Abstract
A modified lectin angiography method was applied to identify ovarian vasculature in 32-34 days old mice treated with gonadotropins. Blood was washed out by perfusion until the inferior vena cava became translucent. Horseradish-peroxidase (HRP)-conjugated Concanavalin A (Con A) solution (10-15 ml) was perfused into the systemic circulation via the left ventricle at the rate of 2-3ml/min. The animals were detained for a 30 min reaction interval. The lumina of the blood vessels were flushed with 5-10 ml of phosphate buffered saline (pH 7.4). The ovaries were excised and fixed by immersion in 4% paraformaldehyde and sectioned serially at thickness of 100, 200 or 400μm using a Microslicer. The binding of HRP-conjugated-Con A to endothelial cells was visualized by using 3, 3'- diaminobenzidine-4 HCl (DAB-4 HCl) reaction. By examining various sections the three-dimensional architecture of the vascular networks of the preovulatory Graafian follicles and corpora lutea can be established. Capillary wreaths of preovulatory Graafian follicles were identified in sections of ovaries removed 11 h after hCG injection. Then, capillary wreaths of the surrounding Graafian follicles became elevated due to the hypertrophic growth of the theca interna and extension of capillary branches into the follicles. Well-developed capillary networks of corpora lutea were found in ovaries removed 24 h after hCG injection. For these observations the 200μm-section was the most useful. Taken together, the present modified lectin angiography is a useful method for visualizing the microvasculature of mouse ovaries.
