Identification of brain regions involved in the arousal from coma and sleep.

2016 Fiscal Year Annual Research Report

• Project/Area Number: 16H06661
• Research Institution: University of Tsukuba
• Principal Investigator: マリシェフサカヤ オリガ 筑波大学, 国際統合睡眠医科学研究機構, 研究員 (20739429)
• Project Period (FY): 2016-08-26 – 2018-03-31
• Keywords: coma / arousal / parabrachial nucleus / lesion / DTA-AAV / inhibition / IVM-Glu-Cl / pons

Outline of Annual Research Achievements

On a first year of project we focused on generation of mouse model of coma. First we designed and produced AAV that caries diphtheria-toxin-subunit A (DTA). The DTA-AAV plasmid and virus was created in our laboratory and it allows focal lesion of neurons without undesired side effects, such as inflammation. We microinjected DTA-AAV to the parabrachial (PB) nucleus of the pons, which is responsible for vigilance state maintenance. Lesion of this area by DTA-AAV produced vestibular-ataxia state on day 3 and developed in to minimally-conscious state following by coma on day 6 (7) and death by the day 7. All animals (n=5) displayed very similar phenotype with a small difference in severity of coma, mainly varying in onset of symptoms and lifespan.

Current Status of Research Progress

3: Progress in research has been slightly delayed.

Reason

Our initial microinjections produced severe inhibition of vigilance state resulting in early coma and death. One of possible reasons is a serotype of AAV, that transfect larger area including glia and produces much bigger inhibition than necessary. Another possibility is that area of inhibition includes more lesion area, such as PB and PC, or PB and DR. This data resulted in carrying-over the research project to the next year.

Strategy for Future Research Activity

We generated new AAV with less lethal effects to produce sufficient inhibition of vigilance state, but not exceeding it. On the other hand we performed more animal experiments with microinjections of AAV restricting to the PB nucleus.