Figure 7. Overlapping distribution between ZBP1 and ß-actin mRNA granules: effects of 15 min KCl depolarization.

A, Triple-label fluorescence detection of ß-actin mRNA (red), ZBP1 (green), and F-actin, using phalloidin (blue). By conventional digital imaging ZBP1 and ß-actin mRNA both exhibited a granular pattern in dendrites with evidence for overlapping, but not highly colocalized, signals. A 3-D deconvolution and volume restoration were performed on two dendritic subregions that appeared to contain ZBP1 and ß-actin mRNA signal in dendritic spines (orange and blue boxes).

B, Enlargement of orange-boxed region depicts a granule within a bulbous dendritic spine that contains both ß-actin mRNA (red) and ZBP1 (green). In the dendritic shaft the granules containing ZBP1 and ß-actin mRNA varied in size from small puncta to larger clusters or aggregates.

C, Enlargement of blue-boxed region also depicts granules that contain both ZBP1 and ß-actin mRNA signal within the spine and shaft (see also video 4).

D, Quantitative colocalization analysis of the spatial relationship between pixels containing ZBP1 (green) and ß-actin mRNA (red). KCl treatment resulted in an overall 31.7% increase in the percentage of ZBP1 pixels that also contained ß-actin mRNA (green bars), with statistical significance observed in proximal (25% increase) and middle (66% increase) dendritic regions. KCl treatment resulted in an 11.4% increase in the percentage of ß-actin mRNA pixels that also contained ZBP1 (red bars), with statistical significance observed in middle dendritic regions (25% increase). Bars show group means ± SEM; p <= 0.01*, two-tailed Mann-Whitney t test. Black asterisks denote significance as compared with control untreated neurons.