cAMP-mediated regulation of murine intestinal/pancreatic Na+/HCO3- cotransporter subtype pNBC1.
Basolateral Na(+)-HCO(3)(-) cotransport is essential for intestinal anion secretion, and indirect evidence suggests that it may be stimulated by a rise of intracellular cAMP. We therefore investigated the expression, activity, and regulation by cAMP of the Na(+)-HCO(3)(-) cotransporter isoforms NBC1 and NBCn1 in isolated murine colonic crypts. Na(+)-HCO(3)(-) transport rates were measured fluorometrically in BCECF-loaded crypts, and mRNA expression levels and localization were determined by semiquantitative PCR and in situ hybridization. Acid-activated Na(+)-HCO(3)(-) cotransport rates were 5.07 +/- 0.7 mM/min and increased by 62% after forskolin stimulation. NBC1 mRNA was more abundant in colonic crypts than in surface cells, and crypts expressed far more NBC1 than NBCn1. To investigate whether the cAMP-induced Na(+)-HCO(3)(-) cotransport activation was secondary to secretion-associated changes in HCO(3)(-) or cell volume, we measured potential forskolin-induced changes in intracellular pH and assessed Na(+)-HCO(3)(-) transport activity in CFTR -/- crypts (in which no forskolin-induced cell shrinkage occurs). We found 30% reduced Na(+)-HCO(3)(-) transport rates in CFTR -/- compared with CFTR +/+ crypts but similar Na(+)-HCO(3)(-) cotransport activation by forskolin. These studies establish the existence of an intracellular HCO(3)(-) concentration- and cell volume-independent activation of colonic NBC by an increase in intracellular cAMP.
Bachmann
O
O
Department of Internal Medicine, University of Tübingen, Germany.
Rossmann
H
H
Berger
U V
UV
Colledge
W H
WH
Ratcliff
R
R
Evans
M J
MJ
Gregor
M
M
Seidler
U
U
D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002712:Chlorides / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D003106:Colon / Q000378:metabolism*; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D015870:Gene Expression / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D017403:In Situ Hybridization; D051379:Mice; D018181:Mice, Inbred CFTR; D026902:Potassium Channel Blockers / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D012964:Sodium / Q000378:metabolism*; D027982:Sodium-Bicarbonate Symporters / Q000235:genetics / Q000378:metabolism*
D001639:Bicarbonates; D002712:Chlorides; D026902:Potassium Channel Blockers; D012333:RNA, Messenger; C498899:Slc4a4 protein, mouse; D027982:Sodium-Bicarbonate Symporters; D005576:Colforsin; D012964:Sodium; D000242:Cyclic AMP