AJP - Gastrointestinal and Liver Physiology, Vol 267, Issue 1 59-G66, Copyright © 1994 by American Physiological Society
Primary culture of salamander intestinal epithelial cells from nests: whole cell Na+ currents induced by valine
J. F. White
Department of Physiology, Emory University, Atlanta, Georgia 30322.
Methods are described for isolating the cell nests, subepithelial clusters
of germinative cells, from salamander intestinal mucosa and for growing the
nests in culture into polarized monolayers of intestinal epithelial cells.
Cells were viable in culture for up to 3 wk. The capacity of the monolayer
cells to engage in membrane transport was evaluated using the patch-clamp
technique in the whole cell mode. L-Valine (25 mM) induced an inward
current in small intestinal cells of 25.8 +/- 5.7 pA and depolarized the
cell membrane 14.5 +/- 1.6 mV. L-Alanine and L-phenylalanine were similarly
effective, whereas D-valine was ineffective. The Km of the transporter for
valine was 90 mM. Replacement of bath Na with
tris(hydroxymethyl)aminomethane eliminated the inward current induced by
valine. The basal (solute-independent) inward current was also reduced by
Na+ replacement. Glucose did not induce a Na+ current. In contrast to the
effect of valine on small intestinal cells, large intestinal cells were
unresponsive to valine. It is concluded that the cultured small intestinal
cells possess Na-amino acid but not Na-sugar cotransport. This profile of
behavior is characteristic of undifferentiated small intestinal cells.
Primary cultures of salamander small intestinal cells should be useful for
studying enterocyte function and the developmental biology of the small
intestinal mucosa.
