| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1998: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1997: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Research Abstract |
Aquaporin 4 (AQP4) is a predominant water channel protein in mammalian brains, and is localized in astrocyte plasma membrane. Although the regulation of AQP4 is believed to be important for homeostasis of water in brain, its regulatory mechanisms of AQP4 are not known. In this report we have investigated the effect of protein kinase C (PKC) activator on the expression of AQP4 mRNA in cultured rat astrocytes. Cultured rat astrocytes constitutively expressed AQP4 mRNA.Treatment of the cells with 0.1 muM of phorbol ester 1 2-O-tetradecanoylphorbol 13-acetate (TPA), an activator of PKC, caused a rapid decrease in AQP4 mRNA.This effect was time- and dose-dependent. On the other hand, TPA-induced decrease in AQP4 mRNA was inhibited by a relatively specific PKC inhibitor, 1-(5-isoquinoline sulfonyl)-2-methylpiperazin (H7) in a dose-dependent manner. Moreover, prolonged treatment of the cells with TPA eliminated subsequent decrease in AQP4 mRNA by TPA.These results strongly suggest that the TPA-induced decrease in AQP4 mRNA is mediated by PKC activation. To test whether the effect of TPA requires protein synthesis, astrocytes were pretreated with cycloheximide, an inhibitor of protein synthesis. Pretreatment of the cells with cycloheximide did not inhibit the decrease in AQP4 mRNA induced by TPA.To test whether TPA-induced decrease in AQP4 is due to decrease in the mRNA stability, we examined the effect of actinomycin D, an inhibitor of transcription, on TPA-treated cells. The stability of AQP4 mRNA did not decrease by the pretreatment of the cell with actinomycin D.These results suggest th atAQP4 mRNA is inhibited by TPA via PKC activation without de novo protein synthesis, and that the inhibition of AQP4 mRNA could be at transcriptional level.
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