Bumetanide increases Cl--dependent short-circuit current in late distal colon: Evidence for the presence of active electrogenic Cl- absorption.
Mammalian colonic epithelia consist of cells that are capable of both absorbing and secreting Cl-. The present studies employing Ussing chamber technique identified two opposing short-circuit current (Isc) responses to basolateral bumetanide in rat distal colon. Apart from the transepithelial Cl--secretory Isc in early distal colon that was inhibited by bumetanide, bumetanide also stimulated Isc in late distal colon that had not previously been identified. Since bumetanide inhibits basolateral Na+-K+-2Cl- cotransporter (NKCC) in crypt cells and basolateral K+-Cl- cotransporter (KCC) in surface epithelium, we proposed this stimulatory Isc could represent a KCC-mediated Cl- absorptive current. In support of this hypothesis, ion substitution experiments established Cl- dependency of this absorptive Isc and transport inhibitor studies demonstrated the involvement of an apical Cl- conductance. Current distribution and RNA sequencing analyses revealed that this Cl- absorptive Isc is closely associated with epithelial Na+ channel (ENaC) but is not dependent on ENaC activity. Thus, inhibition of ENaC by 10 μM amiloride or benzamil neither altered the direction nor its activity. Physiological studies suggested that this Cl- absorptive Isc senses dietary Cl- content; thus when dietary Cl- was low, Cl- absorptive Isc was up-regulated. In contrast, when dietary Cl- was increased, Cl- absorptive Isc was down-regulated. We conclude that an active Cl- extrusion mechanism exists in ENaC-expressing late distal colon and likely operates in parallel with ENaC to facilitate NaCl absorption.
Department of Pediatrics, University of Florida, Gainesville, FL, United States of America.
Lieqi
L
Tang
Department of Pediatrics, University of Florida, Gainesville, FL, United States of America.
Xiefan
X
Fang
Department of Pediatrics, University of Florida, Gainesville, FL, United States of America.
Steven P
SP
Winesett
Department of Pediatrics, University of Florida, Gainesville, FL, United States of America.
Catherine Y
CY
Cheng
Department of Internal Medicine, Yale School of Medicine, New Haven, CT, United States of America.
Henry J
HJ
Binder
Department of Pediatrics, University of Florida, Gainesville, FL, United States of America.
Scott A
SA
Rivkees
Department of Pediatrics, Gastroenterology, Hepatology, and Nutrition, University of Florida, Gainesville, FL, United States of America.
Sam X
http://orcid.org/0000-0003-0252-9357
SX
Cheng
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001464:Barium / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology*; D002712:Chlorides / Q000378:metabolism* / Q000494:pharmacology; D003106:Colon / Q000187:drug effects* / Q000378:metabolism*; D062686:Epithelial Sodium Channel Blockers / Q000494:pharmacology; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D005905:Glyburide / Q000494:pharmacology; D008297:Male; D009924:Organ Culture Techniques; D017207:Rats, Sprague-Dawley; D012964:Sodium / Q000378:metabolism
D002712:Chlorides; D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels; C013407:benzamil; D002034:Bumetanide; D001464:Barium; D000584:Amiloride; D012964:Sodium; D005905:Glyburide