Na(+)-K(+)-ATPase-mediated basolateral rubidium uptake in the maturing rabbit cortical collecting duct.
Within the renal cortical collecting duct (CCD), transepithelial Na(+) absorption and K(+) secretion are linked to basolateral Na(+)-K(+)-ATPase activity. Our purpose was to examine the developmental changes in basolateral Na(+)-K(+)-ATPase-mediated (86)rubidium (Rb) uptake, its inhibitor sensitivity and relationship to pump hydrolytic activity and Na(+) transport. Multiple CCDs ( approximately 6 mm) from maturing rabbits were affixed to coverslips, preincubated at 37 degrees C for 10 min (+/-1-2.5 mM ouabain or 10 or 100 micro M Schering-28080, an inhibitor of H(+)-K(+)-ATPase), and then transferred to prewarmed incubation solution containing tracer amounts of (86)Rb (+/-inhibitors). After 1 min at 37 degrees C, tubular samples were rinsed and permeabilized and isotope counts were measured to calculate basolateral Rb uptake. Ouabain-inhibitable Rb uptake, an index of basolateral Na(+)-K(+) pump activity, increased approximately 3-fold during the 1st 8 wk of postnatal life (P < 0.03). The approximately 2-fold increase in absolute rate of Rb uptake between 1 and 6 wk (2.64 +/- 0.45 to 5.02 +/- 0.32 pmol. min(-1). mm(-1)) did not reach statistical significance. The rate of basolateral Rb uptake increased further after the 6th wk of life to 7.29 +/- 0.53 pmol. min(-1). mm(-1) in adult animals (P < 0.03 vs. 6 wk). Schering-28080 failed to inhibit Rb uptake, implying that functional H(+)-K(+)-ATPase is absent at the basolateral membrane. Na(+)-K(+)-ATPase hydrolytic activity, determined by using a microassay that measured inorganic phosphate release from [gamma-(32)P]ATP under maximum velocity (V(max)) conditions, also increased in the differentiating CCD (from 316.2 +/- 44.4 pmol. h(-1). mm(-1) at 2 wk to 555.9 +/- 105.1 at 4 wk to 789.7 +/- 145.0 at 6 wk; r = 1.0 by linear regression analysis; P < 0.005). The parallel approximately 2.5-fold increases in Na(+)-K(+)-ATPase activity and ouabain-sensitive Rb uptake between 2- and 6-wk postnatal age suggest that the developmental increase in basolateral transport capacity is due predominantly to an increase in enzyme abundance. The signals mediating the developmental increase in Na(+)-K(+)-ATPase activity in the CCD remain to be defined.
Constantinescu
A R
AR
Department of Pediatrics, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, New Brunswick, New Jersey, USA.
Lane
J C
JC
Mak
J
J
Zavilowitz
B
B
Satlin
L M
LM
D000465:Algorithms; D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006868:Hydrolysis; D007093:Imidazoles / Q000494:pharmacology; D066298:In Vitro Techniques; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D009399:Nephrons / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D011817:Rabbits; D012413:Rubidium / Q000378:metabolism*; D012414:Rubidium Radioisotopes; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013696:Temperature
D004791:Enzyme Inhibitors; D007093:Imidazoles; D012414:Rubidium Radioisotopes; C035235:Sch 28080; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium