hGATA1 and hGATA-2 decreases dendritic arborization and spine density.

Neuronal complexity in the hippocampal neurons transfected with AAV-hGATA1 or AAV-hGATA2 was analyzed based on three-dimensional projections of confocal stack images through complete neuritic extension of GFP(+) process. The neurons were transfected with AAV on 4 DIV and analyzed on 24 DIV. (a) An averaged image of multiple confocal planes across GFP(+) cells. (b) Sholl analysis of GFP(+) cells. Cells transfected with AAV-ctl presented neuritis with an increased ramification compared with cells transfected with AAV-hGAT1 and AAV-hGATA2 (^*P<0.05). (c) Representative images are shown of high-magnification Z-stack projections of dendrites of GFP(+) cells from AAV-ctl- and AAV-hGATA-transfected cells. A continuous stretch of dendritic processes was imaged. (d) The spine density (arrowheads) of proximal (P) and distal (D) segments was significantly decreased in AAV-hGATA-transfected cells compared to AAV-ctl-transfected rats (^*P<0.05, ^**P<0.01 compared to proximal region of AAV-ctl; ^#P<0.05, ^###P<0.001 compared to distal region of AAV-ctl; n = 9–12 neurons in each group). The data were expressed as the number of spines per 10 µm. (e) MAP2, PSD-95, Tuj1 and GAP-43 expression in the cells described in (a) determined by Western blot analysis. Expression levels are depicted relative to the level of cells that were not exposed to virus (: no virus) for comparison. (n = 3–4 biological replicates per group). ^*P<0.05, ^**P<0.01, ^***P<0.001 compared to control vector. Scale bars: a, 30 µm; c, 10 µm. Student's t-test.