In support of this interpretation, the recovery of synaptic vesicle numbers following treatment with TTX (Figure 8E) is accompanied by a reduction of synapsin 1 site 2,3 phosphorylation in the DKO neurons (Figures 8I and 8J). We have used biochemical, electrophysiological, genetic, and microscopy tools to elucidate the function of dynamin isoforms in synaptic transmission. Our studies demonstrate a major function for dynamin 3 presynaptically that overlaps and synergizes with that of dynamin 1. However, whereas absence of dynamin 3 worsens the phenotype produced by the loss of dynamin 1, both at the organismal

and synaptic levels, nervous system development is not grossly affected by http://www.selleckchem.com/products/r428.html the lack of both isoforms. Neurons lacking both dynamins develop, differentiate, and establish synapses see more in vitro. Most strikingly, nerve terminals can recycle

synaptic vesicles in their absence, implying that dynamin 2 alone and/or dynamin-independent mechanisms are sufficient to support basic synaptic function. These results collectively demonstrate that neither dynamin 1 nor 3 is essential for regenerating synaptic vesicles but rather contributes to the efficiency of this process. The overlapping function of dynamin 1 and 3 in nerve terminals is supported by their similar localizations and interactions and by the more striking structural and functional defects of the presynapse observed in dynamin 1, 3 DKO neurons relative to dynamin 1 KO neurons. Furthermore, the neonatal lethal phenotype of the DKO far exceeded

the severity of the dynamin 1 single KO phenotype (Ferguson et al., 2007) in spite of the lack of an obvious phenotype in dynamin 3 KO mice. Although this genetic interaction could conceivably arise due to multiple mechanisms, our data suggest a synergistic function of dynamin 1 and 3 in synaptic vesicle endocytosis. This interpretation is further supported by the strong enrichment of dynamin 3 at presynaptic terminals of dynamin 1 KO neurons (Ferguson BCKDHB et al., 2007). Unique functions of dynamin 1 and 3 relative to dynamin 2 likely exist, including differential interactions with other proteins and perhaps phosphorylation-based regulatory mechanisms, but these functions are not essential for the basic mechanism of synaptic vesicle endocytosis. A selective enrichment of dynamin 3 in dendritic spines was reported previously (Gray et al., 2003 and Lu et al., 2007). However, we have shown here that the signal produced by the dynamin 3 antibody used in those studies is not abolished in dynamin 3 KO neurons (Figure 1E).