Fm. Liu et Fa. Gesek, alpha(1)-Adrenergic receptors activate NHE1 and NHE3 through distinct signaling pathways in epithelial cells, AM J P-REN, 280(3), 2001, pp. F415-F425
The Na+/H+ exchanger (NHE) regulates intracellular pH, cell volume, Na+ abs
orption and H+ secretion in epithelial cells of the renal proximal tubule (
PT). alpha (1)-Adrenergic receptors (ARs) increase NHE activity in PT cells
. The purpose of this study was to determine the mechanism of alpha (1)-AR
activation of NHE isoforms expressed in PT cells. Northern and Western blot
ting demonstrate transcripts and protein expression of NHE1 and NHE3 in PT
cells. An anti-NHE1 antibody predominately labels protein expressed at basa
l and lateral membranes. In contrast, NHE3 protein is expressed exclusively
at the apical membrane. To determine NHE isoforms regulated by alpha (1)-A
Rs, antisense oligodeoxynucleotides (AS-ODNs) specific for NHE1 and NHE3 is
oforms were introduced into cells with streptolysin O permeabilization. Cel
ls incubated with AS-ODNs a total of three times exhibited a reduction in p
rotein expression of similar to 85%. Na uptake and changes in intracellular
pH (pH(i)) were used as measures of NHE activity in PT cells. alpha (1)-AR
stimulation increased Na uptake from 8.5 to 13.8 nmol . min(-1) . mg prote
in(-1). AS-ODNs to NHE3 significantly reduced alpha (1)-AR stimulated Na up
take and increases in pHi; no effect was observed in sense-ODN-treated cell
s. Inhibition of NHE1 but not NHE3 expression abolishes amiloride-suppressi
ble NHE activity. alpha (1)-AR stimulation of NHE1 is inhibited by the prot
ein kinase C (PKC) inhibitor calphostin C whereas NHE3 activity is abolishe
d by the mitogen-activated protein kinase (MAPK) inhibitor PD-98059. In PT
cells transfected with MAPK kinase MEKK1(COOH), a truncated version of MEKK
1 that activates MAPK, NHE3 but not NHE1 activity is stimulated. We conclud
e that alpha (1)-ARs activate distinct signaling pathways to regulate speci
fic NHE isoforms localized on opposite membranes in polarized renal epithel
ial cells. alpha (1)-AR activation of NHE1 is regulated by PKC whereas NHE3
is controlled by MAPK and serves to separately regulate pH(i), Na absorpti
on, and proton excretion in PT cells.