Danias John, Shen Fran, Goldblum David, Chen Bin, Ramos-Esteban Jerome, Podos Steven M, Mittag Thom
Department of Ophthalmology, Mt. Sinai School of Medicine, 1 Gustave L. Levy Place, New York, NY 10029, USA.
Invest Ophthalmol Vis Sci. 2002 Mar;43(3):587-94.
To determine the number and cytoarchitecture of retinal ganglion cells (RGCs) in the female Wistar rat, by using a newly devised procedure for rapid RGC counting in the entire retina that avoids assumptions about RGC spatial arrangement.
RGCs of normal female Wistar rats were retrogradely labeled with a fluorescent tracer. Automated counting was accomplished by applying standard imaging software to analysis of all labeled cells in retinal flatmounts. The method was validated by comparison of automated and manual counts of 70,000 RGCs in frames covering the density range in the normal rat retina of 600 to 3600 RGC/mm(2). RGC numbers were determined for each retina and compared with the contralateral retina of the same animal. RGC density maps were constructed for each retina. RGC size distribution was determined.
Automated RGC counting showed a good linear correlation with manual counting (R(2) = 0.9416). Mean total RGC count in 10 rat eyes was 97,609 +/- 3,930 (SEM) per eye. Contralateral eyes differed by an average of 4.1% (3983 plus minus 5098 RGCs). Size analysis calculated from cell areas confirmed that the majority of rat RGCs are between 7 and 21.5 microm in equivalent diameter. The RGC counts for all frames at the same eccentricity in all 10 of the retinas showed that variability increased with eccentricity and increased further as the fractional area of the retina sampled at each eccentricity was reduced. There was also significant variability in the spatial density of the RGCs at the same eccentricity location between different eyes. Comparison of total RGC counts between left and right eyes estimated from RGC counts in sectors of the retina (hemiretinas or quadrants) showed increased variability compared with counting all the RGCs in a retina.
RGCs in the Wistar rat display significant variability in their cytoarchitecture. Such variability can make quantification by sampling problematic for diffuse, and particularly, for focal RGC losses resulting from experimental interventions, unless virtually the entire RGC population is counted.
通过使用一种新设计的在整个视网膜中快速计数视网膜神经节细胞（RGC）的方法，该方法无需对RGC的空间排列做假设，来确定雌性Wistar大鼠视网膜神经节细胞的数量和细胞结构。
用荧光示踪剂对正常雌性Wistar大鼠的RGC进行逆行标记。通过应用标准成像软件对视网膜平铺标本中所有标记细胞进行分析来完成自动计数。通过比较覆盖正常大鼠视网膜中600至3600个RGC/mm²密度范围的帧中70,000个RGC的自动计数和手动计数，对该方法进行了验证。确定每个视网膜的RGC数量，并与同一只动物的对侧视网膜进行比较。为每个视网膜构建RGC密度图。确定RGC大小分布。
RGC自动计数与手动计数显示出良好的线性相关性（R² = 0.9416）。10只大鼠眼睛的平均RGC总数为每只眼睛97,609 ± 3,930（SEM）。对侧眼睛平均相差4.1%（3983 ± 5098个RGC）。根据细胞面积计算的大小分析证实，大多数大鼠RGC的等效直径在7至21.5微米之间。在所有10个视网膜中，相同偏心率下所有帧的RGC计数表明，变异性随偏心率增加，并且随着在每个偏心率下采样的视网膜部分面积减小而进一步增加。在不同眼睛的相同偏心率位置，RGC的空间密度也存在显著变异性。根据视网膜扇形区域（半视网膜或象限）中的RGC计数估计左右眼之间的RGC总数比较显示，与计数视网膜中的所有RGC相比，变异性增加。
Wistar大鼠的RGC在细胞结构上表现出显著变异性。这种变异性会使通过采样进行量化对于弥漫性尤其是实验干预导致的局灶性RGC损失存在问题，除非几乎对整个RGC群体进行计数。
