明胶墨汁灌注展示大鼠视网膜血管的方法

Method of gelatin-ink perfusion to visualize rat retinal vessel

目的对大鼠视网膜墨汁灌注方法进行改良以充分显示微血管。方法78只SD大鼠分两部分进行实验。第一部分中,72只大鼠根据灌注液中明胶的浓度分为纯墨汁组(n=18)、1%明胶墨汁组(n=18)、3%明胶墨汁组(n=18)、5%明胶墨汁组(n=18)。每组动物根据灌注压强分为90、115、140、165、190、215mmHg6个亚组。墨汁从升主动脉进行灌注。用视网膜铺片和切片检测微血管的灌注情况。根据第一部分结果,在第二部分中,6只大鼠先后灌注3%明胶墨汁和5%明胶墨汁各20ml。用上面相同的方法检测微血管的灌注情况。结果在165mmHg压强下灌注37℃的3%明胶墨汁时,视网膜周边部的血管充填充分,但中央部的血管充填不充分。在215mmHg压强下灌注37℃的5%明胶墨汁时,视网膜周边部的血管充填不充分,但中央部的血管充填充分。在165mmHg压强下灌注3%明胶墨汁20ml,再在215mmHg压强下灌注5%明胶墨汁20ml能使视网膜中央部和周边部的大小血管都充填充分,且墨汁无外渗、在水溶液中不褪色、灌注血管与周边组织对比明显。结论在165mmHg恒压下灌注37℃的3%明胶墨汁20ml,再在215mmHg恒压下灌注37℃的5%明胶墨汁20ml是一种较好的显示大鼠视网膜微血管的形态学方法。

Objective To improve the ink perfusion method, thereby enabling it to more completely show the rat retinal microvessels. Methods The present study included two parts. The first part comprised of 72 rats that were randomly divided into four groups： only ink group （ n = 18 ）, 1% gelatin-ink group （ n = 18 ）, 3% gelatin-ink group （n = 18）, and 5% gelatin-ink group （n =18）. The animals of each group were subdivided into 90, 115, 140, 165, 190 and 215 mmHg subgroups, chosen based on their mean arterial pressure. The ink perfusion was performed via the ascending aorta, and rat retinal microvessels were subsequently detected in whole-mount retinas and retinal sections. The second part was based on the results of the first part, in which 6 rats were infused sequentially with 20 ml ink plus 3% gelatin and then 20 ml ink plus 5% gelatin. The rat retinal microvessels were detected as that in the first part. Results Under the perfusion condition that is 40 ml of 37 ℃ ink plus 3% gelatin and 165 mmHg perfusion pressure, the vessels of the peripheral retina were completely shown, whereas only part of the central retinal vessels was shown. In contrast, the perfusion conditions that is 40 ml of 37 ℃ ink plus 5% gelatin and 215 mmHg perfusion pressure, resulted in the central retinal vessels shown very well, but not the peripheral retinal ones. However, when the animals were first perfused with 20 ml of 37 ℃ ink plus 3% gelatin at 165 mmHg perfusion pressure, and subsequently perfused with 20 ml of 37 ℃ ink plus 5% gelatin at 215 mmHg perfusion pressure, both the central retinal vessels and the peripheral retinal vessels were shown very well, without ink leakage in the retina. In addition, the contrast between vessels and surrounding tissues was distinctly improved. Conclusion Perfusing animals with 20 ml of 37 ℃ ink plus 3% gelatin at 165 mmHg perfusion pressure, and subsequently with 20 ml of 37 ℃ ink plus 5% gelatin at 215 mmHg perfusion pressure, is hereby demonstrated as good method for morphological characterization of rat retinal microvessels.