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Real-time PCR was carried out using SYBR Green PCR Master Mix (RR091A, Takara, Japan) and the quantitative analysis was performed using LightCycler Roche 480 qPCR instrument

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Since acetylcholine does not cross the blood-brain barrier, brain levels of acetylcholine are highly dependent on choline balance and choline availability in the brain, and can therefore be enhanced by increasing plasma and brain choline levels

We eventually succeeded in creating a model that had between 12- to 20- fold higher 5-OP levels, substantially higher folds than observed in mammalian system