title: H-K-ATPase activity in PNA-binding intercalated cells of newborn rabbit cortical collecting duct.
abstract: Functional and immunocytochemical studies indicate that intercalated cells in the adult rabbit cortical collecting duct (CCD) possess an H-K-adenosinetriphosphatase (H-K-ATPase). Because growing subjects must retain K+ and excrete H+, we sought to determine whether H-K-ATPase is present in the CCD early in life and, if so, to assess its activity and polarity. H-K-ATPase activity was defined as the initial rate of Sch-28080-inhibitable K+-dependent cell pH (pHi) recovery observed, in the absence of Na+, in response to an in vitro acid load. Transporter activity was assayed in intercalated cells labeled with the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein and apical cell surface marker rhodamine peanut lectin (PNA) in split-open CCDs isolated from neonatal and adult New Zealand White rabbits. In Na+-free N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered solutions (nominal absence of CO2/HCO3-), the rate of K+-dependent pH(i) recovery from a NH4Cl-induced acid load was similar in newborn (0.056 +/- 0.015 pH U/min, n = 9) and adult (0.060 +/- 0.019 pH U/min; n = 9, P = not significant) cells. This rate of K+-dependent pH(i) recovery was significantly reduced by 10-20 pM Sch-28080, an inhibitor of gastric H-K-ATPase, in both newborns (0.009 +/- 0.003 pH U/min, n = 7) and adults (0.013 +/- 0.007 pH U/min, n = 9) (P < 0.05 compared with rates in absence of inhibitor). To determine whether the location of the transporter is consistent with a role in K+ absorption and H+ secretion, pH(i) recovery of acutely acid-loaded intercalated cells in neonatal CCDs (n = 7) microperfused and bathed in the absence of Na+ and K+ was monitored after selective addition of K+ to either the luminal or basolateral membrane. Addition of 5 mM K+ led to a significantly greater rate of pH(i) recovery when it was added to the luminal rather than the peritubular solution (0.049 +/- 0.005 vs. 0.018 +/- 0.005 pH U/min, P < 0.05). We conclude that PNA-binding intercalated cells of the neonatal CCD possess H-K-ATPase activity, predominantly located in the apical membrane. This provides a mechanism for H secretion and K+ retention, processes required for growth.
authors.affiliation: Department of Pediatrics, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: A
authors.initials: A
authors.lastname: Constantinescu
authors.forename: R B
authors.initials: RB
authors.lastname: Silver
authors.forename: L M
authors.initials: LM
authors.lastname: Satlin
mesh_terms: D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016764:Cell Polarity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism*; D006859:Hydrogen / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000494:pharmacology; D066298:In Vitro Techniques; D007672:Kidney Cortex; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D037102:Lectins / Q000378:metabolism*; D018942:Macrolides; D019887:Peanut Agglutinin; D010477:Perfusion; D011188:Potassium / Q000494:pharmacology; D011817:Rabbits; D012964:Sodium / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D007093:Imidazoles; D037102:Lectins; D018942:Macrolides; D019887:Peanut Agglutinin; C035235:Sch 28080; C057620:bafilomycin A; D006859:Hydrogen; D012964:Sodium; D017506:H(+)-K(+)-Exchanging ATPase; D011188:Potassium