Sr8278

The REV-ERBα antagonist SR8278 (Tocris Bioscience) was dissolved in ethanol to a concentration of 50 μg/μL, as described in a previous study^{24 (link)}, and the REV-ERBα agonist GSK4112 (Sigma‒Aldrich) was dissolved in DMSO (Sigma‒Aldrich) to a concentration of 100 μM. SR8278 (16 μg/mouse) and GSK4112 (32 ng) were directly microinjected using a 26-gauge cannula (PlasticOne) into the DR with a Hamilton syringe at a rate of 0.1 μl/min 3 h before social interaction tests.

17β-estradiol was purchased from Aladdin (Shanghai, China). SR8278 was purchased from Tocris Bioscience (Ellisville, MO). ELISA kits for mouse GH and PRL were obtained from Elabscience Biotechnology (Wuhan, China). Anti-GAPDH antibody was purchased from Abcam (Cambridge, MA), and anti-PER2 antibody from Affinity BioReagents (Golden, CA). Anti-HIF-1α antibody was purchased from Genetex (San Antonio, TX), and anti-CCNB2 antibody from Bioworld (Visalia, CA). Anti-CDC20 and anti-ESPL1 antibodies were obtained from Bioss Biotechnology (Woburn, MA). pcDNA3.1, pcDNA3.1-Per2, pRL-TK and luciferase reporters of Ccnb2 (-2000/100 bp), Cdc20 (-2000/100 bp) and Espl1 (-2000/100 bp) were obtained from Transheep Technologies (Shanghai, China). siPer2 (siRNA targeting Per2, sequence is shown in Table S2) was synthesized by IGE Biotechnology (Guangzhou, China). Adenoviral vector encoding siPer2 was obtained from HanBio Biotechnology (Shanghai, China).

GSK4112, SR8278, SR1001, SR1078, lithium chloride and staurosporine (STS) were purchased from Tocris. Drugs were dissolved in DMSO or water and stored as concentrated solutions at −80°C prior to use.

The local injection of SR8278 into the midbrain towards the VTA was performed 3 h before each behavioral test under a dim red light. We followed the time-regiment, for handling animal care and slow microinjection of SR8278 to the VTA 3 h before behavioral tests, as shown previously [13 (link)]. SR8278 (Tocris Bioscience) was dissolved in ethanol to a concentration of 50 µg/µL. SR8278 (20 µg/mouse) or the corresponding vehicle (ethanol) was directly microinfused into the VTA using a 33-gauge injector cannula (model C315I; PlasticOne) attached to a 10-µL Hamilton syringe at a rate of 0.1 µL/min. For the microinfusion, mice were anesthetized with sodium pentobarbital (50 mg/kg, i.p.), mounted on a stereotaxic apparatus (Stoelting), and unilaterally implanted with a 26-guage stainless steel cannula (model C315G, PlasticOne) into the midbrain towards VTA (AP −3.2 mm, ML −0.5 mm, DV −3.5 mm). A 32-gauge dummy cannula was inserted into each guide cannula to prevent clogging. Once the jewelry screws were implanted in the skull as anchors, the whole assembly was affixed to the skull with resin cement.