Somatic Genetics Analysis of Sleep in Adult Mice

Guodong Wang; Qi Li; Junjie Xu; Shuai Zhao; Rui Zhou; Zhenkang Chen; Wentong Jiang; Xue Gao; Shuang Zhou; Zhiyu Chen; Quanzhi Sun; Chengyuan Ma; Lin Chen; Bihan Shi; Ying Guo; Haiyan Wang; Xia Wang; Huaiye Li; Tao Cai; Yibing Wang; Zhineng Chen; Fengchao Wang; Qinghua Liu

doi:10.1523/JNEUROSCI.0089-22.2022

Somatic Genetics Analysis of Sleep in Adult Mice

J Neurosci. 2022 Jul 13;42(28):5617-5640. doi: 10.1523/JNEUROSCI.0089-22.2022.

Authors

Guodong Wang^{1

2

3}, Qi Li^{1

2}, Junjie Xu^{1

2

4}, Shuai Zhao⁵, Rui Zhou^{1

2

6}, Zhenkang Chen⁷, Wentong Jiang^{1

2

3}, Xue Gao^{1

2}, Shuang Zhou^{1

2

4}, Zhiyu Chen^{1

2}, Quanzhi Sun^{1

2}, Chengyuan Ma⁸, Lin Chen^{1

2}, Bihan Shi^{1

2}, Ying Guo^{1

2}, Haiyan Wang^{1

2}, Xia Wang^{1

2}, Huaiye Li^{1

2}, Tao Cai^{1

2}, Yibing Wang^{1

2}, Zhineng Chen⁵, Fengchao Wang^{9

2}, Qinghua Liu^{9

2

3

10}

Affiliations

¹ National Institute of Biological Sciences, Beijing, 102206, China.
² Tsinghua Institute of Multidisciplinary Biomedical Research, Tsinghua University, Beijing, 102206, China.
³ Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, 100730, China.
⁴ College of Life Sciences, Beijing Normal University, Beijing, 100875, China.
⁵ Institute of Automation, Chinese Academy of Sciences, Beijing, 100080, China.
⁶ College of Biological Sciences, China Agriculture University, Beijing, 100094, China.
⁷ Children's Medical Center Research Institute, UT Southwestern Medical Center, Dallas, Texas 75235.
⁸ Chinese Institute of Brain Science, Beijing, 102206, China.
⁹ National Institute of Biological Sciences, Beijing, 102206, China liuqinghua@nibs.ac.cn wangfengchao@nibs.ac.cn.
¹⁰ International Institute for Integrative Sleep Medicine, University of Tsukuba, Tsukuba, 305-8575, Japan.

Abstract

Classical forward and reverse mouse genetics require germline mutations and, thus, are unwieldy to study sleep functions of essential genes or redundant pathways. It is also time-consuming to conduct EEG/EMG-based mouse sleep screening because of labor-intensive surgeries and genetic crosses. Here, we describe a highly accurate SleepV (video) system and adeno-associated virus (AAV)-based adult brain chimeric (ABC)-expression/KO platform for somatic genetics analysis of sleep in adult male or female mice. A pilot ABC screen identifies CREB and CRTC1, of which constitutive or inducible expression significantly reduces quantity and/or quality of non-rapid eye movement sleep. Whereas ABC-KO of exon 13 of Sik3 by AAV-Cre injection in Sik3-E13^flox/flox adult mice phenocopies Sleepy (Sik3 ^Slp/+ ) mice, ABC-CRISPR of Slp/Sik3 reverses hypersomnia of Sleepy mice, indicating a direct role of SLP/SIK3 kinase in sleep regulation. Multiplex ABC-CRISPR of both orexin/hypocretin receptors causes narcolepsy episodes, enabling one-step analysis of redundant genes in adult mice. Therefore, this somatic genetics approach should facilitate high-throughput analysis of sleep regulatory genes, especially for essential or redundant genes, in adult mice by skipping mouse development and minimizing genetic crosses.SIGNIFICANCE STATEMENT The molecular mechanisms of mammalian sleep regulation remain unclear. Classical germline mouse genetics are unwieldy to study sleep functions of essential genes or redundant pathways. The EEG/EMG-based mouse sleep screening is time-consuming because of labor-intensive surgeries and lengthy genetic crosses. To overcome these "bottlenecks," we developed a highly accurate video-based sleep analysis system and adeno-associated virus-mediated ABC-expression/KO platform for somatic genetics analysis of sleep in adult mice. These methodologies facilitate rapid identification of sleep regulatory genes, but also efficient mechanistic studies of the molecular pathways of sleep regulation in mice.

Keywords: AAV-PHP.eB; CRISPR/Cas9; KO; adult brain chimeric; sleep phenotypes; somatic genetics analysis.