The anterior pituitary gland (adenohypophysis) comprises anterior and intermediate lobes (the pars distalis and pars intermedia) arising from placodal ectoderm at the anterior neural ridge. Signaling molecules including SHH, FGF, WNT, BMP and Notch are involved in regulating primordial pituitary proliferation and lineage determination. However, morphogenic events and molecular mechanisms governing anterior and intermediate lobe specification are not clear. Pituitary expression of proopiomelanocortin (POMC), the common precursor for adrenocorticotropin (ACTH) of pars distalis corticotropes and alpha-melanocyte-stimulating hormone (alpha-MSH) of pars intermedia melanotropes, provides a unique marker for anterior and intermediate lobe morphogenesis. We performed time-lapse confocal microscopy lineage tracing in live zebrafish embryos expressing GFP driven by the pomc promoter and show distinct migration pathways of POMC cells destined to the anterior and intermediate lobes. Using morpholino oligonucleotides, we show that hypomorphic FGF3 down-regulation induces specific defects of pars intermedia POMC cells while pomc, growth hormone and prolactin expression remain intact in the pars distalis. This lineage-specific process is independent of the FGF3 effect on early pituitary specifying transcription factors as indicated by normal Lim3 and Pit1 expression in hypomorphic FGF3 morphants. These findings suggest that the FGF3 signal, in addition to its previously described role of regulating progenitor proliferation and survival, delineates the melanotrope and corticotrope lineage boundary, contributing to establishment of the pituitary pars distalis and pars intermedia.