【摘要】：一氧化氮(NO)是一種重要的信號轉導分子,參與腦內許多生理和生理病理過程。大量證據(jù)表明伴隨著缺氧和缺血的腦損傷,海馬神經元會產生過量的NO。缺氧缺血時,離子跨膜重新分布,增加的鈣內流能明顯地影響NO合成酶的活性和產生NO的量,從而改變神經組織中的NO水平。然而,目前仍不清楚內源性的NO與引起鈣內流的離子通道之間的關系。另外,對NO通過何種機制來介導鈣內流也不十分清楚。L型鈣通道在海馬錐體神經元上分布很廣,約30-50%的鈣離子總電流來自L型鈣通道。因此,調控L型鈣通道的活動將顯著影響鈣離子的內流。 在本研究中,我們探討了內源性一氧化氮對大鼠海馬神經元L型鈣通道的作用及其機制。當從鉗制電位-50mV去極化至0mV時,NOS的底物L-精氨酸(L-Arg,1-3mM)增加了61%的L鈣電流(n=30,p0.01)。另外,L-Arg使從-20mV到10mV的電流電壓曲線(Ⅰ-Ⅴ)左移,說明這時的通道對去極化的電壓變得更敏感。尼菲地平(NFDP)是二氫吡啶類的L型鈣通道的阻斷劑。20uMNFDP將L鈣電流減少了60%(n=3,p0.05),L-Arg不再使剩余的電流增大,說明L-Arg的作用是通過L型鈣通道起作用的。與L-Arg的作用相反,NOS的抑制劑N~G-Nitro-L-Arginine Methyl Ester(L-NAME,1-3mM)明顯減少約51%的L鈣電流(n=8,p0.01),提示在培養(yǎng)的海馬神經元上一氧化氮(NO)對通道有基礎調節(jié)。若預處理1mM L-NAME(n=16)和另一種nNOS抑制劑7-Nitroindazole(7-NINA,n=10),兩者皆能部分阻斷L-Arg對通道電流的增大作用(p0.05)。這些結果表明內源性的NO能使海馬L型鈣通道增強通道開放和提高其對電壓的敏感性。 為了探測S-亞硝化是否參與介導內源性NO引起的通道活動,我們用N-ethylmaleimide (NEM,能二價修飾蛋白硫醇基團從而阻止進一步的S-亞硝
[Abstract]:Nitric oxide (NO) is an important signal transduction molecule involved in many physiological and pathophysiological processes in the brain. There is plenty of evidence that hippocampal neurons produce excess no with hypoxic and ischemic brain damage. During hypoxia-ischemia, the ion redistributes across the membrane, and the increased calcium influx can obviously affect the activity of no synthase and the amount of no produced, thus changing the level of no in nerve tissue. However, the relationship between endogenous no and ion channels causing calcium influx remains unclear. In addition, the mechanism of no mediated calcium influx is not very clear. L-type calcium channel is widely distributed in hippocampal pyramidal neurons, about 30-50% of the total calcium current comes from L-type calcium channel. Therefore, the regulation of L-type calcium channel activities will significantly affect the influx of calcium ions. In this study, we investigated the effect and mechanism of endogenous nitric oxide on L-type calcium channel in rat hippocampal neurons. L-arginine (L-Arginine 1-3mm), the substrate of 0mV, increased by 61% L Ca ~ (2 +) current (n ~ (30) p _ (0.01) when depolarization from clamp potential to -50 MV. In addition, L-Arg shifts the current voltage curve (鈪，

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