Monophosphoryl lipid A (MPLA) is a detoxified derivative of LPS that induces tolerance to LPS and augments host resistance to bacterial infections. Previously we demonstrated that LPS inhibits HCO₃^- absorption in the medullary thick ascending limb (MTAL) through a basolateral TLR4-MyD88-ERK pathway. Here we examined whether pretreatment with MPLA would attenuate LPS inhibition. MTALs from rats were perfused in vitro with MPLA (1 µg/ml) in bath and lumen or bath alone for 2 hr, then LPS was added to (and MPLA removed from) the bath solution. Pretreatment with MPLA eliminated LPS-induced inhibition of HCO₃^- absorption. In MTALs pretreated with MPLA plus a phosphoinositide 3-kinase (PI3K) or Akt inhibitor, LPS decreased HCO₃^- absorption. MPLA increased Akt phosphorylation in dissected MTALs. The Akt activation was eliminated by a PI3K inhibitor and in MTALs from TLR4^-/- or TRIF^-/- mice. The effect of MPLA to prevent LPS inhibition of HCO₃^- absorption also was TRIF-dependent. Pretreatment with MPLA prevented LPS-induced ERK activation; this effect was dependent on PI3K. MPLA alone had no effect on HCO₃^- absorption and MPLA pretreatment did not prevent ERK-mediated inhibition of HCO₃^- absorption by aldosterone, consistent with MPLA's low toxicity profile. These results demonstrate that pretreatment with MPLA prevents the effect of LPS to inhibit HCO₃^- absorption in the MTAL. This protective effect is mediated directly through MPLA stimulation of a TLR4-TRIF-PI3K-Akt pathway that prevents LPS-induced ERK activation. These studies identify detoxified TLR4-based immunomodulators as novel potential therapeutic agents to prevent or treat renal tubule dysfunction in response to bacterial infections.