Figure 4. Over-expressed and endogenous synuclein exert similar effects on peptide release.

(a) Sample fluorescence traces of NPY-pHluorin in cultured rodent neurons stimulated at 50 Hz for 5 s. Individual traces were fit to a plateau with single exponential decay. Events that failed to exhibit fluorescence decay (right) were scored as no decay. (b,d) The latency to decay was combined with non-parametric “no decay” data and the cumulative frequency distribution plotted. (b) Overexpression of α-synuclein (SYN) in rat neurons significantly decreased latency to decay compared to controls transfected with empty vector (con) (p < 0.0001 by Kolmogorov-Smirnov). control, n = 652 events / 5 coverslips; α-syn, n = 586 events / 8 coverslips / 2 cultures (d) Latency to decay of NPY-pHluorin events increased in neurons from synuclein TKO mice relative to wt controls (p < 0.0001). wt, n = 928 events / 10 coverslips; TKO, n = 769 events / 11 coverslips / 3 cultures (c,e) Cumulative frequency histograms for the time constants of fluorescence decay (τ_decay) by NPY-pHluorin, including events with no decay. (c) Overexpression of α-synuclein in rat neurons reduced latency to decay and τ_decay (p < 0.001 by Kolmogorov-Smirnov). (e) Loss of synuclein in neurons from TKO mice increases NPY-pHluorin τ_decay relative to neurons from wild type animals (p < 0.0001). Insets in (c) and (e) indicate mean ± SEM for the latency to decay and τ_decay for decaying events in rat (c) and mouse (e) neurons. ****, p < 0.0001 by Mann-Whitney; U = 130046 (latency) and 149764 (tau) in (c); U = 221111 (latency) and 227995 (tau) in (e)