The research with genetic engineering regarding the circulatory regulation of spinal microvasculature and the effects of anesthetics

从大鼠或小鼠脊髓制备脊髓切片（150 μm厚）。对照液以1.5ml/min的流速持续灌流，以93%O_2 + 7%CO_2（PCO_2 = 40 mmHg，pH = 7.4，37℃）鼓泡。在神经元组织内定位一条内径为5.0-10.0 μm的脑实质内小动脉，并通过实时计算机视频显微镜连续监测其内径。视频显微镜设备包括倒置显微镜。将实质小动脉的图像传输到摄像机（C6790-81，Olympus，Tokyo，Japan），并通过媒体转换器（Physio-Tech，Tokyo，Japan）显示在计算机上。在前列腺素F_（2α）（0.5 μM）的作用下，高碳酸血症（10%CO_2产生的PCO_2 = 50 mmHg）引起脊髓实质内微动脉扩张，并记录脊髓微血管管腔内径的变化。在正常小鼠的脊髓切片中，加入乙酰胆碱（0.1 μM）也会诱导这些小动脉扩张，而在神经元型一氧化氮合酶敲除小鼠的切片中，该浓度的乙酰胆碱不会诱导小动脉扩张。麻醉剂对上述血管扩张反应的影响正在研究中，这些结果表明，在脊髓微动脉中，二氧化碳发挥着血管扩张物质的作用，神经元型一氧化氮合酶参与了乙酰胆碱等生理药物诱导的脊髓血管扩张反应。

Spinal slices (150 μm thick) were prepared from the rat or mouse spinal cords. Slices were continuously superfused with control solution at the flow rate of 1.5 ml/min, bubbled with 93% O_2 + 7% CO_2 (PCO_2 = 40 mmHg, pH = 7.4, 37℃). An intraparenchymal arteriole (5.0-10.0 μm in internal diameter) was located within the neuronal tissue, and its internal diameter was continuously monitored with the live computerized videomicroscopy. The videomicroscopy equipment consisted of an inverted microscope. The image of a parenchymal arteriole was transmitted to video camera (C6790-81, Olympus, Tokyo, Japan) and displayed on a computer via a media converter (Physio-Tech, Tokyo, Japan). Changes of intraluminal diameter in spinal microvessels were recorded on computer image files and then analyzed using the image software.During contraction to prostaglandin F_<2α> (0.5 μM), hypercapnia (PCO_2 = 50 mmHg produced by 10% CO2) induced dilation of intraparenchymal spinal arterioles. Addition of acetylcholine (0.1 μM) also induced dilation of these arterioles in the spinal slice from normal mice, whereas this concentration of acetylcholine did not induce dilation of arterioles in the slice from neuronal nitric oxide synthase knockout mice. Effects of anesthetics on above vasodilator responses are under investigation.These results suggest that in spinal microarterioles, carbon dioxide play a role as a vasodilator substance and that neuronal nitric oxide synthase contributes to spinal vasodilator responses induced by physiological agents including acetylcholine.