Mast cells are found in the brain of many species. Although a considerable body of information is available concerning the development and differentiation of peripheral mast cells, little is known about brain mast cells. In the present study, the ontogeny of mast cells in the dove brain was followed by using three markers: acidic toluidine blue, alcian blue/ safranin, and an antiserum to gonadotropin-releasing hormone (GnRH). Mast cells first appear in the pia on embryonic day (E)13-14 in ovo, then along blood vessels extending from the pia into the telencephalon on posthatch day 4-5, and in the medial habenula at week 3. Medial habenular mast cell numbers increase during development, peaking in peripubertal birds, and declining thereafter. Several measures indicate that mast cells mature within the medial habenula: there is an increase in the intensity of metachromasia, a switch from alcian blue granules in young animals to mixed alcian blue and safranin granules in older animals, and an increase in GnRH-like immunoreactivity. These results were extended by using electron microscopy. The architecture of mast cell granules evolved from electron lucent with small electron dense deposits at E15 to more electron dense granules with complex patterns of internal structure by 2 months. Ultrastructural immunocytochemistry for the GnRH-like peptide at 1 month revealed both immunopositive and negative cells, suggesting that the acquisition of this phenotype is not simultaneous across the population. Thus, immature mast cells infiltrate the central nervous system and undergo in situ differentiation within the neuropil.