Testosterone-Induced Matrix Metalloproteinase Activation Is a Checkpoint for Neuronal Addition to the Adult Songbird Brain

Abstract

Testosterone-induced neuronal addition to the adult songbird vocal control center, HVC, requires the androgenic induction of vascular endothelial growth factor (VEGF), followed by VEGF-stimulated angiogenesis. The expanded vasculature acts as a source of BDNF, which supports the immigration of new neurons from the overlying ventricular zone. In tumorigenesis, a similar process of adult angiogenesis is regulated by matrix metalloproteinase (MMP) activity, in particular that of the gelatinases. We therefore investigated the role of the gelatinases in neuronal addition to the HVC of adult female canaries. In situ zymography of the caudal forebrain revealed that testosterone-induced perivascular gelatinase activity that was most prominent in HVC. High-resolution gels revealed distinct MMP activities that comigrated with MMP2 and MMP9, and PCR cloning yielded MMP2 and MMP9 orthologues of 1465 and 1044 bp, respectively. Quantitative PCR revealed that HVC MMP2 mRNA levels doubled within 8 d of testosterone, whereas MMP9 transcript levels were stable. Moreover, isolated adult canary forebrain endothelial cells secreted MMP2, and VEGF substantially increased endothelial MMP2 gelatinase activity. To assess the importance of androgen-regulated, VEGF-induced MMP2 to adult angiogenesis and neurogenesis, we treated testosterone-implanted females with the gelatinase inhibitor SB-3CT. In situ zymography confirmed that SB-3CT suppressed gelatinase activity in HVC, and histological analysis revealed that SB-3CT-treated birds exhibited a decreased endothelial mitotic index and substantially diminished neuronal recruitment to HVC. These data suggest that the androgenic induction of endothelial MMP2 is a critical regulator of neuronal addition to the adult HVC, and as such comprises an important regulatory step in adult neurogenesis.