Research Paper Volume 11, Issue 16 pp 6336—6357

Inhibition of de novo ceramide biosynthesis affects aging phenotype in an in vitro model of neuronal senescence

Figure 4. Effects of aging and L-CS on Ca2+transient frequency and amplitude.(A) Representative brightfield (left) and fluorescent (middle and right) micrographs of a fluo-4-loaded agedL-CSneuronal culture employed to monitor spontaneous Ca2+transients (scale bar 25 µm). Greyscale fluorescent images show cortical neurons before (middle) and during (right) a Ca2+transient. (B) Time course of somatic spontaneous Ca2+oscillations in the four study groups. Each trace depicts a single neuron representative of at least three independent experiments. (C) Bar graphs depict average transient frequencies of vehicle- or L-CS-treated control and aged neurons (ControlVeh: n=499, ControlL-CSn=253, AgedVehn=367, and AgedL-CSn=293 cells obtained from 15-38 experiments). (D) Bar graphs depict the average Ca2+transient amplitude in the four study groups [samples are the same as in (C)]. (E) Representative greyscale fluorescent micrographs of a fluo-4-loaded primary dendrite before (left) and during (right) a Ca2+transient. (F) Time course of dendritic spontaneous Ca2+oscillations in the AgedVehand AgedL-CScultured neurons. Each trace depicts a single dendrite representative of at least three independent experiments. (G) Bar graphs depict average transient frequencies of AgedVehand AgedL-CSdendrites (AgedVehn=21 and AgedL-CSn=27 dendrites from 12-18 experiments). (H) Bar graphs depict the average dendritic Ca2+transient amplitude in the four study groups [samples are the same as in (G)]. In C and D means were compared by two-way ANOVA followed by Tukey post-hoc test. In G and H means were compared by unpaired Student t-test. ** indicates p<0.01, n.s. indicates not significant.