AJP - Gastrointestinal and Liver Physiology, Vol 252, Issue 4 458-G465, Copyright © 1987 by American Physiological Society

ARTICLES

Stimulation-associated redistribution of H+-K+-ATPase activity in isolated gastric glands

T. Urushidani and J. G. Forte

The objective of this work is to establish a procedure to study the stimulation-dependent membrane redistribution and properties of H+-K+-ATPase in an in vitro model system, rabbit isolated gastric glands. Stimulated (10(-4) M histamine plus 10(-5) M forskolin) and resting (10(-4) M metiamide) glands were homogenized and fractionated into PO (40 g, 5 min), P1 (400 g, 10 min), P2 (14,500 g, 10 min), P3 (48,200 g, 90 min), and supernatant, S3. Significant changes occurred in the distribution of our marker for H+-K+-ATPase (K+-p-nitrophenyl phosphatase) activity: a reduction in activity of P3 and a compensatory increment in P1. P3 showed valinomycin (Val)-dependent vesicular H+ uptake, while H+ uptake in P1 was Val independent. Direct measurements of ATPase revealed that H+-K+-ATPase activity of P3 was Val dependent and decreased by stimulation; H+-K+-ATPase activity of P1 was Val independent and increased by stimulation. Further density gradient purification of P1 showed that membranes lighter than 17% Ficoll contained higher specific H+-K+-ATPase activity, and the observed increase in H+-K+-ATPase associated with stimulation was more pronounced. Also, the lighter fractions from stimulated P1 had much latent H+-K+-ATPase activity that was unmasked by n-octylglucoside. The properties of membrane fractions from isolated glands were consistent with results obtained in vivo: high H+-K+-ATPase activity of P3 from resting glands corresponds to cytoplasmic tubulovesicles lacking KCl transport pathways; high activity of P1 from stimulated glands corresponds to apical plasma membrane vesicles containing KCl transport in addition to the H+-K+-ATPase, and full competency for the generation of HCl.

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