Tamoxifen

C57BL/6J mice were purchased from Charles River Laboratories (Jiaxing, China) and Shanghai SLAC Laboratory Animal Co.Ltd (Shanghai, China). Gli1^CreERT2, Lgr5^{EGFP−CreERT2}, R26R^tdTomato mice were purchased from the Jackson Laboratory. Gli1^CreERT2; R26R^tdTomato and Lgr5^{EGFP−CreERT2}; R26R^tdTomato mice were used to monitor the activation and proliferation of Gli1+ and Lgr5+ HFSCs after treatment. Four or six days after depilation, mice (6–7 weeks old) received three i.p. injections of 200 mg/kg Tamoxifen (MACKLIN, Shanghai, China) dissolved in corn oil (MACKLIN) at 1-day intervals. Mice were housed with a 12 h/12 h light/dark cycle at 22 °C, with free access to food and water. Only male mice were used for the experiments. The experimental study was conducted according to the institutional guidelines with approval from the Review Committee of Zhejiang Chinese Medical University.

To pharmacologically ablate myeloid cells, mice were administered a PLX5622-formulated AIN-76A diet (1.2 g PLX5622 per kilogram of diet, formulated by SYSE Bio) ad libitum for 14 d. To pharmacologically ablate peripheral macrophages, mice were administered a PLX73086-formulated AIN-76A diet (0.2 g PLX73086 per kilogram of diet, formulated by Research Diet) ad libitum for 14 d. Control mice were fed an AIN-76A CD. Since the microglial ablation efficiency by CSF1R inhibition might be different between sexes (Shi et al., 2019 (link)), we utilized male mice for this experiment. To efficiently induce CreER-dependent recombination, tamoxifen (150 mg per kg of body weight) dissolved in olive oil (Macklin, 0815210) was administered via oral gavage for four consecutive days following our previously described procedures (Huang et al., 2018b (link); Zhou et al., 2022 (link); Rao et al., 2021 (link); Xu et al., 2020 (link); Huang et al., 2018a (link)). 2-MeSAMP (10 mM) was injected into the lateral ventricle 90 min before the behavior test (Mastorakos et al., 2021 (link)). CNO (100 μg/mL) was administered via intraperitoneal injection 90 min before the behavior test (Binning et al., 2020 (link)).