Differential roles of AVP and VIP signaling in the postnatal changes of neural networks for coherent circadian rhythms in the SCN

ABSTRACT: Cry double deficient (Cry1,2?/?) mice and Vip receptor 2 deficient (Vipr2?/?) mice were bred with PER2::LUC mice carrying a PER2 luciferase reporter. Wild–type (Cry1,2+/+/Vipr2+/+) PER2::LUC transgenic mice on the C57BL/6J background were used as control. For the measurement of PER2::LUC bioluminescence from a cultured SCN slice, mice of 8–16 weeks or 2–5 days old, kept under LD condition, were euthanized between 8:00 and 16:00 by cervical dislocation and decapitated. The brain was rapidly removed and a coronal SCN slice of 150 ?m or 200 ?m was made. In the co–culture experiment, the SCN slices of 150 ?m thick were obtained from adult mice carrying the PER2::LUC reporter (recipient). The slice was pre-cultured for 3 or 4 days, and then co–cultured with an SCN slice from mice without the reporter system (donor). The donor SCN slice of 200 ?m thick was obtained from WT mice of 7 days old and pre-cultured for one day before the co–culturing. When co-cultured, the graft SCN slice was placed inside out on the surface of recipient SCN slice. Measurement of the bioluminescence was started from the beginning of culturing of the recipient SCN and continued for at least 5 days after the co–culture. AVP receptor antagonists (V1A receptor antagonist: SR49059; TOCRIS, V1B receptor antagonist: SSR149415; Axon Medchem) were dissolved in water (SR49059 and SSR149415: final 2.5 ?M). Antagonists were applied into the medium 5 to 7 days after co–culturing.

SUBMITTER: Daisuke Ono 

PROVIDER: S-BSST212 | bioimages |

REPOSITORIES: bioimages