Another possibility could be the presence of a-synuclein in a multimeric or small aggregate form which would be expected to diffuse less freely. In support of this concept the presynaptic terminal has recently been suggested to be the subcellular location where a-synuclein aggregation begins and microaggregates form. Alpha-synuclein-GFP fusion constructs have been used by many groups to study a-synuclein function in different contexts, but the effect of this construct, if any, on oligomerization in synaptic terminals in vivo is unknown. Previous work in this mouse line does demonstrate the presence of granular cytoplasmic aggregates of Syn-GFP in lysosomes but the PD-166866 (PD166866) relationship of these aggregates to those found in synucleinopathies is not clear. Which of these possibilities give rise to this discrepancy between measured asynuclein mobility between our data and that reported in the literature is not certain. Our work, however, does suggests specific differences in the mobility of a-synuclein in different subcellular compartments in mouse cortical neurons in vivo as compared to other systems that will be interesting to analyze in more detail in the future. Our measurement of the differential 17 alpha-propionate fractional recovery of fluorescence signal after photobleaching in normal terminals and high intensity terminals in vivo suggests that the amount of immobile a-synuclein is different in these two groups. Literature values reported for GFP, GFP-tagged wild-type a-synuclein and GFP-tagged a-synuclein bearing the human disease mutation A30P in presynaptic terminals of cultured hippocampal neurons are a fractional recovery of,70�C80%, similar to what we find for normal terminals in vivo. In contrast, relatively immobile GFP-tagged synapsin had a lower fractional recovery,30�C40%, which is closer to the fractional recovery we measure for high intensity terminals. In worm body wall muscle two different populations of YFP-tagged a-synuclein, one with a higher level of fractional recovery and another with a lower level have been reported.