Transcription of the gene encoding brain-derived neurotropic factor (BDNF) is induced in response to a wide variety of extracellular stimuli via the activation of a complex array of transcription factors. However, to what degree individual transcription factors confer specificity upon the regulation of Bdnf is poorly understood. Previous studies have shown that members of the myocyte enhancer factor 2 (MEF2) transcription factor family bind a regulatory element upstream of Bdnf promoter I and associate with an unknown binding site in Bdnf promoter IV. Here we identify calcium-response element 1 as the MEF2 binding site in promoter IV of the Bdnf gene and determine the requirements for individual MEF2 family members in Bdnf regulation. MEF2A, MEF2C, and MEF2D are all highly expressed in embryonic rat cortical neurons; however, only the Mef2c gene encodes an MEF2 splice variant that lacks the γ repressor-domain. We find that MEF2C variants lacking the γ-domain are particularly sensitive to activation by membrane depolarization, raising the possibility that the MEF2s may differentially contribute to activity-regulated gene expression. We find that only knockdown of MEF2C significantly impairs membrane depolarization-induced expression of Bdnf exon IV. By contrast, knockdown of MEF2D significantly enhanced depolarization-induced expression of Bdnf exon I. Together, these data show that individual members of the MEF2 family of transcription factors differentially regulate the expression of Bdnf, revealing a new mechanism that may confer specificity on the induction of this biologically important gene.