Synaptic vesicles that store monoamines and glutamate differ in protein composition.
Neuromodulators such as the monoamines are known to differ from classical neurotransmitters like glutamate in the time scale of signaling due to activation of slower G protein-coupled receptors. Recent work has suggested that the mode of release also differs between classical and modulatory transmitters. Although many components of neurotransmitter release machinery have been identified, we still understand little about the mechanisms responsible for differences in release. In this study, we address the differences between release of dopamine and glutamate by comparing the composition of synaptic vesicles (SVs) that contain the vesicular monoamine transporter 2 (VMAT2) versus vesicular glutamate transporter 2 (VGLUT2). Previous work has shown that these SV populations differ in frequency dependence, recycling kinetics and biogenesis. Taking advantage of a CRISPR-generated knock-in mouse with a cytoplasmic hemagglutinin (HA) tag at the N-terminus of VMAT2 to immunoisolate monoamine SVs, we find differences in the abundance and isoform expression of many SV protein families. Validation in primary neurons and in brain tissue confirms these differences in SV protein abundance between dopamine and glutamate release sites. Functional analysis reveals that the loss of differentially expressed SCAMP5 selectively impairs the recycling of VGLUT2 SVs, sparing VMAT2 vesicles in the same neuronal population. These findings provide new insights into the molecular diversity of SVs and the mechanisms that regulate the release of dopamine and glutamate, with implications for the physiological role of these transmitters and behavior.