cAMP stimulates apical V-ATPase accumulation, microvillar elongation, and proton extrusion in kidney collecting duct A-intercalated cells.
Kidney proton-secreting A-intercalated cells (A-IC) respond to systemic acidosis by accumulating the vacuolar ATPase (V-ATPase) in their apical membrane and by increasing the length and number of apical microvilli. We show here that the cell-permeant cAMP analog CPT-cAMP, infused in vivo, results in an almost twofold increase in apical V-ATPase accumulation in AE1-positive A-IC within 15 min and that these cells develop an extensive array of apical microvilli compared with controls. In contrast, no significant change in V-ATPase distribution could be detected by immunocytochemistry in B-intercalated cells at the acute time point examined. To show a direct effect of cAMP on A-IC, we prepared cell suspensions from the medulla of transgenic mice expressing EGFP in IC (driven by the B1-subunit promoter of the V-ATPase) and exposed them to cAMP analogs in vitro. Three-dimensional reconstructions of confocal images revealed that cAMP induced a time-dependent growth of apical microvilli, starting within minutes after addition. This effect was blocked by the PKA inhibitor myristoylated PKI. These morphological changes were paralleled by increased cAMP-mediated proton extrusion (pHi recovery) by A-IC in outer medullary collecting ducts measured using the ratiometric probe BCECF. These results, and our prior data showing that the bicarbonate-stimulated soluble adenylyl cyclase (sAC) is highly expressed in kidney intercalated cells, support the idea that cAMP generated either by sAC, or by activation of other signaling pathways, is part of the signal transduction mechanism involved in acid-base sensing and V-ATPase membrane trafficking in kidney intercalated cells.
Păunescu
Teodor G
TG
MGH Center for Systems Biology, Program in Membrane Biology, and Division of Nephrology, Massachusetts General Hospital, and Harvard Medical School, Boston, Massachusetts 02114, USA.
Ljubojevic
Marija
M
Russo
Leileata M
LM
Winter
Christian
C
McLaughlin
Margaret M
MM
Wagner
Carsten A
CA
Breton
Sylvie
S
Brown
Dennis
D
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000136:Acid-Base Equilibrium; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism; D002463:Cell Membrane Permeability; D000242:Cyclic AMP / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D007262:Infusions, Intravenous; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008871:Microvilli / Q000201:enzymology; D011401:Promoter Regions, Genetic; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021381:Protein Transport; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D013873:Thionucleotides / Q000008:administration & dosage / Q000378:metabolism*; D013997:Time Factors; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*
D001639:Bicarbonates; D047428:Protein Kinase Inhibitors; D013873:Thionucleotides; D015124:8-Bromo Cyclic Adenosine Monophosphate; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; C480628:Atp6v1b1 protein, mouse; D025262:Vacuolar Proton-Translocating ATPases; D000262:Adenylyl Cyclases