Figure 6: Aβ disrupts the retrograde barrier of tau by destabilizing MT in the AIS and the integrity of AIS by mislocalizing ankG.
From: HDAC6 and RhoA are novel players in Abeta-driven disruption of neuronal polarity
(a) Confocal image of a neuron electroporated with HDAC6-FLAG and EB3-GFP, showing the region corresponding to the AIS (white rectangle). Scale bar, 10 μm. (b) Representative kymographs of the AIS of neurons co-electroporated with EB3-GFP and FLAG (left panels) or HDAC6-FLAG (right panels) and treated with 0.05% DMSO (ctrl), 100 nM Aβ (Aβ) or 10 μM tubacin (tubacin). The vertical arrow indicates the time (90 s) and the horizontal arrow indicates the direction from the cell body to the growth cone. (c) Quantification of the velocity of EB3-GFP comets in the AIS in neurons co-electroporated with EB3-GFP/FLAG (FLAG) and treated with either 0.05% DMSO (ctrl, 0.17±0.01, n=21), 100 nM Aβ for 1 h (Aβ, 0.24±0.01, n=11), 10 μM tubacin for 1 h (tubacin, 0.22±0.02, n=10) or electroporated with EB3-GFP/HDAC6-FLAG (HDAC6) and treated with 0.05% DMSO (ctrl, 0.14±0.01, n=11) or 100 nM Aβ for 1 h (Aβ, 0.17±0.02, n=12). Statistical significance determined by one-way ANOVA, ***P=0.001 (FLAG ctrl/FLAG Aβ), **P=0.019 (FLAG ctrl/FLAG tubacin), *** P<0.001 (HDAC6 ctrl/FLAG Aβ), **P=0.005 (HDAC6 ctrl/FLAG tubacin), **P=0.011 (FLAG Aβ/HDAC6 Aβ). (d) Immunofluorescence analysis of ankG and tubulin-βIII localization in neurons treated with either 0.05% DMSO (ctrl), 100 nM Aβ for 4 h (Aβ) or 10 μM tubacin (tubacin). Scale bar, 25 μm. (e) Quantification of the length of ankG staining along the axon of neurons treated with either 0.05% DMSO (ctrl, 56.78±4.41, n=52), 100 nM Aβ (Aβ, 74.49±5.94, n=68) or 10 μM tubacin (81.57±3.67, n=42). Statistical significance determined by one-way ANOVA, **P=0.006 (ctrl versus Aβ, ctrl versus tubacin). (f) Somatic current-clamp recording of a representative action potential evoked in neurons at 14 DIV treated for 4 h with 0.05% DMSO (black) or 100 nM Aβ (grey). (g) Phase plots of dV/dt versus V constructed from recording as in f in neurons treated for 4 h with 0.05% DMSO (black) or 100 nM Aβ (grey). (h) Quantification of the firing threshold of action potential in neurons treated with 0.05% DMSO (ctrl, −51.33±1.83, n=10) or Aβ (Aβ, −33.82±2.81, n=10). Statistical significance determined by two-tailed t-test, ***P<0.001. In c,e and h, data are expressed as mean±s.e.m. ANOVA, analysis of variance.
