Effect of E. coli heat-stable enterotoxin on colonic transport in guanylyl cyclase C receptor-deficient mice

Alan N. Charney1, Richard W. Egnor1, Jesline T. Alexander-Chacko1, Valentin Zaharia1, Elizabeth A. Mann2, and Ralph A. Giannella2

1 Nephrology Section, Veterans Affairs Medical Center and New York University School of Medicine, New York, New York 10010; and 2 Division of Digestive Diseases, Veterans Affairs Medical Center and University of Cincinnati College of Medicine, Cincinnati, Ohio 45267


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

We studied the functional importance of the colonic guanylyl cyclase C (GCC) receptor in GCC receptor-deficient mice. Mice were anesthetized with pentobarbital sodium, and colon segments were studied in Ussing chambers in HCO3- Ringer under short-circuit conditions. Receptor-deficient mouse proximal colon exhibited similar net Na+ absorption, lower net Cl- absorption, and a negative residual ion flux (JR), indicating net HCO3- absorption compared with that in normal mice. In normal mouse proximal colon, mucosal addition of 50 nM Escherichia coli heat-stable enterotoxin (STa) increased the serosal-to-mucosal flux of Cl- (Jsright-arrow mCl) and decreased net Cl- flux (JnetCl) accompanied by increases in short-circuit current (Isc), potential difference (PD), and tissue conductance (G). Serosal STa had no effect. In distal colon neither mucosal nor serosal STa affected ion transport. In receptor-deficient mice, neither mucosal nor serosal 500 nM STa affected electrolyte transport in proximal or distal colon. In these mice, 1 mM 8-bromo-cGMP produced changes in proximal colon Jsright-arrow mCl and JnetCl, Isc, PD, G, and JR similar to mucosal STa addition in normal mice. We conclude that the GCC receptor is necessary in the mouse proximal colon for a secretory response to mucosal STa.

sodium and chloride ion fluxes; guanosine 3',5'-cyclic monophosphate; anion secretion


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

ACTIVATION OF COLONIC guanylyl cyclase C (GCC) initiates the secretory events caused by the luminal secretion of guanylin and uroguanylin by enterocytes, specifically enterochromaffin cells (2, 6, 7, 25, 29). Escherichia coli heat-stable enterotoxin (STa), which can induce watery diarrhea, also binds to GCC, a transmembrane receptor on intestinal epithelial cells (5, 15). Agonist binding activates guanylyl cyclase, generating cGMP (11, 27). cGMP in turn may interact with cGMP-regulated phosphodiesterases or protein kinases or with cGMP-sensitive apical anion channels (34, 35). Ultimately, the cystic fibrosis transmembrane conductance regulator (CFTR) is activated, and electrogenic anion secretion results. Although the clinical consequences of STa stimulation of intestinal secretion are well known, the physiological importance of intestinal GCC itself (and its endogenous ligands) remains uncertain.

The GCC receptor has been identified along the entire small intestine and colon of the mouse (33), rat (6), opossum (13), and human (4, 22) as well as in a number of other mammals (22). However, STa binding to and activation of GCC does not mean that it is the only means of inducing intestinal secretion. For example, despite apparently similar receptor density, intestinal segments in the rat have markedly different secretory responses to STa (3, 24, 36). Whether prostaglandins or enteric nerves are involved is unclear (14, 20), but there certainly are several steps in the cGMP secretory cascade that are subject to endogenous modulation (3, 14, 20, 35).

We examined the importance of this receptor in two ways. We studied the effects of mucosal STa in GCC receptor-deficient mice. In previous studies (23, 30), GCC receptor-deficient mice exhibited a lack of secretagogue-induced intestinal secretion when examined by the suckling mouse assay. This assay is a valuable screening tool to determine whether STa causes secretion (i.e., by measuring net fluxes and the effects of blood flow, motility, and neuronal and hormonal influences). However, it does not involve a direct measurement of active colonic electrolyte transport. Such measurements in normal and knockout mice would serve to document the importance of the GCC receptor in the secretory cascade and characterize the secretory effect. We also examined whether STa could induce secretion when present on the serosal surface of the epithelium. These experiments were of particular interest in GCC receptor-deficient mice, in which upregulation of a serosal mechanism of action may make an effect easier to detect.

We were also interested in characterizing electrolyte transport in the normal mouse colon. The study of electrolyte transport has benefited greatly from the use of rat, rabbit, and guinea pig intestine. Models of altered electrolyte physiology have been developed (17, 23, 35) in the mouse, including models of gene deletion. Although electrical parameters are often measured and the in situ suckling mouse assay is often used, to date there are few data describing electrolyte transport in the normal mouse intestine. We (17) recently described the electrolyte transport pattern in the mouse distal colon in a study of the effects of acid-base variables in carbonic anhydrase II-deficient mice. Of interest was the finding that active NaCl absorption in the mouse distal colon was stimulated by changes in ambient pH rather than in CO2, the important acid-base variable in the rat colon (16). Such subtle interspecies differences underscore the importance of directly measuring electrolyte flux. The present study includes the first ion flux measurements reported in mouse proximal colon.


    MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Animals. After approval of the Veterans Affairs Subcommittee for Animal Studies, male and female mice were housed in Plexiglas cages with both food (standard chow pellets) and water available ad libitum. Mice lacking the GCC receptor were generated as previously described (23). In brief, the mouse gene encoding GCC was disrupted at the 5' end and chimeric mice were generated. Chimeras were crossed with Black Swiss mice, and the resulting heterozygous littermates (hybrids of 129/Sv and Black Swiss) were bred to produce homozygous GCC (-/-) and GCC (+/+) mice. Wild-type (normal) and homozygous knockout (GCC receptor-deficient) mice used in these experiments are the progeny of these F2 mice. The absence of GCC mRNA and protein was verified by Northern blot, RT-PCR, and Western blot analyses. Histological examination of major organ systems and arterial blood gas analyses were normal. These mice did not exhibit STa-induced secretion in the suckling mouse assay (23).

Ion flux experiments. Flux experiments were conducted as described previously (17). While mice were under pentobarbital sodium anesthesia (5 mg/100 g body wt), the entire colon was removed and rinsed with 0.9% saline. Pairs of resected, unstripped proximal and distal colons were mounted in modified Ussing half-chambers exposing a surface area of 0.385 cm2. Tissues were studied under short-circuit conditions. Periodic bipolar pulses of 0.5 mV yielded electrical current values that were used to calculate tissue conductance (G). Tissues were paired for ion flux studies only when differences in G were not >25%. The short-circuit current (Isc) divided by G yielded the active transport potential difference (PD), which was referenced to the mucosal side.

Unidirectional fluxes of Na+ and Cl- were measured by adding 2 µCi 22Na+ and 1 µCi 36Cl- (100 Ci/g specific activity, New England Nuclear, Boston, MA) to the mucosal side of each tissue pair and the serosal side of the other. Mucosal-to-serosal (Jmright-arrow s) and serosal-to-mucosal (Jsright-arrow m) fluxes (expressed in µeq · cm-2 · h-1) were measured for 16 min after an initial 30-min equilibrium period. Twelve minutes were allowed for each new steady state. Net flux (Jnet) was calculated as Jmright-arrow s - Jsright-arrow m, and the residual ion flux (JR) was equal to Isc - (JnetNa - JnetCl).

Solutions. Reagent grade chemicals were purchased from Sigma Chemical (St. Louis, MO) unless otherwise indicated. All solutions were maintained at 37°C. The HCO3- Ringer contained (in mmol/l) 10 glucose, 96 NaCl, 4 KCl, 2.4 Na2HPO4, 0.4 NaH2PO4, 1 CaSO4, 1.2 MgSO4, 21 NaHCO3, and 18 Na+ gluconate. This solution was gassed with 5% CO2-95% O2, yielding a PCO2 of 35 mmHg and a pH of 7.40.

In both GCC (+/+) and GCC (-/-) mice, pure E. coli STa (32) was added to either the mucosal or serosal bathing solution at a final concentration of 50 or 500 nM. The first dose, 50 nM, was used in the flux experiments because a dose-response analysis (between 0.25 and 500 nM) in normal mice indicated that this was the lowest concentration that produced a maximal response in Isc (Fig. 1). In GCC (-/-) mice, 8-bromo-cGMP (8-BrcGMP) at a final concentration of 1 mM was added to the serosal bathing solution as a positive control. In several GCC (+/+) mice, bumetanide was added to the serosal solution at a final concentration of 0.1 mM and then Na+ butyrate was added to both bathing solutions at a final concentration of 25 mM.


View larger version (13K):
[in this window]
[in a new window]
 
Fig. 1.   Dose-response effect of mucosal heat-stable enterotoxin (STa) on short-circuit current (Isc) in proximal and distal colon of normal mice. The result of a representative experiment is depicted. STa was added to the mucosal bathing solution at final concentrations of 0.25-500 nM. Isc was measured in proximal or distal colonic segments for a period of 28 min, and the difference between the baseline and the maximal value of Isc was plotted. The lowest concentration of STa that produced a maximal response in Isc in each segment was 50 nM.

Data analysis. All data are expressed as means ± SE and were compared by paired or unpaired Student's t-test. Two-tailed values of P < 0.05 were considered significant.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Colonic transport in GCC (+/+) mice. No differences were found in the ion flux rates across the proximal colon of male and female mice; therefore, all data were combined. The effect of mucosal 0.25-500 nM STa on the Isc of the proximal and distal colon is shown in Fig. 1. The lowest concentration that produced a maximal response in Isc in both segments was 50 nM, and this concentration was used in all flux experiments. The increase in Isc occurred immediately after STa addition and was sustained for at least 72 min.

As shown in Table 1, under control conditions GCC (+/+) mice exhibited a serosal positive PD, net Na+ absorption, and minimal net Cl- absorption in proximal colon when perfused with HCO3--Ringer. Mucosal addition of 50 nM STa had no effect on Na+ transport but reduced JnetCl to near 0 because of an increase in Jsright-arrow mCl. This was accompanied by increases in Isc, PD, and G. JR became more positive, suggesting that electrogenic HCO3- secretion was increased as well. Subsequent addition of STa to the serosal bathing solution produced no further changes in Na+ or Cl- transport or in Isc, PD, G, or JR.

                              
View this table:
[in this window]
[in a new window]
 
Table 1.   Effect of mucosal and serosal STa on proximal colon transport in GCC (+/+) normal mice

In a separate group of normal mice, STa was initially added to the serosal bathing solution. Serosal STa addition of up to 500 nM had no effect on Isc in proximal colon. As shown in Table 2, no changes in proximal colon Na+ or Cl- absorption or in Isc, PD, or JR were observed after 50 nM serosal STa, although G increased slightly. Subsequent mucosal addition of STa stimulated Jsright-arrow mCl, reduced JnetCl, and increased Isc, PD, and G. Again, JR became more positive. A small decrease in net Na+ absorption was observed because of an increase in Jsright-arrow mNa, the latter probably reflecting a time-dependent increase in the passive permeability to Na+. These effects were similar to those observed in tissues that were exposed to mucosal STa in the absence of serosal STa.

                              
View this table:
[in this window]
[in a new window]
 
Table 2.   Effect of serosal and mucosal STa on proximal colon transport in GCC (+/+) normal mice

In the distal colon, addition of up to 250 nM mucosal or serosal STa had no effect on measured ion fluxes (data not shown). Mucosal STa at 500 nM increased Isc minimally (by <0.5 µeq · cm-2 · h-1), and serosal STa at 500 nM had no effect on Isc.

To determine whether active Cl- secretion was present in the proximal or distal colon under basal conditions, we measured unidirectional Cl- fluxes after serosal addition of bumetanide. In proximal colon, bumetanide did not inhibit Jsright-arrow mCl [7.8 ± 0.4 vs. 7.0 ± 0.3 µeq · cm-2 · h-1; n = 6; P = not significant (NS)]. Subsequent addition of Na+ butyrate, which inhibits basal Cl- secretion even in those tissues not affected by bumetanide (3), also had no effect on Jsright-arrow mCl. In the distal colon, neither bumetanide nor butyrate reduced Jsright-arrow mCl (results with control, bumetanide, and butyrate were 10.4 ± 0.3, 10.4 ± 0.7, and 10.1 ± 0.5 µeq · cm-2 · h-1, respectively; n = 6; P = NS). These inhibitors did not affect Isc, PD, or G in the proximal or distal colon.

Colonic transport in GCC (-/-) mice. Table 3 shows the basal flux rates and effects of mucosal STa in GCC receptor-deficient mice. When perfused with HCO3--Ringer under control conditions, GCC (-/-) mice exhibited a serosal positive PD and net Na+ absorption in proximal colon similar to GCC (+/+) mice. However, net Cl- absorption was 0 compared with minimal absorption in normal mice [-0.9 ± 0.8 (n = 11) vs. 1.0 ± 0.5 µeq · cm-2 · h-1 (n = 20 derived from Tables 1 and 2); P < 0.03], and a negative JR was observed, indicating net HCO3- absorption [-3.4 ± 0.9 (n = 11) vs. -0.4 ± 0.8 µeq · cm-2 · h-1; n = 20; P < 0.02].

                              
View this table:
[in this window]
[in a new window]
 
Table 3.   Effects of mucosal STa and cGMP on proximal colon transport in GCC (-/-) receptor-deficient mice

Mucosal addition of 50 nM STa in GCC (-/-) mice did not affect Cl- fluxes but caused a small increase in Jsright-arrow mNa and G and a decrease in JnetNa. Isc, PD, and JR were unaffected. In fact, mucosal concentrations of STa up to 500 nM did not increase Isc in these tissues. Subsequent addition of 8-BrcGMP as a positive control stimulated Jsright-arrow mCl, causing net Cl- secretion, and increased Isc, PD, and G. JR became positive, indicating that HCO3- secretion was now present. JnetNa was reduced to near 0 primarily because of an increase in Jsright-arrow mNa.

As in normal mice, serosal addition of up to 500 nM STa in GCC (-/-) mice had no effect on Isc in the proximal colon. Subsequent addition of 8-BrcGMP again increased Jsright-arrow mCl, stimulated Cl- secretion, and caused increases in Isc, PD, and G (data not shown). In the distal colon, addition of up to 500 nM mucosal or serosal STa had no effect on Isc.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

The development of genetically altered mice has increased the use of mouse intestine in physiological research. The control data in this study are the first measures of active electrolyte transport in murine proximal colon. Together with our recent publication describing (17) the mouse distal colon, a more complete picture of electrolyte transport has emerged. Both proximal and distal colon exhibit a lumen-positive PD and net Na+ absorption when perfused with HCO3--Ringer. Like human, rat, and rabbit colon, the mouse colon has a proximal-to-distal gradient of conductance, although it appears to be somewhat more permeable than these tissues (3, 31). Of interest is that mouse distal colon absorbs Cl- at a greater rate than the proximal colon. It is unclear whether this reflects the experimental conditions or whether there are fewer or different Cl- absorptive processes along the apical membranes of proximal colonic epithelial cells.

The values for Na+ transport and various electrical parameters were similar in normal and GCC receptor-deficient mice. However, net Cl- absorption was lower in GCC (-/-) mice than in normal mice, and a negative JR was observed (indicating net HCO3- absorption). These differences most likely reflect a lower level of apical membrane Cl-/HCO3- exchange in receptor-deficient mice. Another possibility is that a basal HCO3- secretory process, normally active, is absent or reduced in GCC (-/-) mice. Guanylin is secreted by mouse colonic mucosa, and active Cl- and HCO3- secretion could be present in normal mice under basal conditions. However, if a basal anion secretory process is absent, JnetCl should be greater, not smaller, in GCC (-/-) mice than in GCC (+/+) mice (because of a lower Jsright-arrow mCl).

Indeed, when basal secretion was tested for in normal mice with bumetanide and butyrate, no evidence for active, basal Cl- secretion was found in either proximal or distal colon. Bumetanide inhibits Na+-K+-2Cl- cotransport and thereby inhibits Cl- entry across the basolateral membrane (1). Butyrate appears to compete for apical anion channels in intestinal epithelia (10). By the use of these same inhibitors, rat distal colon (but not proximal colon) exhibits active, basal Cl- secretion (3, 10). An increase in intracellular HCO3- concentration also inhibits active Cl- secretion in rat distal colon (9) but does not affect basal Jsright-arrow mCl in mouse distal colon (17), confirming the present findings.

Another finding in this study is the confirmation that GCC receptor-deficient mice do not respond to mucosal exposure to STa. Although the suckling mouse assay was initially used to confirm the gene disruption that generated these mice (23, 30), this assay does not involve measurement of unidirectional or active electrolyte transport. In addition, there is a minimal but reproducible degree of specific binding of STa to colonic mucosal cell membranes in GCC (-/-) mice (23). This suggests the presence of an apical membrane receptor other than GCC and/or a serosal membrane receptor (see below). However, our data indicate that GCC (-/-) mice do not exhibit any of the STa-induced changes in colonic Isc, PD, G, Jsright-arrow mCl, or JnetCl that are typical of normal mice. This finding appears to exclude a second apical membrane receptor and the possibility that prostaglandin release mediates a portion of the secretory response in the mouse, as has been found in pig colon (20). Because mucosal nerves were present in our unstripped mouse preparation, a role for the enteric nervous system, which may be significant in rat colon (36), also appears unlikely. Our data clearly indicate that mucosal STa cannot access any secretory step between guanylyl cyclase activation and anion flux across the apical membrane other than the GCC receptor itself and, at least in vitro, no other secretory pathway.

We also tested for the integrity of the secretory pathway beyond the GCC receptor. The addition of cGMP in GCC (-/-) mice produced a secretory effect identical to STa in normal mice and confirmed that the gene product deletion was limited to the receptor alone. Two other deletions elsewhere in the secretory pathway have been described. Humans with cystic fibrosis (CF) and transgenic CF mice both lack the CFTR anion channel, and their colons are nearly or completely unresponsive to mucosal STa and guanylin, respectively (8, 18). However, the duodena of these CF patients and CFTR knockout mice do respond to STa or uroguanylin. The patients have normal increases in net HCO3- secretion and Isc in response to 100 nM mucosal STa, and the mice exhibit small but reproducible increases in Isc at uroguanylin concentrations between 1 and 10 µM (21, 26).

The jejuna of cGMP-dependent protein kinase II-deficient mice also have a small but significant Isc response to (100 nM) mucosal STa, although not to cGMP (35). In contrast with the models of anion channel deletion described above, the proximal colons of these animals have a nearly normal response to STa both in vivo and in vitro. These findings apparently indicate the existence of a colonic (and possibly small intestinal) secretory pathway independent of this kinase, perhaps involving a cGMP-inhibitable phosphodiesterase or activation of a cAMP-dependent kinase (35). Although our findings in GCC (-/-) mice do not have a bearing on this possibility, they do suggest that some steps in this secretory pathway can be bypassed and others cannot. These three models (with specific deficiencies in the receptor, protein kinase, or anion channel) offer unique opportunities to examine this secretory mechanism and its relation to other transport pathways, absorptive and secretory (28).

Finally, our data suggest that STa has no effect on proximal or distal colonic electrolyte transport when present on the serosal surface. The finding has meaning because, as noted above, there is a minimal degree of specific binding of STa to colonic mucosal cell membranes in GCC (-/-) mice (23). Moreover, uroguanylin, which activates GCC when present on the luminal surface of intestinal epithelial cells, also is secreted into the bloodstream. On reaching the kidneys, uroguanylin can cause natriuresis and kaliuresis (12, 19). This suggests that uroguanylin may have two functions: aiding the digestive process by causing luminal fluid secretion as chyme passes along the bowel and signaling the kidney as intestinal absorption proceeds to expedite Na+ and K+ excretion. Although it is doubtful that a serosal GCC receptor and second signaling pathway exist in the mouse colon, serosal activation of guanylyl cyclase (or another secretory pathway) may be present in other tissues or species.

In conclusion, we have described the transport pattern in normal mouse proximal colon, which is similar but not identical to the rat, rabbit, and human colon. We also found that the proximal colon of GCC (-/-) mice does not respond to mucosal STa, although the remainder of this cGMP-stimulated secretory pathway is intact. Neither normal nor receptor-deficient mice respond to serosal STa. The GCC (-/-) mouse and other models of genetically engineered breaks in the cGMP secretory pathway are valuable tools to study the pathogenesis of intestinal secretion.


    ACKNOWLEDGEMENTS

This material is based on work supported by the Office of Research and Development, Medical Research Service, Department of Veterans Affairs.


    FOOTNOTES

Address for reprint requests and other correspondence: A. N. Charney, Nephrology Section, VA Medical Center, 423 E. 23rd St., New York, NY 10010.

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 8 June 2000; accepted in final form 1 September 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Cabantchik, ZI, and Greger R. Chemical probes for anion transporters of mammalian cell membranes. Am J Physiol Cell Physiol 262: C803-C827, 1992[Abstract/Free Full Text].

2.   Cetin, Y, Kuhn M, Kulaksiz H, Adermann K, Bargsten G, Grube D, and Forssmann WG. Enterochromaffin cells of the digestive system: cellular source of guanylin, a guanylate cyclase-activating peptide. Proc Natl Acad Sci USA 91: 2935-2939, 1994[Abstract].

3.   Charney, AN, Giannella R, and Egnor RW. Effects of short-chain fatty acids on cyclic-3',5'-guanosine monophosphate-mediated colonic secretion. Comp Biochem Physiol B Biochem Mol Biol 124: 169-178, 1999[ISI].

4.   Cohen, MB, Guarino A, Shukla R, and Giannella RA. Age-related differences in receptors for Escherichia coli heat-stable enterotoxin in the small and large intestine of children. Gastroenterology 94: 367-373, 1988[ISI][Medline].

5.   Cohen, MB, Moyer MS, Luttrell M, and Giannella R. The immature rat small intestine exhibits an increased sensitivity and response to Escherichia coli heat stable enterotoxin. Pediatr Res 20: 555-560, 1986[Abstract].

6.   Cohen, MB, Witte DP, Hawkins JA, and Currie MG. Immunohistochemical localization of guanylin in the rat small intestine and colon. Biochem Biophys Res Commun 209: 803-808, 1995[ISI][Medline].

7.   Currie, MG, Fok KF, Kato J, Moore RJ, Hamra FK, Duffin KL, and Smith CE. Guanylin: an endogenous activator of intestinal guanylate cyclase. Proc Natl Acad Sci USA 89: 947-951, 1992[Abstract].

8.   Cuthbert, AW, Hickman ME, MacVinish LJ, Evans MJ, Colledge WH, Ratcliff R, Seale PW, and Humphrey PP. Chloride secretion in response to guanylin in colonic epithelial from normal and transgenic cystic fibrosis mice. Br J Pharmacol 112: 31-36, 1994[Abstract].

9.   Dagher, PC, Balsam L, Weber JT, Egnor RW, and Charney AN. Modulation of chloride secretion in the rat colon by intracellular bicarbonate. Gastroenterology 103: 120-127, 1992[ISI][Medline].

10.   Dagher, PC, Egnor RW, Taglietta-Kohlbrecher A, and Charney AN. Short-chain fatty acids inhibit cAMP-mediated chloride secretion in rat colon. Am J Physiol Cell Physiol 271: C1853-C1860, 1996[Abstract/Free Full Text].

11.   Field, M, Graf LH, Laird WJ, and Smith PL. Heat-stable enterotoxin of Escherichia coli: in vitro effects on guanylate cyclase activity, cyclic GMP concentration, and ion transport in small intestine. Proc Natl Acad Sci USA 75: 2800-2804, 1978[Abstract].

12.   Forte, LR, Fan X, and Hamra FK. Salt and water homeostasis: uroguanylin is a circulating peptide hormone with natriuretic activity. Am J Kidney Dis 28: 296-304, 1996[ISI][Medline].

13.   Forte, LR, Krause WJ, and Freeman RH. Escherichia coli enterotoxin receptors: localization in opossum kidney, intestine, and testis. Am J Physiol Renal Fluid Electrolyte Physiol 257: F874-F881, 1989[Abstract/Free Full Text].

14.   Gazzano, H, Wu HE, and Waldman SA. Activation of particulate guanylate cyclase by Escherichia coli heat-stable enterotoxin is regulated by adenine nucleotides. Infect Immun 59: 1552-1557, 1991[ISI][Medline].

15.   Giannella, R, Luttrell M, and Thompson M. Binding of Escherichia coli heat-stable enterotoxin to receptors on rat intestinal cells. Am J Physiol Gastrointest Liver Physiol 245: G492-G498, 1983[Abstract/Free Full Text].

16.   Goldfarb, DS, Egnor RW, and Charney AN. Effects of acid-base variables on ion transport in rat colon. J Clin Invest 81: 1903-1910, 1988[ISI][Medline].

17.   Goldfarb, DS, Sly WS, Waheed A, and Charney AN. Acid-base effects on electrolyte transport in CAII-deficient mouse colon. Am J Physiol Gastrointest Liver Physiol 278: G409-G415, 2000[Abstract/Free Full Text].

18.   Goldstein, JL, Sahi J, Bhuva M, and Layden TJ. Escherichia coli heat stable enterotoxin-mediated colonic Cl- secretion is absent in cystic fibrosis. Gastroenterology 107: 950-956, 1994[ISI][Medline].

19.   Greenberg, RN, Hill M, Crytzer J, Krause WJ, Eber SL, Hamra FK, and Forte LR. Comparison of effects of uroguanylin, guanylin, and Escherichia coli heat-stable enterotoxin STa in mouse intestine and kidney: evidence that uroguanylin is an intestinal natriuretic hormone. J Investig Med 45: 276-283, 1997[ISI][Medline].

20.   Hayden, UL, Greenberg RN, and Carey HV. Role of prostaglandins and enteric nerves in Escherichia coli heat-stable enterotoxin (STa)-induced intestinal secretion in pigs. Am J Vet Res 57: 211-215, 1996[ISI][Medline].

21.   Joo, NS, London RM, Kim HD, Forte LR, and Clark LL. Regulation of intestinal Cl- and HCO3- secretion by uroguanylin. Am J Physiol Gastrointest Liver Physiol 274: G633-G644, 1998[Abstract/Free Full Text].

22.   Krause, WJ, Cullingford GL, Freeman RH, Eber SL, Richardson KC, Fok KF, Currie MG, and Forte LR. Distribution of heat-stable enterotoxin/guanylin receptors in the intestinal tract of man and other mammals. J Anat 184: 407-417, 1994[ISI][Medline].

23.   Mann, EA, Jump ML, Wu J, Yee E, and Giannella RA. Mice lacking the guanylyl cyclase C receptor are resistant to STa-induced secretion. Biochem Biophys Res Commun 239: 463-466, 1997[ISI][Medline].

24.   Nobles, M, Diener M, and Rummel W. Segment-specific effects of the heat-stable enterotoxin of E. coli on electrolyte transport in the rat colon. Eur J Pharmacol 202: 201-211, 1991[ISI][Medline].

25.   Perkins, A, Goy MF, and Li Z. Uroguanylin is expressed by enterochromaffin cells in the rat gastrointestinal tract. Gastroenterology 113: 1007-1014, 1997[ISI][Medline].

26.   Pratha, VS, Hogan DL, Martensson BA, Bernard J, Zhou R, and Isenberg JI. Identification of transport abnormalities in duodenal mucosa and duodenal enterocytes from patients with cystic fibrosis. Gastroenterology 118: 1051-1060, 2000[ISI][Medline].

27.   Rao, MC, Guandalini S, Smith PL, and Field M. Mode of action of heat-stable Escherichia coli enterotoxin. Biochim Biophys Acta 632: 35-46, 1980[ISI][Medline].

28.   Schulz, S. Targeted gene disruption in the development of mouse models to elucidate the role of receptor guanylyl cyclase signaling pathways in physiological function. Methods 19: 551-558, 1999[ISI][Medline].

29.   Schulz, S, Green CK, Yuen PST, and Garbers DL. Guanylyl cyclase is a heat-stable enterotoxin receptor. Cell 63: 941-948, 1990[ISI][Medline].

30.   Schulz, S, Lopez MJ, Kuhn M, and Garbers DL. Disruption of the guanylyl cyclase-c gene leads to a paradoxical phenotype of viable but heat-stable enterotoxin-resistant mice. J Clin Invest 100: 1590-1595, 1997[Abstract/Free Full Text].

31.   Sellin, JH, and DeSoignie R. Rabbit proximal colon: a distinct transport epithelium. Am J Physiol Gastrointest Liver Physiol 246: G603-G610, 1984[Abstract/Free Full Text].

32.   Staples, SJ, Asher SE, and Giannella RA. Purification and characterization of heat-stable enterotoxin produced by a strain of E. coli pathogenic for man. J Biol Chem 255: 4716-4721, 1980[Abstract/Free Full Text].

33.   Swenson, ES, Mann EA, Jump ML, Witte DP, and Giannella R. The guanylin/STa receptor is expressed in crypts and apical epithelium throughout the mouse intestine. Biochem Biophys Res Commun 225: 1009-1014, 1996[ISI][Medline].

34.   Vaandrager, AB, Bot AGM, and de Jonge HR. Guanosine 3',5'-cyclic monophosphate-dependent protein kinase II mediates heat-stable enterotoxin-provoked chloride secretion in rat intestine. Gastroenterology 112: 437-443, 1997[ISI][Medline].

35.   Vaandrager, AB, Bot AGM, Ruth P, Pfeifer A, Hofmann F, and de Jonge HR. Differential role of cyclic GMP-dependent protein kinase II in ion transport in murine small intestine and colon. Gastroenterology 118: 108-114, 2000[ISI][Medline].

36.   Young, A, and Levin RJ. Segment heterogeneity of rat colonic electrogenic secretion in response to the bacterial enterotoxin Escherichia coli STa in vitro. Exp Physiol 76: 979-982, 1991[Abstract].


Am J Physiol Gastrointest Liver Physiol 280(2):G216-G221