Astrocyte CCN1 stabilizes neural circuits in the adult brain - Nature

"All animal experiments were approved by the Salk Institute Institutional Animal Care and Use Committee (IACUC). Rats and mice were typically housed with a standard 12 h:12 h light:dark cycle in the Salk Institute animal facilities, with lights on at 06:00 and lights off at 18:00. Dark-reared mice were housed with a 24 h dark cycle since birth. Mice and rats were provided access to food and water ad libitum. Humidity ranged from 38-62% and temperature from 20 to 22 °C."

"For bulk RNA=sequencing experiments, astrocyte-Ribotag mice were generated by crossing Gfap-cre hemizygous females (B6.Cg-Tg (GFAP-cre)73.12Mvs/J, Jax 012886) to homozygous flox-Rpl22-HA males (B6N.129-Rpl22tm1.1Psam/J, Jax 011029). Male mice hemizygous for cre and heterozygous for flox-Rpl22-HA ( Rpl22-HA+;Gfap-cre+) were used for all experiments. Wild-type C57Bl6/J mice were used (Jax 000664) for experiments."

"For adult knockout experiments, Ccn1fl/fl mice were a gift from L. Lau and were maintained on a C57Bl6/J background. These mice were crossed to mice expressing tamoxifen-inducible Cre recombinase under an astrocytic promoter for temporal elimination ( Aldh1;creERT2, Jax 029655 (ref. )). Experimental mice were homozygous for the Ccn1 floxed allele and either cre− or cre+ (wild type or Ccn1-cKO). For adult cKO experiments, mice were injected intraperitoneally with 75 mg kg −1 of tamoxifen (MP Biomedicals 156738) for 5 consecutive days at 1 month of age."

All animal procedures received IACUC approval and animals were housed under controlled light cycles, humidity (38–62%), and temperature (20–22 °C) with ad libitum food and water. Dark-reared mice experienced 24 h darkness from birth. Astrocyte-Ribotag mice were produced by crossing Gfap-cre hemizygous females to flox-Rpl22-HA homozygous males, and male Rpl22-HA+;Gfap-cre+ mice were used for bulk RNA sequencing. Wild-type C57Bl6/J males were used for snRNA-seq and smFISH validation. Ccn1fl/fl mice were crossed to Aldh1;creERT2 for temporal astrocyte-specific deletion, with specified tamoxifen regimens for juvenile and adult cKOs. Sample sizes were power-calculated, sexes were recorded, and experimenters were blinded. Sprague-Dawley rats were used at P1–2.

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