Nerve Growth Factor Inhibits Na+/H+ Exchange and ... formula ... Absorption through Parallel Phosphatidylinositol 3-Kinase-mTOR and ERK Pathways in Thick Ascending Limb

摘要

In the medullary thick ascending limb, inhibiting the basolateral NHE1 Na⁺/H⁺ exchanger with nerve growth factor (NGF) induces actin cytoskeleton remodeling that secondarily inhibits apical NHE3 and transepithelial absorption. The inhibition by NGF is mediated 50% through activation of extracellular signal-regulated kinase (ERK). Here we examined the signaling pathway responsible for the remainder of the NGF-induced inhibition. Inhibition of absorption was reduced 45% by the phosphatidylinositol 3-kinase (PI3K) inhibitors wortmannin or and 50% by rapamycin, a specific inhibitor of mammalian target of rapamycin (mTOR), a downstream effector of PI3K. The combination of a PI3K inhibitor plus rapamycin did not cause a further reduction in the inhibition by NGF. In contrast, the combination of a PI3K inhibitor plus the MEK/ERK inhibitor U0126 completely eliminated inhibition by NGF. Rapamycin decreased NGF-induced inhibition of basolateral NHE1 by 45%. NGF induced a 2-fold increase in phosphorylation of Akt, a PI3K target linked to mTOR activation, and a 2.2-fold increase in the activity of p70 S6 kinase, a downstream effector of mTOR. p70 S6 kinase activation was blocked by wortmannin and rapamycin, consistent with PI3K, mTOR, and p70 S6 kinase in a linear pathway. Rapamycin-sensitive inhibition of NHE1 by NGF was associated with an increased level of phosphorylated mTOR in the basolateral membrane domain. These findings indicate that NGF inhibits absorption in the medullary thick ascending limb through the parallel activation of PI3K-mTOR and ERK signaling pathways, which converge to inhibit NHE1. The results identify a role for mTOR in the regulation of Na⁺/H⁺ exchange activity and implicate NHE1 as a possible downstream effector contributing to mTOR's effects on cell growth, proliferation, survival, and tumorigenesis.