Analysis on regulation of renal microcirculation with an in-vivo visualisation system

肾脏微循环的可视化研究目前仅限于离体肾单位样本、病理性肾积水模型或颊囊移植模型。这是因为在生理条件下进入肾微循环在技术上非常困难。我们已经克服了这一障碍，通过开发一个CCD内视频显微镜。为了进一步研究肾小球功能，我们已经开始应用荧光进行可视化。本研究的目的是可视化肾脏微循环的活体视频显微镜和双光子显微镜在两周STZ诱导的糖尿病大鼠。60 mg/kg的STZ可使大鼠发生中度糖尿病。糖尿病早期大鼠肾小球小动脉直径（aff）14.0 ± 1.9 μ m，eff9.4 ±0.7μ m，明显大于正常对照组（aff 11.9±0.8 μ m，eff8.8 ±0.7 μ m）（P<0.05）。糖尿病大鼠每100 g肾重量的肾流量较大，这是用跨声波流量计测量的。用FITC葡聚糖（500 k）和罗丹明葡聚糖（20 k，尺寸小于5 nm）在双光子显微镜下观察糖尿病大鼠的肾滤过。FITC葡聚糖留在肾小球循环中，而罗丹明葡聚糖被过滤出肾小球。注射罗丹明B葡聚糖后约60秒可见肾小管。糖尿病大鼠的亮度大于正常对照大鼠。两者合计，超滤观察早期糖尿病将阐明生理和分子生物学活体视频显微镜和双光子显微镜。

Visualisation studies on renal microcirculation so far have been limited on isolated nephron samples, pathological hydronephrotic kidney models or cheek pouch transplant models. It is because making an access to the renal microcirculation under physiological conditions is technically very difficult. We have overcome this obstacle by developing a CCD intravideomicroscope. To investigate glomerular functions further, we have started applying fluorescence for visualisation. The purpose of this study is to visualise renal microcirculation by intravital videomicroscopy and two-photon microscopy in two-week STZ-induced diabetic rats. With 60 mg per kilogramme of STZ, rats become diabetic to the moderate degree. Blood glucose level after 2 weeks is about 400 mg/dl.We found with our intravital videomicroscope that diameters of glomerular arterioles in early diabetic rats (afferent (aff) 14.0 ± 1.9, efferent (eff) 9.4±0.7μ m) were larger than those of normal control (aff 11.9±0.8, eff 8.8±0.7μ m (p<0.05). Renal flow per 100 g kidney weight was larger for diabetic rats, which was measured with a transonic ultrasound flowmeter. From these results, diabetic rats were under hyperfiltrated state.Such renal filtration was visualised with a two-photon microscope administering FITC dextran (500k) and Rhodamine dextran (20k, size less than 5 nm). FITC dextran stayed in the glomerular circulation, while rhodamine dextran was filtered out of the glomeruli. The renal tubules were visualized about 60 seconds after rhodamine B dextran injection. The brightness is greater for diabetic rats than normal control rats. Taken together, hyperfiltration observed in early diabetes will be elucidated physiologically and molecular biologically by intravital videomicroscopy and two-photon microscopy.