AJP - Cell Physiology, Vol 269, Issue 2 C424-C434, Copyright © 1995 by American Physiological Society
Drugs activating G proteins disturb cycling of ADH-dependent water channels in toad urinary bladder
A. Boom, B. Flamion, M. Abramow and R. Beauwens
Laboratoire de Physiologie et de Physiopathologie, Universite Libre de Bruxelles, Belgium.
In the toad urinary bladder, antidiuretic hormone (ADH)-mediated changes in
water permeability depend on exocytic insertion and endocytic retrieval of
water channels into and from the apical membrane, respectively. Because
GTP-binding proteins (G proteins) are well-recognized regulators of
vesicular trafficking throughout the cell, we tested the hypothesis that
drugs interfering with G protein would modify the hydrosmotic response to
ADH and the ADH-regulated formation of endosomes, as assessed by luminal
incorporation of a fluid-phase marker [fluorescein isothiocyanate
(FITC)-dextran, 70 kDa]. Mastoparan (4 microM) and compound 48/80
(poly-p-methoxyphenylethylmethylamine; 50 micrograms/ml), added to the
luminal side of the toad urinary bladder, as well as AlF3 added to the
serosal side (400 microM), inhibited ADH- and 8-bromoadenosine 3',5'-cyclic
monophosphate-induced transepithelial water flow by > 50% and
simultaneously enhanced cellular incorporation of FITC-dextran by >
200%. The pattern of FITC-dextran uptake observed using fluorescence
microscopy both in scraped cells and in the intact bladder was granular,
suggesting fluid-phase endocytosis. Mastoparan and AlF3, which are both
probes of G proteins, increased FITC-dextran uptake only in the presence of
ADH and a transepithelial osmotic gradient, i.e., under conditions where
water channel-carrying endosomes presumably cycle. Therefore, we suggest
that the ADH-dependent cycling of water channels could be controlled by one
or more G proteins associated with the apical membrane and/or the water
channel-carrying vesicles.
