Supplementary Figure 6: Limited Cre recombinase activity in peripheral organs, systemic physiologic parameters and liver and kidney analyses, and visual behavior testing after pericyte ablation with DT.

(a-d) Representative confocal microscopy images from 3 pericyte-CreER; Ai14 mice 14 days after TAM treatment (40 mg/kg per day for 7 days as in main figures) showing limited tdTomato expression in kidney (a), liver (b), heart (c) and skeletal muscle (d). PDGFRβ is used as pericyte marker, Lectin is used to label endothelium, and nuclei were stained with DAPI. Images in a-d representative of n = 3 independent mice with similar results. Bar = 50 µm. (e) Heart rate, respiratory rate (n = 4 vehicle-treated, n = 5 DT-treated mice/group), arterial pO₂, pCO₂, pH (n = 5 vehicle-treated, n = 6 DT-treated mice/group), and glucose levels (n = 4 vehicle-treated, n = 3 DT-treated mice/group); (f), liver analyses: alkaline phosphatase, ALP; alanine aminotransferase, ALT; aspartate aminotransferase, AST; creatine phosphokinase, CPK; albumin, total protein, total bilirubin. n = 4 vehicle-treated, n = 3 DT-treated mice/group. (g) kidney analyses: blood urea nitrogen, creatinine, sodium, calcium (n = 4 vehicle-treated, n = 3 DT-treated mice/group), and potassium (n = 5 mice/group) were not altered in TAM-treated pericyte-CreER; iDTR mice at 3 days post-DT or vehicle treatment. Mean ± S.E.M., Significance by two-tailed Student’s t-test. (Supplementary Figure 1, 2 and 3). (h) Visual behavior testing. TAM-treated pericyte-CreER; iDTR mice treated with DT (developing pericyte ablation) or vehicle (controls) underwent visual behavior testing at 15 days post-DT using a visual cliff test. Briefly, a transparent Plexiglass square arena was divided into two equal parts by aligning the middle of the arena with the edge of a table. The side sitting on the tabletop was considered a “shallow” side and the other one that is positioned over the floor area (50 cm high) a “deep” side. A patterned floor consisting of 3 cm black and white checked paper was placed bellow the arenas: on the shallow side the checked paper was placed immediately below the Plexiglass surface, while on the deep side the paper was placed on the floor creating an illusion of a cliff. Each animal was placed on the shallow side and the total time the animal spent exploring each side of the arena was recorded within a 5 min trial, and the percentage time spent in the shallow side was analyzed. As a positive control, blind C3H/HeJ mice were also tested. Circles represent percent time spent on shallow side of arena for each mouse tested; boxes represent mean ± S.D.. n = 8 C3H/HeJ control mice, n = 6 vehicle-treated, and n = 3 DT-treated pericyte-CreER; iDTR mice; one way ANOVA followed by Bonferroni posthoc test.