Sperm association in the rat epididymis

During dissolution of drops obtained by puncture of rat epididymis in distal caput, mid corpus, proximal and distal cauda we observed in the last tworegions the appearance of a new sperm association which we call rosette and the formation in the same two regions of dense clusters of non-motile degenerating spermatozoa already described in the rat was deferens. Rosettesbecome organized in the proximal cauda by head to head sperm contact through a filamentous PAS positive material. By video microscopy was observed thedilution of a dense drop of cauda epididymal content in a balanced salt solution at 36 degrees C. In less than 3 min, rosettes become dispersed intosingle sperm with a display of intense motility which starts immediately after dilution. A glycoprotein with cohesive properties secreted by the epididymal epithelium is postulated as the factor promoting rosette formation. We confirm that dense clusters correspond to degenerating spermatozoa and describe its first appearance in the epididymal proximal cauda with increase in number toward was deferens

The zona-free hamster oocyte penetration test: a simple procedure using a DNA fluorescent stain to detect the male pronucleus formation

In order to increase the value of the zona-free hamster oocyte penetration test, a comparetively simple and fast method using the fluorochrome Hoechst 33342 was developed. human spermatozoa were washed and incubated 1 hr mediumBWW for capacitation. hamster oocytes were stripped of cumulus oophorus and zona pellucida with hyaluronidase and trypson, washed and used immediately. Thirthy oocytes were placed in a drop of BWW containing 3,5.10(6)/ml of human spermazoa under mineral oil. The sperm-oocyte preparation was incubated for 3 hr at 37 degrees C, during the last 15 min of incubation, the fluorochrome Hoechst 33342 (H) was added and incubation was allowed to proceed until the incubation time was over. Observations showed that the female pronucleus, eccentrically placed, gives a bright green-bluish fluorescence whereas chromatin of sperm heads shows different stages of decondensation and also a bright fluorescence. This inexpensive method has given consistent results in a large number of cases and provides an additional new approach to the <> as a proof of the capacity to form a <>