The depolarization-evoked, Ca2+-dependent release of exosomes from mouse
cortical nerve endings: new insights into synaptic transmission.
Abstract
Background and purpose Exosomes, nanosized extracellular vesicles,
emerged as players in the cell-to-cell communication in the central
nervous system (CNS), having a role in the modulation of the synaptic
activity. This study aimed at evaluating whether exosomes can be
actively released from presynaptic nerve terminals. Experimental
Approach Mouse cortical synaptosomes were exposed to a depolarizing
stimulus (25 mM KCl medium) and exosomes were isolated from the
synaptosomal supernatants. Exosomes were characterized by dynamic light
scattering, transmission electron microscopy, western blot and flow
cytometry analyses. We also evaluated whether and how removing external
Ca2+ ions or activating presynaptic GABAB receptors by exposing
synaptosomes to (±)-baclofen (10 µM) affects the 25 mM KCl-evoked
release of exosomes. Key Results The structural and biochemical analysis
unveiled that synaptosomal supernatants contained vesicles having the
size and the shape of exosomes, immunopositive for the exosomal markers
TSG101, flotillin-1, CD63 and CD9. The content of these proteins in the
exosomal fraction isolated from synaptosomal supernatants increased upon
the exposure of nerve terminals to a depolarizing stimulus and occurred
in a Ca2+-dependent fashion, mimicking the release of glutamate.
(±)-Baclofen significantly reduced glutamate exocytosis but failed to
affect the release of exosomes from cortical synaptosomes. Finally,
presynaptic exosomes were shown to carry selected NMDA and AMPA
receptors subunits. Conclusion and implications Our study unveils the
Ca2+-dependent, depolarization-evoked release of exosomes from mouse
cortical nerve terminals, which is insensitive to presynaptic
release-regulating GABAB receptors. These findings add new insights into
the mechanisms of exosomes-mediated communication in CNS.