The Journal of Neuroscience, March 14, 2007, ():
Brain-Derived Neurotrophic Factor Promotes Long-Term Potentiation-Related Cytoskeletal Changes in Adult Hippocampus
J. Neurosci. Rex et al.
27: 3017
Supplemental Data
Files in this Data Supplement:
- supplemental material
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Supplemental Figure 1. In situ labeling of F-actin with rhodamine-phalloidin in adult hippocampal slices does not alter normal synaptic potentiation. A) Hippocampal slice schematic shows stimulation (S) and recording electrode placements. B) Low-power photomicrograph of field CA1 in a hippocampal slice that received topical application of rhodamine-phalloidin followed by TBS at each of two stimulation sites. The dashed-line box indicates the location and size (550 μm2) of the sample region used for labeled-spine analysis. Calibration bar: 50 µm. C) Representative fEPSPs (set to 40% of maximum population spike-free amplitude) collected from the two pathways (S1 and S2) after topical application (20 min) of phalloidin show that synaptic potentials had normal shapes and amplitudes. Traces recorded 15 min after TBS (15’) show that the patterned stimulation produced robust elevations of fEPSP amplitudes and slopes compared to baseline (0’). Calibration bars: 0.5 mV, 5 ms. D) Group data for fEPSP slopes (means ± SEM) indicate that LTP in phalloidin-treated slices stabilized at values (~140% of baseline) equivalent to those described for untreated slices in earlier reports. E) Composite theta burst responses and their facilitation within a theta train are comparable in control (topical aCSF-treated) and phalloidin-treated slices. F) Facilitation of burst responses (normalized to area of first burst response; mean ± SEM) was similar for groups of control and phalloidin treated slices (p>0.7, 2-way repeated measures ANOVA).
- supplemental material
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Supplemental Figure 2. Greater numbers of phalloidin-labeled spines are detected with widefield as compared to confocal microscopy. Acute hippocampal slices from young adult rat were infused with rhodamine-phalloidin for 20 min and then stimulation was applied to converging afferents to CA1 str. radiatum. Tissue was processed as described in Methods and labeling was evaluated using either widefield fluorescence microscopy with extended focal imaging or with laser scanning confocal microscopy. A,B) Micrographs captured by laser scanning confocal microscopy show in situ phalloidin-labeling in hippocampal slices that received control (CON, A) or theta burst (TBS, B) stimulation. Calibration bar = 10 μm. C-D) Plots show raw spine counts (within the 550 μm2 CA1 sampling zone) across 10-pixel intensity unit bins for similarly treated slices imaged by (C) laser scanning confocal microscopy or (D) widefield fluorescence microscopy with CCD camera. Comparison of the two plots reveals that greater numbers of faintly labeled spines were detected by widefield microscopy.
