Figure 6.

Gradient analysis of CHT-containing vesicles. A, Sucrose velocity gradient sedimentation of whole brain LP2 fraction membranes demonstrates that CHT exists on similar vesicle populations as VAChT (boxed region) and synaptophysin. B, The synaptic vesicle-enriched fractions 5–8 (CHT and VAChT in box) from the sucrose velocity gradient in A were pooled and resolved on a self-generating iodixanol gradient. Once again, CHT and VAChT are distributed in an identical pattern that overlaps with a subset of the synaptophysin immunoreactivity. C, Glycerol velocity gradient separation of the striatal LS1 fraction yields a pattern of distribution for CHT that is indistinguishable from that of VAChT. The box highlights the close match between CHT and VAChT in fractions 3–5 that represent the peak of synaptic vesicle proteins because synaptophysin, VMAT2, VAMP2, and Rab3A also are found in these fractions, but DAT and GAT1 are excluded. Equivalent results were obtained in four separate experiments.