1 Intestinal Disease Research
Program, Faculty of Health Sciences, McMaster University, Hamilton,
Ontario, Canada L8N 3Z5;
2 Institute of Neurobiology, Evidence
suggests that stress may be a contributing factor in intestinal
inflammatory disease; however, the involved mechanisms have not been
elucidated. We previously reported that acute stress alters epithelial
physiology of rat intestine. In this study, we documented
stress-induced macromolecular transport across intestinal epithelium.
After exposure of Wistar-Kyoto rats to acute restraint stress,
transport of a model protein, horseradish peroxidase (HRP), was
assessed in isolated segments of jejunum. The flux of intact HRP was
significantly enhanced across intestine from stressed rats compared
with controls. Electron microscopy revealed HRP-containing endosomes
within enterocytes, goblet cells, and Paneth cells of stressed rats.
The number and area of HRP endosomes within enterocytes were found to
be significantly increased by stress. HRP was also visualized in
paracellular spaces between adjacent epithelial cells only in intestine
from stressed rats. Atropine treatment of rats prevented the
stress-induced abnormalities of protein transport. Our results suggest
that stress, via a mechanism that involves release of acetylcholine,
causes epithelial dysfunction that includes enhanced uptake of
macromolecular protein antigens. We speculate that immune reactions to
such foreign proteins may initiate or exacerbate inflammation.
enteric nerves; epithelium; permeability; endocytosis; tight
junctions
FOR MANY YEARS, stress and anxiety were thought to have
an etiological role in chronic inflammatory diseases and
gastrointestinal pathophysiology. It was suggested that patients with
inflammatory bowel disease (IBD) and irritable bowel syndrome have an
anxiety-prone personality profile (2, 35). Intestinal dysfunction was
also correlated with depression (39). Others argued that psychological problems occur as a result rather than a cause of chronic health problems (18). A more recent report confirmed that stressful life
events frequently precede disease relapses in patients with IBD (12).
However, the underlying mechanisms remain undefined.
Although numerous experimental studies have provided evidence that
stress is involved in causing gastric ulceration (reviewed in Ref. 15)
and altering intestinal motility (5a, 24, 25, 37), relatively few
studies have examined the effects of stress on mucosal function (4, 7,
13). The mucosa of the gastrointestinal tract consists of the lamina
propria, covered by a single cell layer of epithelial cells (mainly
transporting enterocytes, but also goblet cells, enteroendocrine cells,
and Paneth cells) joined together by tight junctions that create a
barrier restricting uptake of luminal material (21). The lamina propria
contains various immunocytes (mast cells, eosinophils, macrophages,
neutrophils, lymphocytes, etc.), and their numbers increase during
inflammation. These cells react nonspecifically to certain bacterial
products or specifically to foreign protein antigens. In addition, the mucosa is highly innervated: networks of nerves surround the crypts, and nerve fibers extend into the villi with varicosities in close proximity to the epithelium (14). Convincing evidence has been presented that enteric nerves regulate the transport function of the
epithelium (8). Signals can be communicated from the central nervous
system to the gut via extrinsic nerves or their connections to
intrinsic nerves in the plexus regions of the gut wall (38). Although
less is known about the nature of extracellular signals regulating
epithelial barrier function, it is reasonable to hypothesize that
similar neural circuits might be involved.
We previously reported (31) that stress causes intestinal mucosal
pathophysiology in Wistar-Kyoto rats, a stress-susceptible strain (26).
After exposure of rats to restraint stress, with or without cold at
8°C, jejunal preparations exhibited a secretory state, indicated by
an increased baseline short-circuit current (Isc) that was
due to net Cl The current investigation was designed to determine whether the
stress-induced epithelial barrier defect, previously identified for
small inert probes, extends to biologically relevant macromolecules such as protein antigens that might trigger an inflammatory/immune response. We examined protein uptake across isolated segments of
intestine in Ussing chambers to eliminate any possible indirect effects
of stress (e.g., changes in blood flow, motility, or mucus secretion)
that might affect protein transport in vivo. The study was focused at
the cellular level, on ultrastructural visualization of the
transepithelial transport pathway. Under carefully controlled conditions, we found that stress stimulated protein uptake via both the
transcellular and paracellular pathways. Our studies also showed that
acetylcholine released during the stress response was critical in
enhancing uptake of macromolecules across the epithelium.
Animals
ABSTRACT
Top
Abstract
Introduction
Methods
Results
Discussion
References
INTRODUCTION
Top
Abstract
Introduction
Methods
Results
Discussion
References
secretion.
Stress also resulted in increased conductance and permeability to two
small probes, mannitol and Cr-EDTA. Reduced temperature enhanced the
magnitude of the intestinal responses to stress but did not alter them
qualitatively. A subsequent study (30) demonstrated that although both
Wistar-Kyoto rats and the parent Wistar strain responded to stress with
intestinal epithelial transport abnormalities, the stress-induced
changes were more profound in Wistar-Kyoto rats, apparently due to a
defect of intestinal cholinesterase activity resulting in
hyperresponsiveness to cholinergic stimulation. In confirmation of this
hypothesis, atropine treatment of Wistar-Kyoto rats prevented the
transport abnormalities.
METHODS
Top
Abstract
Introduction
Methods
Results
Discussion
References
Ussing Chamber Studies
The jejunal segment was carefully stripped of external muscle and cut into four to eight pieces, which were returned to a large volume of 37°C oxygenated buffer. This rinsing removed any mediators that may have been released during the stripping process. The pieces were then mounted in Ussing chambers (opening of 0.6 cm2), taking care to avoid Peyer's patches. (This entire procedure was completed within 5 min and tissues were always kept under physiological conditions of temperature, pH, and oxygenation to avoid damage or deterioration that may influence macromolecular transport. With this experimental approach, tissues were viable for at least 3 h with consistent Isc responses to added agonists or field stimulation.) In the chambers, tissues were bathed in 37°C oxygenated Krebs buffer (10 ml on each side) containing (in mM) 115 NaCl, 1.25 CaCl2, 1.2 MgCl2, 2.0 K2PO4, and 25 NaHCO3, pH 7.35 ± 0.02. The serosal buffer included 10 mM glucose as an energy source, osmotically balanced by 10 mM mannitol in the mucosal (luminal) buffer. The chambers contained agar-salt bridges to monitor the potential difference (PD) across the tissues and inject Isc to maintain zero PD as measured via an automatic voltage clamp (W-P Instruments, Narco Scientific, Downsview, ON, Canada). Conductance was calculated from Ohm's law.Permeability
The inert probe, 51Cr-EDTA (6 µCi/ml) (Radiopharmacy, Chedoke-McMaster Hospital, Hamilton, ON, Canada), was added to the mucosal buffer of the Ussing chambers, balanced by an equivalent concentration of unlabeled Cr-EDTA in the serosal buffer. Samples, 1.0 ml from the serosal buffer and 0.05 ml from the mucosal buffer, were obtained at 30-min intervals or at the beginning and end of the experiment, respectively. Buffers were replaced as required to keep the volume constant. The radioactivity of 51Cr-EDTA was measured in a gamma-counter. Transepithelial fluxes were calculated by standard formulas and were expressed as nanomoles per hour per square centimeter.Protein Transport
We used horseradish peroxidase (HRP) as a model protein because it has a molecular mass of 40 kDa, similar in size to antigenic proteins known to stimulate immune responses in sensitive individuals (9). Intact HRP can be quantitatively measured using an enzymatic assay, and the reaction product is easily visualized as electron-dense material in ultrastructural studies. HRP (10Electron Microscopy
Tissues were removed at 60 and 120 min, fixed, and processed for visualization of HRP reaction product by electron microscopy. Methods for HRP product identification were modified from Graham and Karnovsky (16). The tissues were fixed in 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.4) for 2 h at 22°C, rinsed for 18 h (4°C) with 0.05 Tris buffer (pH 7.6), and then washed three times, 5 min each time. Peroxidase activity was demonstrated by incubating the tissues for 15 min in 0.5 mg/ml diaminobenzidine in 0.05 M Tris buffer (pH 7.6, 22°C) and subsequently incubating them for 15 min in the same buffer containing 0.01% H2O2. Tissues were then processed for routine transverse electron microscopy. Quantitative analysis was performed on coded high-magnification photomicrographs; 80 micrographs (i.e., at least 80 enterocytes) were evaluated per rat group (20 per rat in 4 rats per group). The number and diameter of HRP-containing endosomes were determined in 4 × 6-µm windows in the apical region (between the microvilli and the nucleus) of villus enterocytes, and the area occupied by such endosomes was calculated. Preliminary studies determined that endogenous peroxidase was not evident in epithelial cells or lamina propria of tissues from control or stressed rats.Statistical Analysis
ANOVA and subsequent Newman-Keuls analyses were used to compare groups; single comparisons were performed using Student's t-test. Pearson's coefficient was used to determine correlations. A value of P <0.05 was accepted as statistically significant. ![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Tissue Conductance and Permeability to Cr-EDTA
Compared with controls, jejunal tissues obtained from stressed rats exhibited a significantly increased (P
|
|
Protein Transport
Figure 2A illustrates the difference in fluxes of HRP among stressed and control rats and stressed rats administered atropine, in either of two salt preparations. Transport of HRP was also dramatically enhanced by stress, with the flux value approximately fourfold that across control tissues (43.6 ± 7.1 vs. 10.3 ± 2.1 pmol · h
|
Electron Microscopy
Transcellular pathway. Eighty high-power electron photomicrographs were evaluated per rat group to obtain qualitative and quantitative information on the uptake and pathway of HRP transport across the epithelium. No epithelial damage was observed in tissues from any rat group. HRP was clearly evident within endosomes in villus enterocytes of rats in all groups (Fig. 3, A-C). Endosomes appeared more dense and numerous in enterocytes from stressed rats (Fig. 3B) compared with controls (Fig. 3A) and atropine sulfate-treated stressed rats (Fig. 3C). A lower-power photomicrograph from a stressed rat (Fig. 4C) shows a large number of HRP-containing endosomes located throughout enterocytes and goblet cells. In the crypts, HRP was observed in endosomes within enterocytes (not shown) and Paneth cells and also aggregated on the microvilli of these cells (Fig. 3D).
|
|
|
Paracellular pathway. In tissues from control rats, HRP was not demonstrated in the paracellular spaces between adjacent epithelial cells (Fig. 4A). In contrast, photomicrographs from 75% of stressed rats revealed HRP within paracellular spaces and tight junctions (Fig. 4B). This finding was most evident at the 120-min time point. Paracellular transport was apparent both in the villus region, where HRP penetrated the entire length of the lateral space (Fig. 6A), and in the crypts (Fig. 6B), where the depth of HRP penetration was incomplete but clearly past the tight junctional region in many instances. HRP was never observed in the paracellular spaces in stressed rats treated with atropine sulfate. HRP fluxes were higher in tissues from rats where paracellular HRP was demonstrated; a significant correlation (r = 0.79, P = 0.002) was found between conductance values of tissues from rats in all groups and HRP fluxes. These findings suggested that the paracellular route contributed to the higher protein transport induced by stress.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Two previous studies have reported that severe physical stress (surgical trauma or burns) perturbs the intestinal barrier (6, 29). To our knowledge, our study is the first to document that a short period of relatively mild stress enhances intestinal epithelial permeability to macromolecules. Overall transport of HRP was increased by stress, and HRP was demonstrated to penetrate jejunal epithelium via both the transcellular and paracellular pathways. The number of HRP endosomes and their relative area inside enterocytes were significantly increased by restraint stress. HRP-containing endosomes were also observed in goblet cells and Paneth cells of stressed rats. HRP was visualized in the paracellular spaces between adjacent epithelial cells in stressed but not in control rats. In addition, our study implicated cholinergic mechanisms in the stress-induced epithelial abnormalities because treatment of rats with the muscarinic antagonist atropine prevented the changes in barrier function.
Our earlier studies (31) documented that Wistar-Kyoto rats react to acute stress with alterations in intestinal epithelial physiology. Ion secretion was stimulated and permeability to ions and the small inert probes, mannitol and Cr-EDTA, was enhanced. The transport abnormalities were maintained over the 3 h of the experiment and did not return to normal values until 24 h later. We also recorded a reduced Isc response to electric transmural stimulation of nerves in tissues from stressed rats. However, normal Isc responses to several secretagogues were not affected by stress, suggesting that stress induced neurotransmitter release from intestinal mucosal neurons.
In a subsequent study (30), we determined that Wistar-Kyoto rats respond to stress with epithelial abnormalities several times greater than those in Wistar rats (the parent strain). Our finding that Wistar-Kyoto rats have reduced activity of cholinesterase in intestinal mucosa suggested a role for cholinergic mechanisms in the gut abnormalities. Both atropine sulfate and atropine methyl nitrate, a quaternary salt that does not cross the blood-brain barrier, inhibited the stress-induced intestinal pathophysiology, suggesting a peripheral rather than a central location for muscarinic receptors. In contrast, hexamethonium had no effect. Those studies implicated acetylcholine released by stress in the changes in intestinal mucosal function.
In this study, we demonstrated that stress resulted in an impairment of the epithelial barrier to macromolecules and that cholinergic mechanisms were also involved in mediating this effect. Both atropine sulfate and atropine methyl nitrate treatment of rats before stress inhibited the fivefold increase in HRP flux caused by acute restraint stress. Ultrastructural analysis showed that the number and size of endosomes in a fixed region of enterocytes were significantly increased after HRP addition to the mucosal surface of tissue from stressed rats. Again, muscarinic blockade with atropine prevented the stress-induced increases. These findings are compatible with those from previous studies that have implicated nerves in the regulation of epithelial barrier function. General and specific neural blockade have been demonstrated to inhibit transepithelial transport of proteins such as bovine serum albumin and ovalbumin (11, 17). Other studies (5, 28) have shown that the cholinergic agonist carbachol increased transepithelial transport of HRP via transcellular and paracellular pathways in rat ileum. Carbachol also stimulated secretion of mucin from goblet cells (27), and stress has also been shown to induce mucus secretion (7). Therefore, cholinergic stimulation may be involved in the uptake of HRP into these cells. We found HRP bound to the apical membrane and within another secretory cell type in crypt epithelium, the Paneth cell. Our studies did not provide information on the possible mechanisms that might account for this effect.
HRP within tight junctions and paracellular spaces was observed in photomicrographs of intestinal tissues obtained from stressed rats but never in photomicrographs of tissues from control rats or stressed rats treated with atropine. The protein was visible along the entire length of the intercellular spaces between villus cells, but penetration appeared to be more limited between crypt cells, possibly due to the reduced ability of the macromolecule to enter the crypts. The presence of HRP in the paracellular regions was more evident in sections fixed at later times, suggesting gradual accumulation of HRP in this pathway. Protein in the paracellular spaces of specific tissues was associated with increased conductance, and the conductance values also correlated with HRP fluxes. Taken together, these findings suggest that stress enhances the permeability of the paracellular pathway, not only to Cr-EDTA but also to macromolecular proteins.
Tight junctions are impermeable to proteins under normal circumstances (21), although physiological regulation of tight junctional permeability to ions and small molecules is recognized. However, under certain conditions, including cholinergic stimulation with carbachol, HRP penetration of the tight junctions has been reported (5). Carbachol causes release of Ca2+ from intracellular stores and activation of protein kinase C (PKC) in cultured epithelial cells (20); activation of PKC has been implicated in loosening of tight junctions (34). Decreased resistance of tight junctions in epithelial monolayers was shown to be associated with F-actin rearrangements and phosphorylation of myosin light chain in response to T cell activation or bacterial attachment (23, 33). Stress may be an extreme situation where cholinergic stimulation in combination with other factors may result in an increase in tight junctional permeability that extends to macromolecules.
Our studies implicate cholinergic nerves in the stressed-induced epithelial pathophysiology but do not rule out other mediators or mechanisms. Evidence indicates there may also be effects via mucosal immune cells such as mast cells (7). Studies have reported that vagal stimulation can cause histamine release from mast cells in rat ileum (3). In addition, acute stress resulted in release of substance P from guinea pig airways and chronic stress caused reduced tissue levels of substance P (1), a neuropeptide that can activate immune cells, including mast cells (32). The electron photomicrographs from this study showed eosinophils and other phagocytes in the lamina propria containing HRP. Reduced density and numbers of eosinophil granules and the presence of free mast cell granules in the lamina propria suggested activation of these cells. However, it is clear that cholinergic nerves are critical in the pathway because atropine treatment of rats prevented the stress-induced epithelial barrier defect.
Our findings may be important for understanding the role of stress in allergic and inflammatory conditions. Stress may increase epithelial permeability facilitating passage of protein antigens, including food antigens and microbial toxins and products, from the gut lumen. The excessive uptake of antigens may trigger an immune response. For example, stress in sensitive individuals may increase allergen uptake and result in a local anaphylactic reaction due to mast cell activation. Release of mast cell mediators, such as histamine, may also be enhanced by cholinergic mechanisms (36) and would further increase epithelial permeability (11) and stimulate secretory activity (10). In addition, there is evidence that intracellular processing of antigens by enterocytes is necessary for T cell suppression of immune responses resulting in oral tolerance (22). Epithelial penetration of proteins via the paracellular route would avoid such protective mechanisms. The consequence of transcellular passage of proteins through cells in the epithelium, such as goblet cells, remains to be determined. Our previous finding that the degree of epithelial dysfunction induced by stress varies in different rat strains together with the results of this study suggest that genetic factors relating to cholinergic sensitivity may be important in regulating the intestinal epithelial barrier and thus determining the predisposition of an individual to stress-induced intestinal inflammation.
![]() |
ACKNOWLEDGEMENTS |
---|
The authors acknowledge the expert technical assistance of M. Benjamin, G. Scholten, and the staff of the Electron Microscopy Unit, Faculty of Health Sciences, McMaster University.
![]() |
FOOTNOTES |
---|
This research was supported by a grant from the Medical Research Council of Canada.
Address for reprint requests: M. H. Perdue, Intestinal Disease Research Program, HSC-3N5C, McMaster Univ., 1200 Main St. West, Hamilton, ON, Canada L8N 3Z5.
Received 16 June 1997; accepted in final form 24 July 1998.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Abe, T., S. Yoshihara, T. Ando, and T. Ichimura. The
influence of frequency of cold-water bathing on the substance P content
of the airways in the guinea pigs. Regul. Pept.,
1 Suppl.: S29, 1992.
2.
Almy, T. P.,
F. Kern, Jr.,
and
M. Tulin.
Alterations in colonic function in man under stress: experimental production of sigmoid spasm in healthy persons.
Gastroenterology
12:
425-436,
1949.
3.
Bani-Sacchi, T.,
M. Barattini,
S. Bianchi,
P. Blandina,
S. Brunelleschi,
R. Fantozzi,
P. F. Mannaioni,
and
E. Masini.
The release of histamine by parasympathetic stimulation in guinea-pig auricle and rat ileum.
J. Physiol. (Lond.)
371:
29-43,
1986[Abstract].
4.
Barclay, G. R.,
and
L. A. Turnberg.
Effect of psychological stress on salt and water transport in the human jejunum.
Gastroenterology
93:
91-97,
1987[Medline].
5.
Bijlsma, P. B.,
A. J. Kiliaan,
G. Scholten,
M. Heyman,
J. A. Groot,
and
J. A. J. M. Taminiau.
Carbachol but not forskolin can increase mucosal-to-serosal transport of intact protein in rat ileum in vitro.
Am. J. Physiol.
271 (Gastrointest. Liver Physiol. 34):
G147-G155,
1996
5a.
Bueno, L.,
S. M. Collins,
and
J. L. Junien.
Stress and Digestive Motility. London: Libby, 1989.
6.
Carter, E. A.,
R. G. Tompkins,
E. Schiffrin,
and
J. F. Burke.
Cutaneous thermal injury alters macromolecular permeability of rat small intestine.
Surgery
107:
335-341,
1990[Medline].
7.
Castagliuolo, I.,
J. T. LaMont,
B. Qiu,
S. M. Fleming,
K. R. Bhaskar,
S. T. Nikulasson,
C. Kornetsky,
and
C. Pothoulakis.
Acute stress causes mucin release from rat colon: role of corticotropin releasing factor and mast cells.
Am. J. Physiol.
271 (Gastrointest. Liver Physiol. 34):
G884-G892,
1996
8.
Cooke, H. J.,
and
R. A. Reddix.
Neural regulation of intestinal electrolyte transport.
In: Physiology of the Gastrointestinal Tract (3rd ed.), edited by L. R. Johnson. New York: Raven, 1994, vol. 2, p. 2083-2132.
9.
Crowe, S. E.,
and
M. H. Perdue.
Gastrointestinal food hypersensitivity: basic mechanisms of pathophysiology.
Gastroenterology
103:
1075-1095,
1992[Medline].
10.
Crowe, S. E.,
P. Sestini,
and
M. H. Perdue.
Allergic reactions of rat jejunal mucosa. Ion transport response to luminal antigen and inflammatory mediators.
Gastroenterology
99:
74-82,
1990[Medline].
11.
Crowe, S. E.,
K. Soda,
A. M. Stanisz,
and
M. H. Perdue.
Intestinal permeability in allergic rats: nerve involvement in antigen-induced changes.
Am. J. Physiol.
264 (Gastrointest. Liver Physiol. 27):
G617-G623,
1993
12.
Duffy, L. C.,
M. A. Zielezny,
J. R. Marshall,
T. E. Byers,
M. M. Weiser,
J. F. Phillips,
B. M. Calkins,
P. L. Ogra,
and
S. Graham.
Relevance of major stress events as an indicator of disease activity prevalence in inflammatory bowel disease.
Behav. Med.
17:
101-110,
1991[Medline].
13.
Empey, L. R.,
and
R. N. Fedorak.
Effect of misoprostol in preventing stress-induced intestinal fluid secretion in rats.
Prostaglandins Leukot. Essent. Fatty Acids
38:
43-48,
1989[Medline].
14.
Furness, J. B.,
and
M. Costa.
The Enteric Nervous System. New York: Churchill Livingston, 1987.
15.
Glavin, G. B.
Restraint ulcer: history, current research and future implications.
Brain Res. Bull.
5:
51-58,
1980[Medline].
16.
Graham, R. C.,
and
M. J. Karnovsky.
Glomerular permeability: ultrastructural cytochemical studies using peroxidases as protein tracers.
J. Exp. Med.
124:
1123-1134,
1966[Medline].
17.
Kimm, M. H.,
G. H. Curtis,
J. A. Hardin,
and
D. G. Gall.
Transport of bovine serum albumin across rat jejunum: role of the enteric nervous sytem.
Am. J. Physiol.
266 (Gastrointest. Liver Physiol. 29):
G186-G193,
1994
18.
Kirsner, J. B. Ulcerative colitis 1970: recent
developments. Scand. J. Gastroenterol. 6, Suppl. 6: 63-91, 1970.
19.
Lim, D. K.,
I. B. Hoskins,
R. W. Rockhold,
and
I. K. Ho.
Comparative studies of muscarinic and dopamine receptors in three strains of rat.
Eur. J. Pharmacol.
165:
279-287,
1989[Medline].
20.
Luo, H.,
R. P. Lindeman,
and
H. S. Chase, Jr.
Participation of protein kinase C in desensitization to bradykinin and to carbachol in MDCK cells.
Am. J. Physiol.
262 (Renal Fluid Electrolyte Physiol. 31):
F499-F506,
1992
21.
Madara, J. L.
Loosening tight junctions: lessons from the intestine.
J. Clin. Invest.
83:
1089-1094,
1989[Medline].
22.
Mayer, L.,
and
D. Eisenhardt.
Lack of induction of suppressor T cells by intestinal epithelial cells from patients with inflammatory bowel disease.
J. Clin. Invest.
86:
1255-1260,
1990[Medline].
23.
McKay, D. M.,
K. Croitoru,
and
M. H. Perdue.
T cell-monocyte interactions regulate epithelial physiology in a coculture model of inflammation.
Am. J. Physiol.
270 (Cell Physiol. 39):
C418-C428,
1996
24.
McRae, S.,
K. Younger,
D. G. Thompson,
and
D. L. Wingate.
Sustained mental stress alters human jejunal motor activity.
Gut
23:
404-409,
1982[Abstract].
25.
Monnikes, H.,
B. G. Schmidt,
H. E. Raybould,
and
Y. Taché.
CRF in the paraventricular nucleus mediates gastric and colonic motor response to restraint stress.
Am. J. Physiol.
262 (Gastrointest. Liver Physiol. 25):
G137-G143,
1992
26.
Pare, W. P.
Stress ulcer susceptibility and depression in Wistar Kyoto (WKY) rats.
Physiol. Behav.
46:
993-998,
1989[Medline].
27.
Phillips, T. E.
Both crypt and villus intestinal goblet cells secrete mucin in response to cholinergic stimulation.
Am. J. Physiol.
262 (Gastrointest. Liver Physiol. 25):
G327-G331,
1992
28.
Phillips, T. E.,
T. L. Phillips,
and
M. R. Neutra.
Macromolecules can pass through occluding junctions of rat ileal epithelium during cholinergic stimulation.
Cell Tissue Res.
247:
547-554,
1987[Medline].
29.
Rhodes, R. S.,
and
M. J. Karnovsky.
Loss of macromolecular barrier function associated with surgical trauma to the intestine.
Lab. Invest.
25:
220-229,
1971[Medline].
30.
Saunders, P. R.,
N. P. M. Hanssen,
D. S. Chandraratne,
and
M. H. Perdue.
Cholinergic nerves mediate stress-induced intestinal transport abnormalities in Wistar-Kyoto rats.
Am. J. Physiol.
273 (Gastrointest. Liver Physiol. 36):
G486-G490,
1997
31.
Saunders, P. R.,
U. Kosecka,
D. M. McKay,
and
M. H. Perdue.
Acute stressors stimulate ion secretion and increase epithelial permeability in rat intestine.
Am. J. Physiol.
267 (Gastrointest. Liver Physiol. 30):
G794-G799,
1994
32.
Shanahan, F.,
J. A. Denburg,
J. Fox,
J. Bienenstock,
and
A. D. Befus.
Mast cell heterogeneity: effects of the neuroenteric peptides on histamine release.
J. Immunol.
135:
1331-1337,
1985
33.
Spitz, J.,
R. Yuhan,
A. Koutsouris,
C. Blatt,
J. Alverdy,
and
G. Hecht.
Enteropathogenic Escherichia coli adherence to intestinal epithelial monolayers diminishes barrier function.
Am. J. Physiol.
268 (Gastrointest. Liver Physiol. 31):
G374-G379,
1995
34.
Stenson, W. F.,
R. A. Easom,
T. E. Riehl,
and
J. Turk.
Regulation of paracellular permeability in Caco-2 cell monolayers by protein kinase C.
Am. J. Physiol.
265 (Gastrointest. Liver Physiol. 28):
G955-G962,
1993
35.
Sullivan, A.
Psychogenic factors in ulcerative colitis.
Am. J. Dig. Dis.
2:
651-656,
1935.
36.
Wang, L.,
A. M. Stanisz,
B. K. Wershil,
S. J. Galli,
and
M. H. Perdue.
Substance P induces ion secretion in mouse small intestine through effects on enteric nerves and mast cells.
Am. J. Physiol.
269 (Gastrointest. Liver Physiol. 32):
G85-G92,
1995
37.
Wittman, T.,
F. Crenner,
F. Angel,
L. Hanusz,
C. Ringwald,
and
J. F. Grenier.
Long duration stress. Immediate and late effects on small and large bowel motility in rat.
Dig. Dis. Sci.
35:
495-500,
1990[Medline].
38.
Wood, J. D.
Enteric neurophysiology.
Am. J. Physiol.
247 (Gastrointest. Liver Physiol. 10):
G585-G598,
1984
39.
Young, S. J.,
D. H. Alpers,
C. C. Norland,
and
R. A. Woodruff.
Psychiatric illness and the irritable bowel syndrome. Practical implications for the primary physician.
Gastroenterology
70:
162-166,
1976[Medline].