Neurodegeneration, Neuroprotection, and Disease-Oriented NeuroscienceResearch PaperOrexin neurons in hypothalamic slice cultures are vulnerable to endoplasmic reticulum stress
Highlights
▶Effect of ER stress inducers was examined in hypothalamic slice cultures. ▶ER stress inducers caused selective loss of orexin. ▶Loss of orexin at early stage resulted from depletion of stored orexin. ▶Loss of orexin at later stage resulted from degeneration of orexin neurons. ▶ER stress might be relevant to pathogenic events in narcolepsy.
Section snippets
Preparation of hypothalamic slice cultures
Organotypic slice cultures of rat hypothalamus were prepared as described in our previous studies (Katsuki and Akaike, 2004, Michinaga et al., 2010b. Experimental procedures were approved by our institutional ethical committee concerning animal experiments, and were conformed to the Guide and Use of Laboratory Animals as adopted and promulgated by the US National Institutes of Health. Briefly, brains were obtained from 7–8-day-old Wistar/ST rats (Nihon SLC, Shizuoka, Japan) under deep
Orexin neurons decrease more prominently than other neuropeptide-containing neurons during ER stress
To examine whether ER stress could affect orexin-immunoreactive neurons, we used conventional ER stress inducers such as tunicamycin and thapsigargin. Tunicamycin, by preventing protein N-glycosylation (Kaneko and Tsukamoto, 1994), promotes accumulation of misfolded proteins that leads to ER stress and initiation of UPR. Thapsigargin inhibits Ca2+-ATPase of ER (Perez-Sala and Mollinedo, 1995) and depletes ER luminal Ca2+, which leads to protein unfolding and misfolding due to Ca2+-dependent
Discussion
Here we explored potential relationship between ER stress and the integrity of orexin neurons. For this purpose, we prepared hypothalamic slice cultures from rats at 7 or 8 days old, and after 18 days of cultivation, treated them with various drugs. This corresponds to the usage of brains of 25 or 26-day-old rats, provided that developmental events proceed in vitro in the same manner as those in vivo. In this context, a study on developmental changes of orexin expression has shown that
Acknowledgments
This work was supported by a Grant-in-Aid for Scientific Research from The Ministry of Education, Culture, Sports, Science and Technology, Japan.
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