Solid growing tumors often show pronounced extracellular acidosis which influences intracellular signal cascades and thereby tumor phenotype. We analysed the impact of acidosis on ERK1/2 and p38 kinase in AT1 cells and compared it to non tumor cells. Cells were exposed to a control (pH 7.4) or an acidic extracellular environment (pH 6.6) for 3 h. In AT1 cells acidosis induced ERK1/2- and p38 phosphorylation without significant changes in the expression level. This was not the case in CHO- or OK-cells. Blockade of the membrane proton receptor OGR-1 with Cu2+, did not prevent the effect of acidosis but potentiated it. EGF-receptor (EGFR) signalling was not required, because inhibition of EGFR proved ineffective. Furthermore, EGFR expression in CHO-cells did not alter pH-responsiveness. Similarly, the inhibitor of protein kinase(s) C, BIM, could not prevent acidosis-induced signalling. These findings indicate that acidosis induced signaling on p38 kinase and ERK1/2 in tumor cells does not depend on the classical modules EGFR or PKC, neither seems it to be transduced by a proton receptor. Furthermore, non-tumor cells seem to respond less sensitive to acidosis. Unveiling the underlying mechanism may provide a therapeutical strategy for tumor targeting drugs.