During antidiuresis, the cells of the renal medulla are exposed to high osmolarity and low oxygen tension. Cell survival under these harsh environmental conditions critically depends on Prostaglandin E2, synthesized by cyclooxygenase (Cox)-1 and 2. In MDCK cells, used as model-system for renal medullary cells, the expression of Cox-2 is not constitutive, but is induced under conditions of hypertonic stress. In the present study, we investigated the signal transduction pathways involved in hypertonicity-induced Cox-2 expression in MDCK cells, using various pharmacological inhibitors for signaling pathways, and found evidences for the existence of an EGFR-Raf-MAPK cascade. Inhibition of EGFR resulted in a reduced abundance of Cox-2-mRNA and -protein following osmotic stress. Various MAP kinases act downstream of the EGFR. The kinases p38, ERK1/2 and JNK are all known to be activated by hypertonicity. While inhibition of JNK had no significant effect, inhibition of ERK1/2 and p38 abolished hypertonicity-induced Cox-2 expression. Inhibition of EGFR or Cox-2 itself resulted in increased incidence of apoptosis and reduced cell survival following osmotic stress. These results indicate, that transactivation of EGFR and the MAPKs ERK1/2 and p38 plays a crucial role in Cox-2 expression and survival of MDCK cells under hypertonic conditions.