Figure 5
FRET imaging of Rap1 activity in neurons with or without the CaMKII inhibitor. (a and b) Time-lapse FRET imaging of Rap1 activity in dissociated hippocampal neurons treated with the CaMKII inhibitor (b) or its inactive analogue (a). Time stamps (hour) are shown in the upper-left-hand corner of the images. Two spines (marked as [1] and [2] for [A] and [3] and [4] for [B]) are shown as enlarged images in the lower rows. The arrows indicate local Rap1 activation. Bar, 5 µm and 1 µm for the lower- and higher-magnification images, respectively. (c) Quantification of the changes in FRET efficiency in persistent spines, changes indicative of the suppression of transient Rap1 activation by the inhibition of CaMKII activity. Data are presented as the mean ± SEM, (neurons at 16–21 DIV, n = 24 spines from 3 independent time-lapse sessions for both control [KN92] and KN93), **p < 0.01, t-test. (d) Difference in FRET efficiency between newly generated spines and adjacent persistent spines. Data are presented as the mean ± SEM, (neurons at 16–21 DIV, n = 15 spines from 3 independent time-lapse sessions for control [KN92], n = 14 spines from 3 independent time-lapse sessions for KN93), **p < 0.01, t-test.
