Abstract
Brain-derived neurotrophic factor (BDNF) regulates diverse brain functions via TrkB receptor signaling. Due to the expression of TrkB receptors, astrocytes can internalize extracellular BDNF proteins via receptor-mediated endocytosis. Endocytosed BDNF can be re-secreted upon stimulation, but the molecular mechanism underlying this phenomenon remains unrecognized. Our study reveals that vesicle-associated membrane protein 3 (Vamp3) selectively regulates the release of endocytic BDNF from astrocytes. By using quantum dot (QD)-conjugated mature BDNF (QD-BDNF) as a proxy for the extracellular BDNF protein, we monitored the uptake, transport, and secretion of BDNF from cultured cortical astrocytes. Our data showed that endocytic QD-BDNF particles were enriched in Vamp3-containing vesicles in astrocytes and that ATP treatment sufficiently triggered either the antero- or retrograde transport and exocytosis of QD-BDNF-containing vesicles. Downregulation of Vamp3 expression disrupted endocytic BDNF secretion from astrocytes but did not affect uptake or transport. Collectively, these results provide evidence of the selective ability of astrocytic Vamp3 to control endocytic BDNF secretion during BDNF recycling.
Highlights
Brain-derived neurotrophic factor (BDNF) regulates diverse brain functions via tropomyosin-related kinase B (TrkB) receptor signaling
The intracellular uptake of quantum dot (QD)-BDNF particles into astrocytes was mediated by receptor-mediated endocytosis, as (1) QD-Bovine serum albumin (BSA) treatment resulted in no intracellular QD particles (Fig. 1C), and (2) the number of intracellular QD-conjugated mature BDNF complex (QD-BDNF) particles (Fig. 1D) from astrocytes was significantly reduced by shRNA-mediated genetic knockdown (KD) of TrkB expression (Fig. S1)
We showed the direct uptake and recycling of mature form of BDNF (mBDNF) in astrocytes by utilizing QD-BDNF as a proxy for the extracellular BDNF protein
Summary
Brain-derived neurotrophic factor (BDNF) regulates diverse brain functions via TrkB receptor signaling. Monitoring the direct uptake, transport and ATP-dependent exocytosis of endocytic mBDNF in astrocytes utilizing recombinant mBDNF proteins linked to quantum dots (QDs) revealed that Vamp[3] was selectively involved in the exocytosis of endocytic mBDNF. Our QD-linked mBDNF sensor was sufficient for examining the transport and activity-dependent secretion of endosomes, as reported p reviously[7,17,18], due to the excellent photostability and high signal-to-noise ratio of QDs in live cells. These results support the notion that mBDNF recycling in astrocytes serves as an additional source of extracellular BDNF, which is crucial for activity-dependent synaptic plasticity
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