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Brain-derived neurotrophic factor (BDNF) is a major neuronal growth factor that is widely expressed in the central nervous system. It is synthesized as a glycosylated precursor protein, (pro)BDNF and post-translationally converted to the mature form, (m)BDNF. BDNF is known to be produced and secreted by cortical glutamatergic principal cells (PCs); however, it remains a question whether it can also be synthesized by other neuron types, in particular, GABAergic interneurons (INs). Therefore, we utilized immunocytochemical labeling and reverse transcription quantitative PCR (RT-qPCR) to investigate the cellular distribution of proBDNF and its RNA in glutamatergic and GABAergic neurons of the mouse cortex. Immunofluorescence labeling revealed that mBDNF, as well as proBDNF, localized to both the neuronal populations in the hippocampus. The precursor proBDNF protein showed a perinuclear distribution pattern, overlapping with the rough endoplasmic reticulum (ER), the site of protein synthesis. RT-qPCR of samples obtained using laser capture microdissection (LCM) or fluorescence-activated cell sorting (FACS) of hippocampal and cortical neurons further demonstrated the abundance of BDNF transcripts in both glutamatergic and GABAergic cells. Thus, our data provide compelling evidence that BDNF can be synthesized by both principal cells and INs of the cortex.