Figure 5: Interfering with Tau N-terminal-dependent vesicle-binding rescues Tau-induced presynaptic deficits in cultured rat hippocampal neurons.

Autaptic rat hippocampal neuronal cultures were transduced with AAV viral vectors expressing GFP or Tau variants. (a) Autaptic neuronal culture transduced with AAV-EGFP is visualized by bright-field illumination (left) and GFP fluorescence (right). (b–d) Electrophysiological recordings using patch clamp in response to 10 consecutive high frequency stimulation trains (10 Hz for 10 s with 30 s interval). The representative traces are shown in (c) and the relative first EPSCs were plotted to train numbers (b,d). In (d), Tau^P301L expressing neurons were acutely treated with 5 μM Tat-NT^Tau or control Tat-mCherry peptides before patch clamp recordings. The experiments were independently repeated at least three times and the number of neurons recorded is indicated in the graph. Data present mean±s.e.m. (e) Colloidal coomassie staining of purified Tat-NT^Tau peptide; NT^Tau corresponds to the N-terminal domain (aa 1–113) of Tau. (f) Tat-NT^Tau peptide competes with full-length Tau for vesicle binding in vitro. Intact synaptic vesicles (SVs) were immobilized to Dynabeads using anti-Synaptobrevin 2 (Syb) antibodies. Following immobilization, SVs were incubated with recombinant full-length Tau (40 nM) with or without the presence of 0.1, 1.0 or 4.0 μM purified Tat-NT^Tau peptide. Immunoblots were probed for full-length Tau, Tat-NT^Tau and the synaptic vesicle marker Syb. Note that Tat-NT^Tau reduced the amount of full-length Tau bound to SVs in a dose-dependent manner.