共同研究報告書


研究区分 一般研究

研究課題

 哺乳類冬眠中の脳内熱産生メカニズムの光学的解析
新規・継続の別 新規
研究代表者／所属 生命創成探究センター バイオフォトニクス
研究代表者／職名 JSPS Postdocs Fellow
研究代表者／氏名 張菁圃

研究分担者／氏名／所属／職名
　
氏　　名
所　　属
職　　名

1

 榎木亮介 イオフォトニクス研究グループ 准教授

2

 山口良文 北大低温研

3

 李明亮 イオフォトニクス研究グループ 特任助教　　

研究目的 Hibernators use the strategy to survive harsh environments， such as low temperature and food scarcity， by reducing metabolism and body temperature. Body temperature is mainly controlled by brown adipose tissue (BAT) that is rich in mitochondria and highly expresses uncoupling protein 1 (UCP1)， where heat is generated. I found that UCP1 is also expressed in the hamster brain. However， it is unclear whether the brain UCP1 is functional. We will measure the intracellular temperature and study the role of brain UCP1 in the thermoregulation and the physiological functions during hibernation.
　　
研究内容・成果 To address the role of UCP1 in the brain， we first confirmed the expression of UCP1 in neurons in the paraventricular nucleus of the hypothalamus (PVN) and in astrocytes near the third ventricle， which is innervated by tanycytes， in the hamster， but not in the mouse， using multiple immunohistochemical techniques. We next performed quantitative analysis to characterize the expression of brain UCP1 during the hibernation cycle. We found that hypothalamic UCP1 expression was upregulated upon to the long-term cold exposure and returned to the same level as that of warm control during the deep torpor， whereas the amount of BAT UCP1 peaked during the deep torpor. It suggested that there is a BAT independent thermoregulatory mechanism in the brain.

To investigate the thermogenic capabilities of hypothalamic astrocytes and neurons， we first conducted the acute brain slice calcium imaging and found that both astrocytes and neurons in the hypothalamus were responded to the cold stimulation. Next， we will measure the intracellular temperature by delivering the genetically encoded temperature indicator (GETI) B-gTEMP [Kai Lu， et al.， 2022] into the cytosol and mitochondria of hypothalamic astrocytes or neurons in Syrian hamsters with adeno-associated virus transfection. The temperature inside cytoplasm and mitochondria will be monitored in acute brain slice under multi-photon microscopy under temperatures ranging from 5 to 35 degrees. In the past year， we have established the virus injection techniques and preliminarily confirmed the expression of B-gTEMP in the hamster brain. To further understand the physiological role of UCP1 in real hibernation conditions， we will perform in vivo fibre photometry recording from hypothalamic astrocytes expressing B-gTEMP during hibernation cycles. To this aim， we have constructed the fibre photometry systems and established operations for long-term recording.

To specify the necessity of UCP1， all the experiments mentioned above will be conducted in wild-type and UCP1 knockout (KO) hamsters. The UCP1-KO hamsters were successfully generated by Prof. Yamaguchi at the Institute of Low Temperature Science (ILTS) at Hokkaido University， and the brain samples were collected and the expression of UCP1 was preliminarily examined by immunostaining.
　　
成果となる論文・学会発表等 [1] CHANG Ching-Pu， YAMAGUCHI Yoshifumi， WATANABE Masatomo， NEMOTO Tomomi， ENOKI Ryosuke. “Identification of uncoupling protein 1 (UCP1) in the hypothalamus of Syrian hamster brain”. Abstract for “The 46th Annual Meeting of the Japan Neuroscience Society. Sendai， Japan. August 1-4， 2023.”

[2] CHANG Ching-Pu. “Optical Analysis of Brain Thermogenesis in Mammalian Hibernator.” Oral present for “Hibernation Biology Meeting in Sapporo and Jozankei Onsen. October 11-12， 2023”