Sasha Blaug, Kevin Hybiske,
Jonathan Cohn, Gary Firestone, Terry Machen and Sheldon.Miller (2001).
ENaC and CFTR dependent ion and fluid transport in mammary epithelia.
Amer. J. Physiol. 281, C633-648.
epithelial 31EG4 cells (MEC) were grown as monolayers on filters to analyze the
apical membrane mechanisms that help mediate ion and fluid transport across the
epithelium. RT-PCR showed the presence of cystic fibrosis transmembrane
conductance regulator (CFTR) and epithelial Na(+) channel (ENaC) message, and
immunomicroscopy showed apical membrane staining for both proteins. CFTR was
also localized to the apical membrane of native human mammary duct epithelium.
In control conditions, mean values of transepithelial potential (apical-side
negative) and resistance (R(T)) are -5.9 mV and 829 Omega x cm(2), respectively.
The apical membrane potential (V(A)) is -40.7 mV, and the mean ratio of apical
to basolateral membrane resistance (R(A)/R(B)) is 2.8. Apical amiloride
hyperpolarized V(A) by 19.7 mV and tripled R(A)/R(B). A cAMP-elevating cocktail
depolarized V(A) by 17.6 mV, decreased R(A)/R(B) by 60%, increased short-circuit
current by 6 microA/cm(2), decreased R(T) by 155 Omega x cm(2), and largely
eliminated responses to amiloride. Whole cell patch-clamp measurements
demonstrated amiloride-inhibited Na(+) currents [linear current-voltage (I-V)
relation] and forskolin-stimulated Cl(-) currents (linear I-V relation). A
capacitance probe method showed that in the control state, MEC monolayers either
absorbed or secreted fluid (2--4 microl x cm(-2) x h(-1)). Fluid secretion was
stimulated either by activating CFTR (cAMP) or blocking ENaC (amiloride). These
data plus equivalent circuit analysis showed that 1) fluid absorption across MEC
is mediated by Na(+) transport via apical membrane ENaC, and fluid secretion is
mediated, in part, by Cl(-) transport via apical CFTR; 2) in both cases,
appropriate counterions move through tight junctions to maintain
electroneutrality; and 3) interactions among CFTR, ENaC, and tight junctions
allow MEC to either absorb or secrete fluid and, in situ, may help control
luminal [Na(+)] and [Cl(-)].