A New Twist in the Brain-Gut Axis DAVID C. WHITCOMB,
MD, PHD,* IAN L. TAYLOR,
ABSTRACT: Gastrointestinal functions are precisely regulated by hormonal and neural negative feedback loops. In addition to the classic hormonal and vago-vagal reflex mechanisms, these studies indicate that there are direct actions of gut hormones on the dorsal vagal complex. The current data demonstrate that pancreatic polypeptide is released into the circulation by vagal-cholinergic dependent mechanisms. It travels to the brainstem in the circulation, traverses the blood-brain barrier through "leaky" regions of this barrier in the area postrema and nucleus of the tractus solitarius and binds to specific receptors in the dorsal vagal complex. By binding to these sites, pancreatic polypeptide can directly inhibit vagal input to the pancreas and other gastrointestinal organs. These observations provide an anatomic basis to explain why pancreatic polypeptide is a more potent inhibitor of the action of central stimulants of pancreatic secretion than it is of the response to peripheral secretogogues. They also establish a novel mechanism by which gut peptides can influence brain function directly. KEY INDEXING TERMS: Brain-gut axis; Brain-gut peptides; Pancreas; Pancreatic polypeptide; Pancreatic secretion; Peptide YY. [Am J Med Sci 1992; 304(5):334-338.]
n the 19th century, the great Russian physiologist Pavlov concluded that the nervous system controlled all digestive functions. This concept became From the "'Department of Medicine, Diuision of Gastroenterology and Hepatology, and the Department of Physiology, Uniuersity of Pittsburgh, Pittsburgh, Pennsyluania, and the tDepartment of Medicine, Diuision of Gastroenterology, Duke Uniuersity Medical Center and the Durham VA Medical Center, Durham, North Carolina. Supported by research grant from the Veterans Administration and Grant # DK 44072 from the National Institutes of Health, Bethesda, MD. Presented at the Subspecialty Meetings jointly sponsored by the Southern Society for Clinical Inuestigation, Southern Section American Federation for Clinical Research, Southern Society for Pediatric Research, and Southern Region for I nuestigatiue Dermatology in New Orleans, LA on January 31, 1992. Correspondence: Ian L. Taylor, MD, Diuision of Gastroenterology, Box 3913, Duke Uniuersity Medical Center, Durham, NC 27710.
untenable when Baylis and Starling discovered secretin in 1902 and coined the term "hormone" to describe a chemical messenger released into the circulation to have effects on a distant target. Over the next three to four decades, much attention was given to hormonal control of gut function to the exclusion of the nervous system. However, recent discoveries have clearly shown the interdependency of the nervous and endocrine systems in controlling gastrointestinal physiology, that is, the endocrine system, itself, is modulated by neural inputs, and the endocrine system, in turn, modulates the nervous system. This review will address how a gut hormone, pancreatic polypeptide (PP), modulates the central nervous system by binding to and modulating the function of the vagal nuclei. Overview of the Digestive Process
Every aspect of the gastrointestinal function is precisely regulated and integrated. The sight, smell, and taste of food induce a cephalic phase that results in the stimulation of a series of secretory and motor events that prepare the gut-a classic Pavlovian experiment. . As the meal is ingested and the stomach fills, a variety of neural reflexes and hormonal events are initiated which depend, in part, on the physical, chemical and nutritional properties of the meal. Pulverized, partially digested nutrients are delivered to the small intestine at a manageable rate that is coordinated with the entry of pancreatic enzymes and bile acids that facilitate the digestive and absorptive functions of the small bowel. The mechanisms involved in coordinating this process are complex and involve both nervous and hormonal systems. The control of pancreatic secretion serves as an excellent model in which to study interactions between the nervous and endocrine systems. Feedback Inhibition of Pancreatic Secretion
Pancreatic exocrine secretion is precisely regulated through several hormonal and neural feedback mechanisms. Regulation of pH and proteolytic enzyme concentration in the intestinal lumen by secretin and cholecystokinin (CCK) are two examples. Secretin is released from the duodenum when the luminal pH drops below 4.5. 1•2 It then circulates to the pancreatic ductal cells and stimulates secretion of a pancreatic fluid rich in bicarbonate (Figure 1A). Bicarbonate secretion continues until the luminal pH is greater than pH 4.5, at which time secretin release stops. Thus, a mechanism November 1992 Volume 304 Number 5
Whitcomb and Taylor
Physiologic Inhibition of Pancreatic Secretion Circulation
~:::~n ? CCK.RF
Ductal cells (bicarbonate)
Figure 1. Feedback regulation of luminal pH and protein concen· tration through hormonal mechanisms. (A) luminal pH < 4.5 (acid) results in release of secretin into the circulation. Secretin acts on pancreatic duct cells to secrete pancreatic juice high in bicarbonate. The bicarbonate neutralized the acid, eliminating the stimulus for secretin release. (B) Luminal proteins result in the accumulation of cholecystokinin releasing factors (CCK-RF) that cause the release of CCK into the circulation. CCK acts on pancreatic acinar cells to secrete zymogen granules high in proteolytic enzymes. The enzymes digest luminal protein and the CCK-RF, eliminating the stimulus for CCK release.
exists to regulate intestinal pH so that it is optimal for pancreatic enzyme function. 3•4 In rats and, possibly, man, the presence of protein in the intestine results in accumulation of proteasesensitive CCK-releasing factors in the intestinal lumen. 5 - S Cholecystokinin is released and circulates to the acinar cells, where it stimulates release of the enzymes that digest luminal proteins including CCK-releasing factors. As a result, the stimulus for CCK release is eliminated (Figure IB). However, if the enzyme activity is saturated with proteins, the CCK-releasing factor is temporarily protected and continues to stimulate CCK release and, therefore, pancreatic enzyme secretion. Protease inhibitors exert an effect similar to undigested protein. When protease inhibitors are present, the CCK-releasing factor is protected, resulting in continued CCK release and pancreatic secretion.5-8 These observations suggest that another negative feedback loop exists to regulate pancreatic enzyme secretion through modulation of CCK release. Although the parasympathetic nervous system appears to contribute 50% or more of the stimulus for pancreatic secretion in many species, including man and rat, the mechanisms for feedback regulation of vagal stimulation are poorly understood.1-3 However, the vagus, itself, is subject to feedback inhibition. THE AMERICAN JOURNAL OF THE MEDICAL SCIENCES
The pancreas appears to be under both stimulatory and inhibitory control following the ingestion of a meal. l.2 The stimulatory mechanisms involve the action of secretin, CCK, and parasympathetic nerves. However, the maximal pancreatic response to CCK and secretin is greater than the response to a meal. 9.10 Further, the pancreatic secretory response to a meal can be inhibited by duodenal distention or by perfusing the duodenum or jejunum with hyperosmolar solutions. 1l- l4 Pancreatic secretion can also be inhibited by perfusion of the ileum and colon with oleic acid, sodium oleate, or hypertonic glucose.15 - 19 Many of these inhibitory events are mediated, at least in part, by hormones. l .2 Hormones in the PP family (pp,20-22 peptide YY [PYY],16-1S neuropeptide Y),23 somatostatin,24.25 enkephalins,26 and pancreastatin27 all inhibit pancreatic secretion. The physiologic importance of the inhibitory effects of pp28 and pyy29 is supported by immunoneutralization studies that demonstrate enhancement of pancreatic secretion. In vivo neutralization of PP or PYY by injection of specific antisera results in significant increases in meal-stimulated pancreatic protein and fluid output. These findings are part of a growing body of evidence that control of pancreatic secretion is under both stimulatory and inhibitory tone during digestion of a meal. Current data suggest an indirect mechanism of action for the hormones that inhibit pancreatic exocrine secretion under physiologic conditions. Two general features characterize these peptides, as illustrated by PP and PYY. First, there are no receptors for these peptides on acinar or ductal cells.23.26.3o Second, studies suggest that the inhibitory effects of pp,31.32 Pyy17.33 and other inhibitory peptides (2) involves inhibition of cholinergic transmission. Therefore, it appears that these hormones regulate the neuronal, rather than hormonal, stimuli of pancreatic secretion. However, the sites of action of PP and PYY are unclear. Recent studies have provided insight into the mechanisms of action of these peptides and have given evidence for a novel mechanism by which these hormones can modulate vagal tone. Physiology of Pancreatic Polypeptide
Pancreatic polypeptide, PYY, and neuropeptide Y are structurally-related 36 amino acid peptides.34 Pancreatic polypeptide is secreted from endocrine cells in the islets of Langerhans and scattered among pancreatic acinar cells, especially in the ventral pancreas.35-37 Pancreatic polypeptide is released from the endocrine pancreas after ingestion of a meal via vagal, cholinergicdependent mechanisms.38.39 Infusion ofPP in amounts that reproduce blood concentrations observed after a meal inhibits pancreatic exocrine secretion in vivo,2o-22 suggesting that the effects of PP on the pancreas are physiologic. Although PP inhibits pancreatic secretion in vivo, it has no effect on CCK-stimulated secretion
To examine this question, we screened the circumventricular organs, including the area postrema (AP) and nucleus tractus solitarius (NTS), using in vitro receptor autoradiography to locate putative PP receptors in the rat brain. 43 A dense distribution of high affinity, saturable binding sites that are specific for PP were observed in the AP, the NTS, and the dorsal motor nucleus of the vagus (Figure 3). These receptors are specific for PP since neither neuropeptide Y or PYY produced significant inhibition of saturable 1251_ PP binding. The identification of specific PP receptors in an area of the brain that has an incomplete BBB and contains the dorsal vagal complex provides an explanation for the indirect mechanism of action of PP. These data also explain why PP was a more potent inhibitor of centrally stimulated pancreatic exocrine secretion.
Figure 2. Schematic diagram of the pancreatic vagal system. The dorsal vagal complex contains the area postrema (AP), nucleus tractus solitarius (NTS), and dorsal motor nucleus of the vagus (DMV). This area receives input from higher centers (ie, hypothalamus), sensory input from afferent vagal fibers, and exposure to humeral factors because of an incomplete blood-brain barrier. Inhibitory agent could theoretically act on afferent fibers (1); at the dorsal vagal complex (2); on vagal efferents (3); on pancreatic ganglia (G; 4); at postgaglionic sites (5); or at other central sites (6).
from isolated pancreatic acini,31,32 suggesting that the inhibitory effects of PP are indirect. The failure to demonstrate receptors for PP on rat acinar cells is in keeping with this suggestion.31 It has been shown that PP partially inhibits potassium depolarization-stimulated cholinergic transmission in pancreatic lobules. 31 ,32 However, postganglionic inhibition of cholinergic transmission does not appear to be the major mechanism of inhibition in the intact animal. 33 The sites on which PP could act to inhibit vagal tone include the afferent vagal fibers, the dorsal vagal complex, efferent vagal fibers, and the intrapancreatic ganglia (Figure 2, sites 1-4). Recent studies in rats33 have shown that PP (and PYY) preferentially and completely inhibit pancreatic secretion stimulated centrally by 2-deoxy-D-glucose. These peptides are much less effective in blocking pancreatic secretion following stimulation of the cut vagus or stimulated directly by injection of CCK or bethanechol (thus eliminating sites 3, 4, and 5 on Figure 2). These data suggest that PP and PYY act proximal to the efferent vagus, possibly on the brain, itself. If PP does act centrally, it must either enter the brain using a specific transport system or cross the bloodbrain barrier (BBB) through one of the circumventricular organs where the BBB is incomplete.40-42
Figure 3. Autoradiographic localization of pancreatic polypeptide (PP) receptors in rat brain. (A) Nissl's stain of a sagittal section of rat brain from near midline. (bar = 5 mm). (B) Dark-field photomicrograph of the tritium-sensitive film that overlaid section from (A) for 10 days (total binding). White areas, representing areas of high concentration of 12SI_bPP binding, are observed in the area postrema (AP), nucleus tractus solitarius (NTS), dorsal motor nucleus ofthe vagus (DMV), and the interpedunclar nucleus (lPN). (C) Darkfield photomicrograph of a control section adjacent to the section shown in (B) that was incubated with 1 /Lmol/l unlabeled bPP (nonspecific binding). (D) Nissl's stain of a transverse section of the rat brain at the level of the area postrema. (bar = 1 mm). (E) Dark-field photomicrograph of the film that overlaid the section from (D) for 10 days (total binding). High concentrations of 12SI_bPP binding are present in the AP and DMV, whereas the NTS exhibited a slightly lower concentration of binding sites. (F) Dark-field photomicrograph of a control section adjacent to the section shown in (B) that was incubated with 1 /Lmol/l unlabeled bPP (nonspecific binding). November 1992 Volume 304 Number 5
Whitcomb and Taylor
brain stem in the circulation, traverses the BBB at the AP and NTS, and binds to specific receptors in the DVC. By binding to these sites, PP may directly inhibit vagal input to the pancreas (Figure 4). These observations establish a novel mechanism by which gut peptides can directly influence brain function and add a new twist to the brain-gut axis. References
Figure 4. Schematic diagram of a feedback inhibition of the vagus by pancreatic polypeptide (PP). Beginning with the cephalic phase of digestion, the dorsal motor nucleus of the vagus (DMV) becomes active, resulting in increased vagal tone. The vagus stimulates pan· creatic secretion and PP release from PP containing endocrine cells through intrapancreatic ganglia (G). Thus, PP travels via the cir· culatory system to the area postremal (AP) and nucleus tractus so· litarius (NTS), where it crosses the blood-brain barrier, binds to specific PP receptors in the dorsal vagal complex, and decreases vagal tone. Thus, PP completes a negative feedback loop to modulate vagal function.
Pancreatic polypeptide has not been identified in the mammalian brain by radioimmunoassay or immunohistochemistry,36.44.45 and thus appears to be strictly a peripheral hormone. Further, the demonstration ofPPspecific binding sites in the brain by in vitro autoradiography does not necessarily mean that circulating PP binds to these brain regions under physiologic conditions. Using an in vivo radioreceptor assay,46 we have demonstrated that PP, circulating at concentrations seen after a meal ('" 30 pM), binds to the brain region that contains the AP, NTS, and the dorsal motor nucleus of the vagus.43 Therefore, both in vivo and in vitro data suggest that the PP family of hormones can interact directly with the dorsal vagal complex. These data have led us to hypothesize that PP functions as endocrine neuromodulator by directly binding to and inhibiting the vagal nuclei in the brainstem (Figure 4). In summary, the AP and NTS are located in a brain region that has an incomplete BBB, forming a portal of entry for circulating hormones into the dorsal vagal complex. This brain region is a likely location for crosstalk between the neural and hormonal signals that modulate gastrointestinal function, including pancreatic exocrine secretion. The current data demonstrate that PP is released into the circulation by vagalcholinergic dependent mechanisms, travels to the THE AMERICAN JOURNAL OF THE MEDICAL SCIENCES
1. Solomon T: Control of exocrine pancreatic secretion, in Johnson L (ed): Physiology of the Gastrointestinal Tract, 2nd ed. New York, Raven Press, 1987, pp 1173-1207. 2. Owyang C, Williams J: Pancreatic secretion, in Yamada T (ed): Textbook of Gastroenterology, 1st ed. Philadelphia, J .B. Lippincott Co., 1991, pp 294-314. 3. Solomon TE: Regulation of pancreatic secretion. Clinics in Gastroenterology 13:657-678,1984. 4. Owyang C, Louie D, Tatum D: Feedback regulation of pancreatic enzyme secretion. J Clin Invest 77:2042, 1986. 5. Louie D, May D, Miller P, Owyang C: Cholecystokinin mediates feedback regulation of pancreatic enzyme secretion in rats. Am J Physiol 250:G252, 1986. 6. Lu L, Louie D, Owyang C: A cholecystokinin releasing peptide mediates feedback regulation of pancreatic secretion. Am J Physiol 256:G430, 1989. 7. Liddle R, Green G, Conrad C, Williams JR: Proteins but not amino acids, carbohydrates or fats stimulate cholecystokinin secretion in the rat. Am J PhysioI251:G243,1986. 8. Miyasaka K, Guan D, Liddle R, Green G: Feedback regulation by trypsin: Evidence for intraluminal CCK-releasing peptide. Am J Physiol 257:G 175-G 181, 1989. 9. Itoh Z, Honda R, Hiwatashi K: Biphasic secretory response of exocrine pancreas to feeding. Am J PhysioI238:G332-G337, 1980. 10. Beglinger C, Fried M, Whitehouse I, Jansen J, Lamers C, Gyr K: Pancreatic enzyme responses to a liquid meal and to hormonal stimulation. Correlation with plasma secretin and cholecystokinin levels. J Clin Invest 75:1471-1476,1985. 11. Dooley C, Valenzuela J: Duodenal volume and osmoreceptors in the stimulation of human pancreatic secretion. Gastroenterology 96:23, 1984. 12. Olsen 0, Schaffalitzky DE, Muckadell OB, Cantor P: Plasma secretin, plasma cholecystokinin, pancreaticobiliary secretion, and fat absorption: Effect of duodenal osmolality and polysorbate 80. Scand J GastroenteroI22:1109-1114, 1987. 13. Dyck W: Influence of intrajejunal glucose on pancreatic exocrine function in man. Gastroenterology 60:864, 1971. 14. Harper A: The control of pancreatic secretion. Gut 13:308, 1972. 15. Laugier R, Sarles H: Action of oleic acid on the exocrine pancreatic secretion of the conscious rat: Evidence for an anti -cholecystokinin-pancreozymin factor. J Physiol (£ond) 271:81-92, 1977. 16. Pappas TN, Debas HT, Taylor IL: Peptide YY: Metabolism and effect on pancreatic secretion in dogs. Gastroenterology 89:13871392,1985. 17. Pappas TN, Debas HT, Goto Y, Taylor IL: Peptide YY inhibits meal-stimulated pancreatic and gastric secretion. Am J Physiol 248:G118-G123,1985. 18. Pappas TN, Debas HT, Chang AM, Taylor IL: Peptide YY release by fatty acids is sufficient to inhibit gastric emptying in dogs. Gastroenterology 91:1386-1389, 1986. 19. Owyang C, Green L, Rader D: Colonic phase of pancreatic and biliary secretion in man. Gastroenterology 84:470, 1983. 20. Adrian TE, Besterman HS, Mallinson CN, Greenberg GR, Bloom SR: Inhibition of secretion stimulated pancreatic secretion by pancreatic polypeptide. Gut 20:37-40, 1978. 21. Lin TM, Evans DC, Chance RC, Spray GF: Bovine pancreatic peptide: Action on gastric and pancreatic secretion in dogs. Am J PhysioI232:E311-E315, 1977.
22. Taylor IL, Solomon TE, Walsh JH, Grossman MI: Pancreatic polypeptide: Metabolism and effect on pancreatic secretion in dogs. Gastroenterology 76:524-528, 1979. 23. Mulholland MW, Lally K, Taborsky GJ: Inhibition of rat pancreatic exocrine secretion by neuropeptide Y: Studies in vivo and in vitro. Pancreas 6:433-440, 1991. 24. Domschke S, Domschke W, Rosch W, Konturek SJ, Sprugel W, Mitznegg P, Wunsch E, Demling L: Inhibition by somatostatin of secretin-stimulated pancreatic secretion in man: A study with pure pancreatic juice. Scand J GastroenteroI12:59-63, 1977. 25. Konturek SJ, Tasler J, Cieszkowski M, Jaworek J, Arimiura A, Schally AW: Studies on the inhibition of pancreatic secretion by luminal somatostatin. Am J PhysioI241:GI09-G115, 1981. 26. Louie DS, Chen HT, Owyang C: Inhibition of exocrine pancreatic secretion by opiates is mediated by suppression of cholinergic transmission: Characterization of receptor subtypes. J Pharmacol Exp Ther 246:132-136,1988. 27. Miyasaka K, Funakoshi A, Kitani K, Tamamura H, Funakoshi S, Fujii N: Inhibitory effect of pancreastatin on pancreatic exocrine secretions. Pancreastatin inhibits central vagal nerve stimulation. Gastroenterology 99:1751-1756, 1990. 28. Shiratori K, Lee K, Chang T, Jo Y, Coy D, Chey W: Role of pancreatic polypeptide in the regulation of pancreatic exocrine secretin in dogs. Am J Physiol 255:G535-G541, 1988. 29. Guan D, Maouyo D, Taylor IL, Gettys TW, Greeley GJ, Morisset J: Peptide-YY, a new partner in the negative feedback control of pancreatic secretion. Endocrinology 128:911-916, 1991. 30. Sheikh SP, Roach E, Fuhlendorff J, Williams J: Localization of Yl receptors for NPY and PYY on vascular smooth muscle sells in rat pancreas. Am J PhysioI260:G250-G257, 1991. 31. Louie DS, Williams JA, Owyang C: Action of pancreatic polypeptide on rat pancreatic secretion: In vivo and in vitro. Am J Physiol 249:G489-G495, 1985. 32. Jung G, Louie DS, Owyang C: Pancreatic polypeptide inhibits pancreatic enzyme secretion via a cholinergic pathway. Am J PhysioI253:G706-G710, 1987. 33. Putnam WS, Liddle RA, Williams JA: Inhibitory regulation of rat exocrine pancreas by peptide YY and pancreatic polypeptide. Am J Physiol 256:G698-G703, 1989.
34. Taylor IL: Pancreatic polypeptide family: Pancreatic polypeptide, neuropeptide Y, and peptide YY, in Schultz ST (ed): Handbook of Physiology, The Gastrointestinal System. Bethesda, MD, American Physiological Society, 1989, pp 475-544. 35. Baetens D, Malaisse-Lagae F, Perrelet A, Orci L: Endocrine pancreas: Three·dimensional reconstruction shows two types of islets of Langerhans. Science 206:1323-1325, 1979. 36. Miyazaki K, Funakoshi A: Distribution of pancreatic polypeptidelike immunoreactivity in rat tissues. Regul Pept 21:37-43, 1988. 37. Taylor IL: Distribution and release of peptide YY in dog measured by specific radioimmunoassay. Gastroenterology 88:731737,1985. 38. Schwartz TW: Pancreatic polypeptide: A hormone under vagal contro!' Gastroenterology 85:1411-1425, 1983. 39. Taylor IL, Feldman M, Richardson CT, Walsh JH: Gastric and cephalic stimulation of human pancratic polypeptide release. Gastroenterology 75:432-437, 1978. 40. Van Houten M: Circumventricular organs: Receptors and mediators of direct peptide hormone action on brain. Advances In Metabolic Disorders 10:269-289, 1983. 41. FenstermacherJ, Gross P, Sposito N, Acuff V, Gruber K: Structure and functional variations in capillary systems within the brain. Ann N Y Acad Sci 529:21-30, 1988. 42. Pardridge WM: Neuropeptides and the blood-brain barrier. Annu Rev Physiol 45:73-82, 1983. 43. Whitcomb DC, Taylor IL, Vigna SR: Characterization of saturable binding sites for circulating pancreatic polypeptide in rat brain. Am J PhysioI259:G687-G691, 1990. 44. DiMaggio DA, Chronwall BM, Buchanan K, O'Donohue TL: Pancreatic polypeptide immunoreactivity in rat brain is actually neuropeptide Y. Neuroscience 15:1149-1157, 1985. 45. Lundberg JM, Terenius L, Hokfelt T, Tatemoto K: Comparativf! immunohistochemical and biochemical analysis of pancreatic polypeptide-like peptides with special reference to presence of neuropeptide Y in central and peripheral neurons. J Neurosci 4:2376-2386, 1984. 46. Whitcomb DC, O'Dorisio TM, Cataland S, Nishikawara MT: Theoretical basis for a new in vivo radioreceptor assay for polypeptide hormones. Am J PhysioI249:E555-E560, 1985.
November 1992 Volume 304 Number 5