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Fig. 4 Notch signaling mediates the regulation of pericyte proliferation by blood flow and Piezo1 (A) Representative images of the d2GFP-labeled ECs with Notch activation in the brain of Yoda1-treated Tg(fli1a:DsRedEx);Tg(Tp1:d2GFP) larvae at 4.5 DPF, compared to the location-matched ECs in the DMSO control larvae. EC is in red, and cell of high Notch activation is in green. (B) Effects of Yoda1 treatment and piezo1 knockout on Notch signaling in brain ECs. TP1+ brain EC number denotes the total number of TP1+ ECs in the entire brain. (C) Representative images of the pericytes and blood vessels in the midbrain of DMSO-, DAPT-, DAPT+NB-, and DAPT+Yoda1-treated Ki(pdgfrb:GAL4-VP16);Tg(4×nrUAS:GFP);Tg(fli1a:DsRedEx) larvae at 4.5 DPF. EC is in red and pericyte in green. All experiments were independent for DMSO controls. (D) Effects of DAPT, DAPT+NB, and DAPT+Yoda1 treatment on midbrain pericyte number. All experiments illustrated were independent for DMSO controls. (E) Effects of DAPT, DAPT+NB, and DAPT+Yoda1 treatment on pericyte coverage of midbrain vessels. All experiments were independent for DMSO controls. (F) Effect of DAPT treatment on brain pericyte proliferation. All experiments were independent for DMSO controls. Images are shown from a top view and partially z-projected. Scale bar, 50 μm for (A) and (C). Images of 4.5-DPF larvae are used for counting the number of pericytes and the pericyte coverage of the vessels in the midbrain, and time-lapse images during 3.0–3.5 DPF are used for counting the mean proliferation frequency of brain pericytes. Data are represented as mean ± SEM. The N values are shown above the aligned plots. Stars represent the results of unpaired two-tailed Student’s t test between groups (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001). See also Figure S6.