Sodium-dependent chloride secretion across rabbit descending colon

224

100

We have used a well-differentiated human colonic cell line, the T4 cell line, as a model system to study the pathways of cellular ion transport involved in vasoactive intestinal polypeptide (VIP)-induced chloride secretion. A modified Ussing chamber was used to study transepithelial Na' and Cl-fluxes across confluent monolayer cultures of the T84 cells grown on permeable supports. In a manner analogous to isolated intestine, the Addition of VIP caused an increase of net Cl-secretion which accounted for the increase in short circuit current (1). The effect of VIP on I,, was dose dependent with a threshold stimulation at 10-10 M VIP, and a maximal effect at 10-8 M.Bumetanide prevented or reversed the response to VIP. Inhibition by bumetanide occurred promptly when it was added to the serosal, but not to the mucosal bathing media. Ion replacement studies demonstrated that the response to VIP required the simultaneous presence of Na', K+, and CF-in the serosal media. Utilizing cellular ion uptake techniques, we describe an interdependence of bumetanide-sensitive 22Na', "Rb+, and MCl-uptake, which is indicative of a Na+,K+iCl-cotransport system in this cell line. This transport pathway was localized to the basolateral membrane. Extrapolated initial velocities of uptake for each of the three ions was consistent with the electroneutral cotransport of 1 Na+:1 K+ (Rb+):2 Cl-. Our findings indicate that VIP-induced Cl-secretion intimately involves a bumetanide-sensitive Na+,K+,CF cotransport system which is functionally localized to the basolateral membrane.

Section: Discussionsupporting

confidence: 75%

Section: Introductionmentioning

confidence: 99%

Vasoactive intestinal polypeptide-induced chloride secretion by a colonic epithelial cell line. Direct participation of a basolaterally localized Na+,K+,Cl- cotransport system.

Dharmsathaphorn¹,

Mandel²,

Masui³

et al. 1985

224

100

“…Alternate explanations include the possibility that Na',K+,Cl-cotransport might not be directly regulated by prostaglandin El, but rather serves to bring Cl-into the cell in the presence of a favorable electrochemical gradient, i.e., after Clis secreted from the cells. The sensitivity of Cl-secretion to loop diuretic has been observed in a variety of epithelia (38)(39)(40)(41)(42)(43)(44) and suggests that similar CF-uptake mechanisms may be shared by many epithelial cell types that secrete CF-.…”

Section: Discussionmentioning

confidence: 99%

Chloride secretory mechanism induced by prostaglandin E1 in a colonic epithelial cell line.

Weymer¹,

Huott²,

Liu³

et al. 1985

137

Confluent T8 monolayers grown on permeable supports and mounted in a modified Ussing chamber secrete chloride (Cl-) in response to prostaglandin El. The threshold stimulation was observed at 10-' M and a maximal effect at 10' M. Unidirectional flux studies showed an increase in both serosal to mucosal and mucosal to serosal CI-fluxes with 10' M prostaglandin El; the increase in serosal to mucosal Cl-flux exceeded the increase in mucosal to serosal flux, resulting in net Cl-secretion.Na' transport was not affected in either direction and the changes in net Cla flux correlated well with the changes in short circuit current. To identify the electrolyte transport pathways involved in the Cla secretory process, the effect of prostaglandin El on ion fluxes was tested in the presence of putative inhibitors. Bumetanide was used as an inhibitor for the basolaterally localized Na',K+,Ca-cotransport system whose existence and bumetanide sensitivity have been verified in earlier studies (Dharmsathaphorn et al. 1984. J. Clin. Invest. 75:462-471). Barium was used as an inhibitor for the K+ efflux pathway on the basolateral membrane whose existence and barium sensitivity were demonstrated in this study by preloading the monolayers with "Rb+ (as a tracer for K+) and simultaneously measuring MRb+ efflux into both serosal and mucosal reservoirs. Both bumetanide and barium inhibited the net chloride secretion induced by prostaglandin El suggesting the involvement of the Na+,K+,Ca-cotransport and a K+ efflux pathways on the basolateral membrane in the Clsecretory process. The activation of another Cl-transport pathway on the apical membrane by prostaglandin El was suggested by Cl-uptake studies. Our findings indicate that the prostaglandin El-stimulated Cl-secretion, which is associated with an increase in cyclic AMP level, intimately involves (a) a bumetanide-sensitive Na+,K+,Cl-cotransport pathway that serves as a C1-uptake step across the basolateral membrane, (b) the stimulation of a barium-sensitive K+ efflux mechanism on the basolateral membrane that most likely acts to recycle K+, and (c) the activation of a Cl-transport pathway on the apical membrane that serves as a Cl-exit pathway.

“…If the chloride absorptive process was neutral sodium-chloride absorption, the removal ofchloride should inhibit sodium absorption (Table I). However, the nature ofchloride secretory process is uncertain in that most chloride transport processes are either electrogenic and sodium-dependent (e.g., cyclic AMP-stimulated chloride secretion in the rabbit distal colon [26]) or electroneutral and sodium-independent (e.g., chloride-bicarbonate exchange in the rat and rabbit distal colon [4,11]). Our results2 are not consistent with either an electrogenic and sodium-dependent or an electroneutral and sodium-independent chloride secretory process but rather with an electroneutral, sodium-dependent process which has been described in the guinea pig ileum (27).…”

Section: Discussionmentioning

confidence: 99%

Ion transport in proximal colon of the rat. Sodium depletion stimulates neutral sodium chloride absorption.

Foster¹,

Budinger²,

Hayslett³

et al. 1986

The model of sodium and chloride transport proposed for the colon is based on studies performed in the distal segment and tacitly assumes that ion transport is similar throughout the colon. In rat distal colon, neutral sodium-chloride absorption accounts for the major fraction of overall sodium absorption and aldosterone stimulates electrogenic, amiloride-sensitive sodium absorption. Since we have demonstrated qualitative differences in potassium transport in proximal and distal segments of rat colon, unidirectional 22Na and 'Cl fluxes were performed under shortcircuit conditions across isolated proximal colon of control and sodium-depleted rats with secondary hyperaldosteronism. In the control group, net sodium absorption (JN:) (7.4±0.5 Ateq/h * cm2) was greater than I,, (1.4±0.1 lueq/h cm2), and JPt was 0 in Ringer solution. Residual flux (J) was -5.2±0.5 teq/h cm2 consistent with hydrogen ion secretion suggesting that neutral sodium absorption may represent sodium-hydrogen exchange. 1 mM mucosal amiloride, which inhibits sodium-hydrogen exchange in other epithelia, produced comparable decreases in J" and JR (4.1±0.6 and 3.2±0.6 ;&eq/h * cm2, respectively) without a parallel fall in I,<. Sodium depletion stimulated JN, JA, and I., by 7.0±1.4, 6.3±1.9, and 0.8±0.2 ;eq/h cm2, respectively, and 1 mM amiloride markedly inhibited JN' and J by 6.0±1.1 and 4.0±1.6 ,ueq/h * cm2, respectively, with only a minimal reduction in I4.. Conclusions: the predominant neutral sodium-absorptive mechanism in proximal colon is sodium-hydrogen exchange. Sodium depletion stimulates electroneutral chloride-dependent sodium absorption (most likely as a result of increasing sodium-hydrogen and chloride-bicarbonate exchanges), not electrogenic chloride-independent sodium transport. The model of ion transport in the proximal colon is distinct from that of the distal colon.