Intracellular signaling through extracellular signal-regulated kinase (ERK) is important in regulating cellular functions in a variety of tissues including the CNS. Although ERK1 and ERK2 have a very similar substrate profile and amino acid sequences, there are strikingly different phenotypes between Erk1- and Erk2-deficient mice. Thus, the question arose as to whether these two proteins are functional homologs that compensate for each other, or whether they have distinct functions. Here, we generated double knock-out mice deficient for Erk2 in the CNS, with ubiquitous homozygous deletion of Erk1, and compared the phenotypes of these mice with those of monogenic Erk2-deficient mice. Although we did obtain double knock-out newborn pups, they survived for not >1 d. These pups appeared normal just after parturition. However, they had no milk in their stomachs even 6-7 h after birth. Intracerebral hemorrhages with varying location and severity were observed. The ventricular zones and corpus callosum of the double knock-out pups did not develop adequately. Neuronal size and nuclear morphology in some brain regions were markedly aberrant in the double knock-out pups compared with controls, while deficiency in Erk2 only caused a mild phenotype. These results suggest that total ERK1/2 activity governs cellular behaviors to ensure proper brain development.