 72
76 63 78 74
63 77 82 75
53 77 68 73
47 57 72 52
Mean S.D.
7 5
32 10
76 18
80 9
74 5
72 7
74 8
67 10
57 11
Somatostatin levels are expressed as the difference between basal level and plateau level (ApM). Basal level is defined as the mean of all samples during phase 2 and plateau level asthe mean of 5 consecutive samples obtained 30 rain after the start of an infusion dose. Motilin and PP levels were averaged for the corresponding time periods but are expressed in % of basal level.
B e c a u s e it h a d b e e n n o t e d i n e a r l i e r e x p e r i m e n t s w i t h h i g h e r d o s e s o f s o m a t o s t a t i n [16], t h a t all g a s t r i c a c t i v i t y s e e m e d t o b e i n h i b i t e d , a s p e c i a l f e a t u r e o f t h e present series was the presence of four recording orifices in the stomach to ensure detection of gastric activity. In none of the experiments was gastric activity seen Somatostotin infusion 1.2
2.4
(pMol/kg.min) 4.8
I
antrum
3
o? 1.... ~t t, q It D
duodenum
1 t 1 kt ii ~i t I ~,,~.+ °~ti~+,,,,d| I, 1 i i Ut| i ~'it+~ lilL I i i itl I 1
13-
12
+ 25cm
n-
+50cm
j~ llI,L~,,
I
I
I
I
1
0
100
200
300
400 MINUTES
Fig. 2. Representation of the manometric recording from the human small intestine obtained in volunteer W.C. during a control period and three consecutive infusion periods of 90 min each. All four gastric channels gave similar results and only one is represented here.
214 TABLE II Characteristics of spontaneous a and of somatostatin-induced activity fronts Spontaneous
Infusion dose (pM/kg per min) 1.2
Duration (min) Frequency of contractions (No./min) Progression velocity (cm/min) b
2.4
4.8
5.25 + 0.34
5.08 + 0.79
5.13 _+0.55
5.58 + 0.74
11.38 5-0.19
11.68 + 0.39
11.43 + 0.18
11.33 + 0.14
6.76 5:1.36
7.19 + 2.72
7.89 + 1.46
6.68 + 1.28
Only the characteristics for the upper jejunal recording site are shown. Numbers refer to means __+S.E.M. a Values obtained during earlier control studies in normal subjects, see reference 20. b Between duodenal and upper jejunal recording site.
during somatostatin infusion, although with this procedure a gastric c o m p o n e n t can be detected in 90% of all spontaneously occurring activity fronts [21].
Discussion It is becoming increasingly evident that in the interdigestive state various physiological parameters show periodic fluctuations, all of them apparently in phase with the migrating m o t o r complex. Indeed, fluctuations of plasma motilin [ 17,22-24] and pancreatic polypeptide [19,24] are related to the M M C . So are gastric acid [25,26] pancreatic and bile [24,25,27,28] secretion. It is tempting to consider, therefore, the existence of a generalized periodicity of the digestive functions with a m o t o r component, a secretory c o m p o n e n t of exocrine functions, and a h o r m o n a l component involved in the regulation of the m o t o r and secretory phenomena. This paper demonstrates that there exists also a relationship between plasma somatostatin and the M M C . Previous attempts [ 15,16] failed to demonstrate such a correlation, p r o b a b l y because of the technical difficulties involved in the somatostatin radioimmunoassay and the short half-life of somatostatin. However, recently Aizawa et al. [29] demonstrated that such a relationship exists in dogs, with peaks of plasma somatostatin coinciding with gastric phase III contractions. This is in agreement with our results in man. Conflicting results have been reported on the effect of somatostatin infusions on the M M C . In dogs, T h o r et al. [11] found a stimulation but Ormsbee et al. [13] and Poitras et al. [14] reported an inhibition of the migrating m o t o r complex. Interestingly, Poitras et al. [14] noted that during somatostatin infusion, activity fronts still occurred but started at a lower level. Aizawa et al. [29] reported an inhibition of gastric interdigestive activity during somatostatin infusion. In man, Lux et al. [15] reported a stimulation of phase 3 activity during infusion of pharmacological doses of somatostatin. These observations were confirmed by us [16] but it was also noted
215 that under these circumstances phase 2 of the MMC was inhibited. Finally, no change in gastric motility was observed in monkeys during somatostatin infusion in the interdigestive or postprandial state [10]. The data reported in this paper partially resolve this controversy in that somatostatin blocks phase 3 in the stomach and stimulates it in the small intestine. The inhibition of phase 3 in the stomach might be related to the decrease in motilin levels, as there is increasing evidence that the target for motilin is the stomach [21]. The inhibition of phase 2 in the small intestine, on the other hand, could be due to the known suppression of other 'digestive' hormones such as gastrin, C C K and secretin by somatostatin. Indeed exogenous infusion of these substances induces a fed-like pattern which is similar to a strong phase 2. These observations are in favour of the concept that the influence of different hormonal factors depends upon the organ involved and upon the phase. In the experiments reported in this paper the rise in somatostatin levels provoked by the two lowest infusion doses might be considered as physiological. Indeed, after a meal we found a short rise of plasma somatostatin from 17 to 25 pM [30]. The short peaks that correspond with phase 3 activity are even higher. Our lowest dose already provoked a complete inhibition of phase 2, and a stimulation of phase 3 activity. This is in favour of the hypothesis that somatostatin is directly involved in the stimulation of intestinal phase 3. However, one can only speculate upon the mechanism by which somatostatin is acting. There are data in the literature suggesting that in normal physiological conditions the occurrence of the AF in the small intestine is inhibited: its cycling frequency in a denervated small bowel loop is higher than in the main intestine [31] and the AF in such a loop is no longer inhibited by feeding [32]. As the effect of somatostatin is generally inhibitory, its function in the regulation of the MMC may well be to inhibit an inhibitor, favoring a free-run of the AF in the small intestine at the moment of its start in the gastroduodenal area. In this way somatostatin may help to facilitate the orderly progression of the AF of the MMC from the stomach into the small intestine. Further work is needed to clarify this point.
Acknowledgements
The authors gratefully acknowledge the skillful technical assistance of T. Degreef, L. Gregoire, S. Spee, S. Vanweerts and R. Vos. These investigations were supported by the Belgian Foundation for Medical Research (FWGO, Brussels) grants No. 3.0040.79 and 3.0046.80 and by the Belgian Ministry for Science, contract No. 80/85-4.
References
1 Vantrappen, G., Janssens, J., Hellemans, J. and Ghoos, Y., The interdigestivemotor complex of normal subjectsand patientswith bacterialovergrowthof the small intestine,J. Clin. Invest.,59 (1977) 1158-1166.
216 2 Vantrappen, G., Janssens, J. and Peeters, T.L., The migrating motor complex, Med. Clin. North Am., 65 (1981) 1311-1329. 3 Kruhlich, L., Dhariwal, A.P.S. and McCann, S.M., Stimulatory and inhibitory effects of purified hypothalamic extracts on growth hormone release from rat pituitary in vitro, Endocrinology, 83 (1968) 783-790. 4 Brazeau, P., Vale, W., Burgus, R., Ling, N., Butcher, M., Rivier, J. and Guillemin, R., Hypothalamic peptide that inhibits the secretion of immunoreactive pituitary growth hormone, Science, 179 (1973) 77-79. 5 Patel, Y.C., Zing,g, H.H., Fitz-Patrick, D. and Srikant, C.B., Somatostatin: some aspects of its physiology and pathophysiology. In S.R. Bloom and J.M. Polak (Eds.), Gut Hormones, Churchill Livingstone, Edinburgh-London-Melbourne-New York, 1982, pp. 339-349. 6 Boden, G., Jacoby, H.I. and Straus, A., Somatostatin interacts with basal and carbachol stimulated antral and duodenal motility, Gastroenterology, 70 (1976) 961. 7 Tansy, M.F., Martin, J.S. and Landin, W.E., The differential action of somatostatin on the motor effector system of the canine gastrointestinal tract, Metabolism, 27 (1978) 1353-1357. 8 Bloom, S.R., Ralphs, D.N. and Besser, G.M., Effect of somatostatin on motilin levels and gastric emptying, Gut, 16 (1975) 834. 9 Johansson, C., Efendic, S. and Wisen, O., Effects of short term somatostatin infusion on gastric and intestinal propulsion in humans, Scand. J. Gastroenterol., 13 (1978) 481-483. 10 Koerker, D.J. and Hansen, B.C., Influence of somatostatin on gastric motility and meal absorption in rhesus monkeys, Macaca mulatta, Metabolism, 30 (1981) 335-339. 11 Thor, P., Krol, R. and Konturek, S.J., Effect of somatostatin on myoelectrical activity of the small bowel, Am. J. Physiol., 235 (1978) E249-E254. 12 Lee, K.Y. and Chert, W.Y., Actions of somatostatin and secretin on stimulated or spontaneous myoelectrical activities of gastric antrum and duodenum, Physiologist, 19 (1976) 266. 13 Ormsbee, H.S., Koehler, S.L. and Telford, G.L., Somatostatin inhibits motilin-induced interdigestive contractile activity in the dog, Dig. Dis., 23 (1978) 781-788. 14 Poitras, P., Steinbach, J.H. and Van Deventer, G., Motilin-independent ectopic fronts of the interdigestive myoelectric complex in dogs, Am. J. Physiol., 239 (1980) GI25-G220. 15 Lux, G., Femppel, J. and Lederer, P., Somatostatin induces interdigestive intestinal motor and secretory complex-like activity in man, Gastroenterology, 78 (1980) 1212. 16 Peeters, T.L., Depraetere, Y. and Vantrappen, G.R., Somatostatin and the interdigestive motor complex (MMC) in man, Gastroenterology, 80 (1981) 1250. 17 Peeters, T.L., Vantrappen G.R. and Janssens, J., Fasting plasma motilin levels are related to the interdigestive motor complex, Gastroenterology, 79 (1980) 716-719. 18 Peeters, T.L., Depraetere, Y. and Vantrappen, G.R., Radioimmunoassay for somatostatin using a simple extraction method, Clin. Chem., 27 (1981) 888-891. 19 Janssens, J., Hellemans, J., Adrian, T.E., Bloom, S.R., Peeters, T.L., Christofides, N. and Vantrappen, G., Pancreatic polypeptide is not involved in the regulation of the migrating motor complex in man, Regul. Peptides, 3 (1982) 41-49. 20 Track, N.S., Watters, L.M., and Gauldie, J., Motilin, human pancreatic potypeptide and gastrin plasma concentrations in fasting subjects, Clin. Biochem., 12 (1979) 109- I 17. 21 Janssens, J., Vantrappen, G., Peeters, T. and Hellemans, J., The activity front of the migrating motor complex of the human stomach but not of the small intestine is motilin dependent Gastroenterology, 82 (1982) 1093. 22 Lee, K.Y., Chey, W.Y., Tai, H. and Yajima, H., Radioimmunoassay of motilin: validation and studies on the relationship between plasma motilin and interdigestive myoelectric activity of the duodenum of dog, Am. J. Dig. Dis., 23 (1978) 789-795. 23 Itoh, Z., Takeuchi, S., Aizawa, 1., Mori, K., Taminato, T., Seino, Y., Imura, H. and Yanaihara, N., Changes in plasma motilin concentration and gastrointestinal contractile activity in conscious dog, Am. J. Dig. Dis., 23 (1978) 929-935. 24 Keane, F.B., DiMagno, E.P., Dozois, R.R. and Go, V.L.M., Relationship among canine interdigestive exocrine pancreatic and biliary flow, duodenal motor activity, plasma pancreatic polypeptide, and motilin, Gastroenterology, 78 (1980) 310-316.
217 25 Steinbach, J.H., Hines, J.C. and Code, C.F., Gastric acid secretion during the interdigestive myoelectric complex. 63rd Annual Meeting of the American Physiological Society, April 6-10, 1979. 26 Vantrappen, G., Peeters, T.L. and Janssens, J., The secretory component of the interdigestive migrating motor complex in man, Scand. J. Gastroenterol., 14 (1979) 663-668. 27 DiMagno, E.P., Hendricks, J.C., Go, V.L.W. and Dozois, R.R., Relationships among canine fasting pancreatic and biliary secretions, pancreatic duct pressure and duodenal phase III motor activity - - Boldryeff revisited, Dig. Dis. Sci., 24 (1979) 689-693. 28 Peeters, T.L., Vantrappen, G. and Janssens, J., Bile acid output and the interdigestive migrating motor complex in normals and in cholecystectomized patients, Gastroenterology, 79 (1980) 678-681. 29 Aizawa, I., Itoh, Z., Harris, V. and Unger, R., Plasma somatostatin-like immunoreactivity during the interdigestive period in the dog, J. Clin. Investig., 68 (1981) 206-213, and Med. Clin. North Am., 65 (1982) 1311-1329. 30 Peeters, T.L., Depraetere, Y., Vantrappen, G. and Janssens, J., Plasma somatostatin in man. In Proceedings of the Serono Symposia, Somatostatin, Athens 1981, 1982, in press. 31 Sarr, M.G. and Kelly, K.A., Presence of interdigestive myoelectric complex in autotransplanted (extrinsically denervated) canine jejunum, Gastroenterology, 78 (1980) 1251. 32 Weisbrodt, N.W., Copeland, E.M., Moore, E.P., Kearly, R.W. and Johnson, L.R., Effect of vagotomy on electrical activity of the small intestine of the dog. Am. J. Physiol., 228 (1975) 650-654.
209
Elsevier BiomedicalPress
Somatostatin and the interdigestive migrating motor complex in man T.L. Peeters, J. Janssens and G.R. Vantrappen Division of Gastroenterology, Department of Medical Research, Catholic University of Leuven, Gasthuisberg, B- 3000 Leuven, Belgium
(Received27 September1982; accepted for publication 3 December 1982) Summary The relationship between somatostatin and the interdigestive migrating motility complex (MMC) was determined in human volunteers. Motor activity was monitored manometrically by means of seven perfused catheters: four in the stomach, one in the duodenum, two in the jejunum. Blood samples were drawn every 10 min and radioimmunoassayed for motilin, pancreatic polypeptide and somatostatin. In four volunteers two activity fronts (AF) were recorded and somatostatin levels correlated to the manometric data. The start of an A F in the upper duodenum was accompanied by somatostatin peaks. Peak values, taken as the mean of the levels in the sample obtained after the start of an AF, the preceding sample and the next one, averaged 32 + 4 pM compared to 12 + 2 pM in the remaining period. In four volunteers somatostatin was infused in doses of 1.2, 2.4 and 4.8 p M / k g per min over three consecutive periods of 90 min, causing dose-dependent increments in plasma somatostatin levels of 7, 32 and 76 pM. In all volunteers and for all doses all gastric activity was completely inhibited. In the intestine phase 2 was abolished but phase 3 stimulated: during somatostatin infusion phase 3 occurred with an interval of 39 _ 6 min. Motilin and PP levels were decreased. As the two lowest infusion doses caused increases in somatostatin levels that might be considered as physiological, somatostatin seems to have a physiological role in the regulation of the migrating motor complex. We propose that it facilitates the progressing of the activity front into the small intestine. intestinal motility; gastric motility; interdigestive motility; motilin; pancreatic polypeptide
Introduction
The interdigestive migrating motor complex (MMC) is a typical motility pattern of the small intestine in the fasting state, consisting essentially of a rest period 0167-0115/83/0000-0000/$03.00 © 1983 ElsevierBiomedicalPress
210 (phase 1), a period of irregular contractions (phase 2), a period of rhythmic contractions occurring at maximal frequency (phase 3) and a period of rapid decline in activity (phase 4). All phases progress aborally, and occur cyclically with a period of about 92 min in man [1]. Several gastrointestinal hormones have been shown to be related to the MMC. From a recent review on this subject, it can be concluded that there is evidence for the involvement of motilin in the initiation of phase 3 in the stomach. Infusion of cholecystokinin, gastrin, glucagon, insulin and secretin disrupts the interdigestive pattern and produces a fed pattern [2]. Somatostatin, originally discovered as the growth hormone release inhibiting factor [3;4], has also been found to influence gastrointestinal function. It is generally accepted that it has a powerful inhibitory effect on the release of gastrointestinal hormones and on gastric and pancreatic secretion [5]. However, its influence on gastrointestinal motility is controversial: depending upon species, organ, dose and various other factors, different and sometimes conflicting results have been reported. In anesthetized dogs, Boden et al. [6] did not observe an effect on gastric antral motility, but Tansy et al. [7] noted a brief inhibitory effect. In man, inhibitory effects [8] on gastric motor function and an initial increase followed by a decrease of the gastric emptying rate [9] were observed. In monkeys, somatostatin had no effect on antral motility [10]. In the small intestine induction [11] as well as inhibition [12-14] of the activity front in the interdigestive period have been reported in dogs. In man, Lux et al. [15], using pharmacological doses of somatostatin, found a decrease in the period of the activity front, and we confirmed this observation [16]. The aim of the present study was to determine whether somatostatin has a physiological role in the regulation of the interdigestive motor activity in man.
Methods
Experimental protocol The study, approved by the ethical committee of the University Hospital, was performed in 8 normal human volunteers, 5 men and 3 women, aged between 22 and 34 years. Upper intestinal motor activity was monitored by means of seven perfused manometric catheters. The four proximal catheters had side-openings 3 cm apart. The distance between the three distal catheters was 25 cm, and catheters four and five were separated by 17 cm. The catheter assembly was positioned under fluoroscopic control so that the four proximal openings were located in the gastric antrum. In this way the 5th opening was in the duodenum and the two distal openings in the upper jejunum, 25 and 50 cm more distally. Details of the recording technique have been published elsewhere [1]. Measurements were started after an overnight fast and blood samples were drawn every 10 min throughout the study. In four volunteers (group I) blood sampling was done until two activity fronts had been recorded. In four other volunteers (group II) somatostatin (Serono, Freiburg, Germany) was infused in doses of 1.2, 2.4 and 4.8
211 p m o l / k g per min over three consecutive periods of 90 min. The infusion was started immediately after the passage of an activity front in the upper small bowel.
Radioimmunoassay Venous blood (5 ml) was collected into plastic tubes containing 0.5 ml of a solution of trisodium citrate (31.3 g/l) and aprotinin (5.109 units/l). Plasma was immediately separated and stored at - 2 0 ° C until radioimmunoassayed for motilin, PP and somatostatin by methods described previously [17-19]. Briefly, motilin was determined by a radioimmunoassay developed to porcine motilin with antibody (antiserum G-71) obtained from Dr. J.C. Brown (Vancouver, Canada). For concentrations from 0 to 150 pM, our assay has a precision of 9 pM (precision is defined as two standard errors). Pancreatic polypeptide was determined using reagents (bovine PP for iodination, antiserum to bovine PP, human PP as standard) obtained from Dr. R.E. Chance, Eli Lilly, Indianapolis, U.S.A., using the procedures described by Track et al. [20]. In a range of 0-120 pM our assay has a precision of 4 pM. Somatostatin was radioimmunoassayed after acid/ethanol extraction using a minor modification of our previously described procedure [18]. Indeed, it was found that after evaporation of the acid/ethanol supernatant, traces of acetic acid remained in the residue which influenced the p H during the incubation for radioimmunoassay. Therefore, a constant amount of N a O H was added to all tubes after evaporation. This amount was determined by titrating five control sera extracted and evaporated together with test samples to pH 7.0. The somatostatin radioimmunoassay has a precision of 5 pM in the 0-150 pM range.
Analysis of pressure records Manometric recordings were visually analysed during the experiment for the presence of intestinal activity fronts and this visual analysis was used to determine the end of the experimental period (group I) or the start of the infusion (group II). For further analysis, the number of contractions per min was counted on the manometric tracings for all recording sites and transformed into graphs such as the one shown in Figure 2. The criteria for the identification of an activity front were similar to those reported previously [1]. The gastric component of the activity front was defined as a burst of contractions, occurring at a rhythm of 3 per min, and followed by a period of quiescence.
Results
For seven of the eight activity fronts recorded in group I subjects, a peak of plasma somatostatin was found to coincide with the start of the front in the upper small bowel. The peak value was defined as the mean of 3 consecutive values, the first one being the plasma level obtained before the 10-min period in which the activity front was detected in the most proximal catheter. The mean peak value
212
( + S.E.) was 32 + 4 pM compared to 12 __.2 pM in the remaining period (P < 0.01). In Figure 1 somatostatin fluctuations are summarized using our previously described transformation technique [17]. In this technique data are obtained for ten equal subdivisions of the period between two activity fronts by interpolation, and expressed as normal variates. For these experiments, the transformation into normal variates, which we used on our motilin and PP data, was dropped. The purpose of the transformation into normal variates is to compensate for the wide interindividual fluctuations in motilin and PP levels, but as somatostatin levels in our volunteers were rather similar, this was not required. Dropping the transformation into normal variates has the advantage that the data are directly expressed in concentrations and more easily visualized. It is again seen that somatostatin shows a peak around the time of the start of the MMC in the gastroduodenal area. The intravenous infusion of somatostatin at doses of 1.2, 2.4 and 4.8 pmol/kg per min over three consecutive periods of 90 min caused dose dependent increments in plasma somatostatin of 7, 32 and 76 pM. Plasma motilin and plasma PP levels were already decreased by the lowest infusion dose to 80 and 74% of their pre-infusion level respectively. Further increases in the somatostatin infusion rate had little effect (Table I). In all subjects and for all doses, intestinal phase 2 was completely abolished and intestinal phase 3 occurred at increased frequency. This is clearly demonstrated by the example in Figure 2. The mean interval between two intestinal activity fronts was 39 + 6 rain compared to a normal period length of 92 min (P < 0.001). The characteristics of somatostatin-induced activity fronts were not different from those reported in earlier control experiments, as is shown in Table II, except for their point of origin.
50
P~
4o
Z
~- 30 0
~ z
20
I
I
I
I
l
I
I
i
50 °/o OF PERIOD LENGTH
I
100
Fig. 1. Analysis of the relationship between somatostatin levels (mean + S.D., n = 8) and the occurrence of activity fronts in the human small intestine. Using the start of phase 3 in the duodenum as a reference point (0.100%), the time interval between two intervals was divided into ten units, and somatostatin values were pooled accordingly. For further explanation see text.
213 TABLE I Hormone levels during somatostatin infusion (1.2, 2.4 and 4.8 p M / k g per rain) Volunteer
Somatostatin (A pM)
Motilin (%)
Pancreatic polypeptide (%)
1.2
2.4
4.8
1.2
2.4
4.8
1.2
2.4
4.8
P.Z. M.C. L.T. W.C.
13 1 8 7
44 25 23 38
98 55 72 78
75 70 83 90
82 70 73 72
76 63 78 74
63 77 82 75
53 77 68 73
47 57 72 52
Mean S.D.
7 5
32 10
76 18
80 9
74 5
72 7
74 8
67 10
57 11
Somatostatin levels are expressed as the difference between basal level and plateau level (ApM). Basal level is defined as the mean of all samples during phase 2 and plateau level asthe mean of 5 consecutive samples obtained 30 rain after the start of an infusion dose. Motilin and PP levels were averaged for the corresponding time periods but are expressed in % of basal level.
B e c a u s e it h a d b e e n n o t e d i n e a r l i e r e x p e r i m e n t s w i t h h i g h e r d o s e s o f s o m a t o s t a t i n [16], t h a t all g a s t r i c a c t i v i t y s e e m e d t o b e i n h i b i t e d , a s p e c i a l f e a t u r e o f t h e present series was the presence of four recording orifices in the stomach to ensure detection of gastric activity. In none of the experiments was gastric activity seen Somatostotin infusion 1.2
2.4
(pMol/kg.min) 4.8
I
antrum
3
o? 1.... ~t t, q It D
duodenum
1 t 1 kt ii ~i t I ~,,~.+ °~ti~+,,,,d| I, 1 i i Ut| i ~'it+~ lilL I i i itl I 1
13-
12
+ 25cm
n-
+50cm
j~ llI,L~,,
I
I
I
I
1
0
100
200
300
400 MINUTES
Fig. 2. Representation of the manometric recording from the human small intestine obtained in volunteer W.C. during a control period and three consecutive infusion periods of 90 min each. All four gastric channels gave similar results and only one is represented here.
214 TABLE II Characteristics of spontaneous a and of somatostatin-induced activity fronts Spontaneous
Infusion dose (pM/kg per min) 1.2
Duration (min) Frequency of contractions (No./min) Progression velocity (cm/min) b
2.4
4.8
5.25 + 0.34
5.08 + 0.79
5.13 _+0.55
5.58 + 0.74
11.38 5-0.19
11.68 + 0.39
11.43 + 0.18
11.33 + 0.14
6.76 5:1.36
7.19 + 2.72
7.89 + 1.46
6.68 + 1.28
Only the characteristics for the upper jejunal recording site are shown. Numbers refer to means __+S.E.M. a Values obtained during earlier control studies in normal subjects, see reference 20. b Between duodenal and upper jejunal recording site.
during somatostatin infusion, although with this procedure a gastric c o m p o n e n t can be detected in 90% of all spontaneously occurring activity fronts [21].
Discussion It is becoming increasingly evident that in the interdigestive state various physiological parameters show periodic fluctuations, all of them apparently in phase with the migrating m o t o r complex. Indeed, fluctuations of plasma motilin [ 17,22-24] and pancreatic polypeptide [19,24] are related to the M M C . So are gastric acid [25,26] pancreatic and bile [24,25,27,28] secretion. It is tempting to consider, therefore, the existence of a generalized periodicity of the digestive functions with a m o t o r component, a secretory c o m p o n e n t of exocrine functions, and a h o r m o n a l component involved in the regulation of the m o t o r and secretory phenomena. This paper demonstrates that there exists also a relationship between plasma somatostatin and the M M C . Previous attempts [ 15,16] failed to demonstrate such a correlation, p r o b a b l y because of the technical difficulties involved in the somatostatin radioimmunoassay and the short half-life of somatostatin. However, recently Aizawa et al. [29] demonstrated that such a relationship exists in dogs, with peaks of plasma somatostatin coinciding with gastric phase III contractions. This is in agreement with our results in man. Conflicting results have been reported on the effect of somatostatin infusions on the M M C . In dogs, T h o r et al. [11] found a stimulation but Ormsbee et al. [13] and Poitras et al. [14] reported an inhibition of the migrating m o t o r complex. Interestingly, Poitras et al. [14] noted that during somatostatin infusion, activity fronts still occurred but started at a lower level. Aizawa et al. [29] reported an inhibition of gastric interdigestive activity during somatostatin infusion. In man, Lux et al. [15] reported a stimulation of phase 3 activity during infusion of pharmacological doses of somatostatin. These observations were confirmed by us [16] but it was also noted
215 that under these circumstances phase 2 of the MMC was inhibited. Finally, no change in gastric motility was observed in monkeys during somatostatin infusion in the interdigestive or postprandial state [10]. The data reported in this paper partially resolve this controversy in that somatostatin blocks phase 3 in the stomach and stimulates it in the small intestine. The inhibition of phase 3 in the stomach might be related to the decrease in motilin levels, as there is increasing evidence that the target for motilin is the stomach [21]. The inhibition of phase 2 in the small intestine, on the other hand, could be due to the known suppression of other 'digestive' hormones such as gastrin, C C K and secretin by somatostatin. Indeed exogenous infusion of these substances induces a fed-like pattern which is similar to a strong phase 2. These observations are in favour of the concept that the influence of different hormonal factors depends upon the organ involved and upon the phase. In the experiments reported in this paper the rise in somatostatin levels provoked by the two lowest infusion doses might be considered as physiological. Indeed, after a meal we found a short rise of plasma somatostatin from 17 to 25 pM [30]. The short peaks that correspond with phase 3 activity are even higher. Our lowest dose already provoked a complete inhibition of phase 2, and a stimulation of phase 3 activity. This is in favour of the hypothesis that somatostatin is directly involved in the stimulation of intestinal phase 3. However, one can only speculate upon the mechanism by which somatostatin is acting. There are data in the literature suggesting that in normal physiological conditions the occurrence of the AF in the small intestine is inhibited: its cycling frequency in a denervated small bowel loop is higher than in the main intestine [31] and the AF in such a loop is no longer inhibited by feeding [32]. As the effect of somatostatin is generally inhibitory, its function in the regulation of the MMC may well be to inhibit an inhibitor, favoring a free-run of the AF in the small intestine at the moment of its start in the gastroduodenal area. In this way somatostatin may help to facilitate the orderly progression of the AF of the MMC from the stomach into the small intestine. Further work is needed to clarify this point.
Acknowledgements
The authors gratefully acknowledge the skillful technical assistance of T. Degreef, L. Gregoire, S. Spee, S. Vanweerts and R. Vos. These investigations were supported by the Belgian Foundation for Medical Research (FWGO, Brussels) grants No. 3.0040.79 and 3.0046.80 and by the Belgian Ministry for Science, contract No. 80/85-4.
References
1 Vantrappen, G., Janssens, J., Hellemans, J. and Ghoos, Y., The interdigestivemotor complex of normal subjectsand patientswith bacterialovergrowthof the small intestine,J. Clin. Invest.,59 (1977) 1158-1166.
216 2 Vantrappen, G., Janssens, J. and Peeters, T.L., The migrating motor complex, Med. Clin. North Am., 65 (1981) 1311-1329. 3 Kruhlich, L., Dhariwal, A.P.S. and McCann, S.M., Stimulatory and inhibitory effects of purified hypothalamic extracts on growth hormone release from rat pituitary in vitro, Endocrinology, 83 (1968) 783-790. 4 Brazeau, P., Vale, W., Burgus, R., Ling, N., Butcher, M., Rivier, J. and Guillemin, R., Hypothalamic peptide that inhibits the secretion of immunoreactive pituitary growth hormone, Science, 179 (1973) 77-79. 5 Patel, Y.C., Zing,g, H.H., Fitz-Patrick, D. and Srikant, C.B., Somatostatin: some aspects of its physiology and pathophysiology. In S.R. Bloom and J.M. Polak (Eds.), Gut Hormones, Churchill Livingstone, Edinburgh-London-Melbourne-New York, 1982, pp. 339-349. 6 Boden, G., Jacoby, H.I. and Straus, A., Somatostatin interacts with basal and carbachol stimulated antral and duodenal motility, Gastroenterology, 70 (1976) 961. 7 Tansy, M.F., Martin, J.S. and Landin, W.E., The differential action of somatostatin on the motor effector system of the canine gastrointestinal tract, Metabolism, 27 (1978) 1353-1357. 8 Bloom, S.R., Ralphs, D.N. and Besser, G.M., Effect of somatostatin on motilin levels and gastric emptying, Gut, 16 (1975) 834. 9 Johansson, C., Efendic, S. and Wisen, O., Effects of short term somatostatin infusion on gastric and intestinal propulsion in humans, Scand. J. Gastroenterol., 13 (1978) 481-483. 10 Koerker, D.J. and Hansen, B.C., Influence of somatostatin on gastric motility and meal absorption in rhesus monkeys, Macaca mulatta, Metabolism, 30 (1981) 335-339. 11 Thor, P., Krol, R. and Konturek, S.J., Effect of somatostatin on myoelectrical activity of the small bowel, Am. J. Physiol., 235 (1978) E249-E254. 12 Lee, K.Y. and Chert, W.Y., Actions of somatostatin and secretin on stimulated or spontaneous myoelectrical activities of gastric antrum and duodenum, Physiologist, 19 (1976) 266. 13 Ormsbee, H.S., Koehler, S.L. and Telford, G.L., Somatostatin inhibits motilin-induced interdigestive contractile activity in the dog, Dig. Dis., 23 (1978) 781-788. 14 Poitras, P., Steinbach, J.H. and Van Deventer, G., Motilin-independent ectopic fronts of the interdigestive myoelectric complex in dogs, Am. J. Physiol., 239 (1980) GI25-G220. 15 Lux, G., Femppel, J. and Lederer, P., Somatostatin induces interdigestive intestinal motor and secretory complex-like activity in man, Gastroenterology, 78 (1980) 1212. 16 Peeters, T.L., Depraetere, Y. and Vantrappen, G.R., Somatostatin and the interdigestive motor complex (MMC) in man, Gastroenterology, 80 (1981) 1250. 17 Peeters, T.L., Vantrappen G.R. and Janssens, J., Fasting plasma motilin levels are related to the interdigestive motor complex, Gastroenterology, 79 (1980) 716-719. 18 Peeters, T.L., Depraetere, Y. and Vantrappen, G.R., Radioimmunoassay for somatostatin using a simple extraction method, Clin. Chem., 27 (1981) 888-891. 19 Janssens, J., Hellemans, J., Adrian, T.E., Bloom, S.R., Peeters, T.L., Christofides, N. and Vantrappen, G., Pancreatic polypeptide is not involved in the regulation of the migrating motor complex in man, Regul. Peptides, 3 (1982) 41-49. 20 Track, N.S., Watters, L.M., and Gauldie, J., Motilin, human pancreatic potypeptide and gastrin plasma concentrations in fasting subjects, Clin. Biochem., 12 (1979) 109- I 17. 21 Janssens, J., Vantrappen, G., Peeters, T. and Hellemans, J., The activity front of the migrating motor complex of the human stomach but not of the small intestine is motilin dependent Gastroenterology, 82 (1982) 1093. 22 Lee, K.Y., Chey, W.Y., Tai, H. and Yajima, H., Radioimmunoassay of motilin: validation and studies on the relationship between plasma motilin and interdigestive myoelectric activity of the duodenum of dog, Am. J. Dig. Dis., 23 (1978) 789-795. 23 Itoh, Z., Takeuchi, S., Aizawa, 1., Mori, K., Taminato, T., Seino, Y., Imura, H. and Yanaihara, N., Changes in plasma motilin concentration and gastrointestinal contractile activity in conscious dog, Am. J. Dig. Dis., 23 (1978) 929-935. 24 Keane, F.B., DiMagno, E.P., Dozois, R.R. and Go, V.L.M., Relationship among canine interdigestive exocrine pancreatic and biliary flow, duodenal motor activity, plasma pancreatic polypeptide, and motilin, Gastroenterology, 78 (1980) 310-316.
217 25 Steinbach, J.H., Hines, J.C. and Code, C.F., Gastric acid secretion during the interdigestive myoelectric complex. 63rd Annual Meeting of the American Physiological Society, April 6-10, 1979. 26 Vantrappen, G., Peeters, T.L. and Janssens, J., The secretory component of the interdigestive migrating motor complex in man, Scand. J. Gastroenterol., 14 (1979) 663-668. 27 DiMagno, E.P., Hendricks, J.C., Go, V.L.W. and Dozois, R.R., Relationships among canine fasting pancreatic and biliary secretions, pancreatic duct pressure and duodenal phase III motor activity - - Boldryeff revisited, Dig. Dis. Sci., 24 (1979) 689-693. 28 Peeters, T.L., Vantrappen, G. and Janssens, J., Bile acid output and the interdigestive migrating motor complex in normals and in cholecystectomized patients, Gastroenterology, 79 (1980) 678-681. 29 Aizawa, I., Itoh, Z., Harris, V. and Unger, R., Plasma somatostatin-like immunoreactivity during the interdigestive period in the dog, J. Clin. Investig., 68 (1981) 206-213, and Med. Clin. North Am., 65 (1982) 1311-1329. 30 Peeters, T.L., Depraetere, Y., Vantrappen, G. and Janssens, J., Plasma somatostatin in man. In Proceedings of the Serono Symposia, Somatostatin, Athens 1981, 1982, in press. 31 Sarr, M.G. and Kelly, K.A., Presence of interdigestive myoelectric complex in autotransplanted (extrinsically denervated) canine jejunum, Gastroenterology, 78 (1980) 1251. 32 Weisbrodt, N.W., Copeland, E.M., Moore, E.P., Kearly, R.W. and Johnson, L.R., Effect of vagotomy on electrical activity of the small intestine of the dog. Am. J. Physiol., 228 (1975) 650-654.