We describe here the identification and molecular characterization of a new brain protein that we named SV2C because it is homologous to the synaptic vesicle proteins SV2A and SV2B, and because it is also recognized by the monoclonal SV2 antibody that led to the initial discovery of SV2A and SV2B. SV2C is more closely related to SV2A (62% identity) than to SV2B (57% identity), and contains 12 transmembrane regions similar to these proteins. To characterize SV2C and compare its properties and localization with those of SV2A and SV2B, we raised an SV2C-specific antibody. Using this antibody, we show that SV2C is an N-glycosylated protein that is concentrated on small synaptic vesicles; in addition, it is found on microvesicles in adrenal chromaffin cells. We evaluated the relative localization of the three SV2 isoforms by staining rat brain sections with antibodies specific for SV2A, SV2B and SV2C. Analysis of the resulting staining patterns confirmed previous conclusions that SV2A is ubiquitously expressed in virtually all synapses. SV2B, although more restricted in distribution, was also found in a wide variety of synapses throughout the brain. In striking contrast to this general localization and to similarly wide distributions of other synaptic vesicle proteins, SV2C was observed only in few brain areas. High levels of SV2C were found primarily in phylogenetically old brain regions such as the pallidum, the substantia nigra, the midbrain, the brainstem and the olfactory bulb. SV2C was undetectable in the cerebral cortex and the hippocampus, and found at low levels in the cerebellar cortex. Our data suggest that closely related members of a synaptic vesicle protein family can either have very general (SV2A) or restricted distributions (SV2C), possibly in order to allow specialization in the regulation of the expression or of the function of these abundant synaptic vesicle proteins.