Du Congwu, Koretsky Alan P, Izrailtyan Igor, Benveniste Helene
Brookhaven National Laboratory, Medical Department, Upton, New York 11973-5000, USA.
J Cereb Blood Flow Metab. 2005 Aug;25(8):1078-92. doi: 10.1038/sj.jcbfm.9600102.
We describe an approach to measure changes in intracellular calcium along with changes in blood volume and oxygenation directly from the exposed rat cortex in vivo during cerebral ischemia and reperfusion. Measurements were made using a catheter-based optical system. The endface of a Y-shaped bifurcated fiber optic bundle was mounted on the cortical surface. It delivered the light at three wavelengths of 548, 555, and 572 nm to the brain through a fast monochromator coupled to a xenon lamp, and collected the calcium-dependent fluorescence emission from Rhod2 at 589 nm (excited at 548 nm) along with the diffuse reflections at the wavelengths of 555 and 572 nm to determine the changes in blood volume and hemoglobin oxygenation. The feasibility of this approach was experimentally examined by inducing transient cerebral ischemia and reperfusion in the rat. The ischemia induced an 8.5%+/-1.7% fluorescence increase compared with the preischemic control values. Blood volume and tissue hemoglobin oxygenation decreased by 57.4%+/-12.6% and 47.3%+/-12.5%, respectively. All signals normalized on reperfusion. The ischemia-induced change in Rhod2-Ca2+ fluorescence was blocked using a calcium channel blocker, nimodipine, confirming that intracellular changes in calcium were responsible for the fluorescence changes. Thus, changes in cerebral hemodynamics and intracellular calcium concentration changes were measured simultaneously, facilitating future studies of the interrelationship between neuronal activation and metabolic and vascular processes in normal and diseased brain.
我们描述了一种在脑缺血和再灌注期间直接从暴露的大鼠活体皮层测量细胞内钙变化以及血容量和氧合变化的方法。测量使用基于导管的光学系统进行。一个Y形分叉光纤束的端面安装在皮层表面。它通过与氙灯耦合的快速单色仪将548、555和572nm三个波长的光传输到大脑，并收集589nm处来自Rhod2的钙依赖性荧光发射（在548nm激发）以及555和572nm波长处的漫反射，以确定血容量和血红蛋白氧合的变化。通过在大鼠中诱导短暂性脑缺血和再灌注，对该方法的可行性进行了实验研究。与缺血前对照值相比，缺血诱导荧光增加8.5%±1.7%。血容量和组织血红蛋白氧合分别下降57.4%±12.6%和47.3%±12.5%。再灌注时所有信号恢复正常。使用钙通道阻滞剂尼莫地平可阻断缺血诱导的Rhod2-Ca2+荧光变化，证实细胞内钙变化是荧光变化的原因。因此，可同时测量脑血流动力学变化和细胞内钙浓度变化，有助于未来对正常和患病大脑中神经元激活与代谢及血管过程之间相互关系的研究。
