Gel shift analysis of a radioactively labeled oligonucleotide containing the CRE consensus sequence with 10 μg protein extract from the anterior hypothalamus of 12- to 13-day-old rats (lanes 2–5). Before incubation with the labeled oligonucleotide, samples were treated as follows. A, Incubation with 100-fold excess of unlabeled oligonucleotide containing the CRE consensus sequence resulted in elimination of the retarded band representing CRE-binding activity (lane 2); B, incubation with an antiserum directed against phosphorylated CREB (βpCREB) caused additional retardation with a supershifted band (lane 3, indicated by arrow in the margin); C, incubation with antiserum directed against CREB-1 (βCREB-1) resulted in nearly complete inhibition of DNA-binding activity (lane 4); D, incubation with buffer alone (lane 5).