312 MORPHOLOGICAL AND MORPHOMETRICS CHARACTERISTICS OF NUCLEAR SPERM IN THE EPIDIDYMAL TRANSIT OF BULLS

Abstract

During the transit through the epididymis, many morphological, physiological, and biochemical characteristics of spermatozoa are modified, as part of the maturation process. Nuclear maturation continues in the epididymis through an increase in formation of protamine disulfide. Thus, penetration through the oocyte membranes could be facilitated for elongated spermatozoa with dramatically condensed chromatin and nuclear integrity. Therefore, size, shape, and nuclear defects could be used to estimate the stage-related nuclear transformations from early spermiogenesis to the end of epididymal transit. Nellore bulls (n = 9), 30-36 months old, with high seminal quality (>80% motile and morphologically normal sperm) were submitted to orchiectomy. Impressions in slides of the caput, corpus, and caudal regions of the epididymis were prepared for evaluation of morphology (Feulgen stain, phase-contrast microscopy at 1000 ×) and morphometry of the nuclear sperm. The slides were captured in a Motic 2300 camera adapted to the microscope and digitally assessed. Nuclear morphology was considered normal (without visible alterations), head defect (variations in shape and form), and nuclear defects (abnormal chromatin condensation and presence of vacuoles). Base, width, length (μm), and area (μm²) were estimated in least 60 sperm nuclei. No difference (P > 0.05) among epididymal regions for normal nuclei (70.3 ± 3.1%), head defects (3.4 ± 0.5%), and nuclear defects (5.3 ± 1.3%) were seen. The base was higher (2.68 ± 0.5 μm, P < 0.01) in the caput than corpus (2.44 ± 0.4 μm) and caudal regions (2.41 ± 0.4μm). Normal nuclei were associated (P < 0.01) with width (r = 0.20), length (r = 0.27), and area (r = 0.44) in the caput and with width (r = 0.21), length (r = 0.40), and area (r = 0.33) in the corpus of epididymis. Epididymal transit affected (P < 0.001) the measures and nuclear status that accounted for regression analysis: normal nucleus (40.254 + 3.027, length; R² = 0.20), head defects (0.922 + 1.097, width + 0.093 × area; R2 = 0.26), and nuclear defects (6.993-0.496, length + 0.454, base; R² = 0.23). The results suggest that important variations occur in the nuclear status during the epididymal transit in the bovine spermatozoa. The higher measures in the sperm base of the caput suggest a narrowing probably indicating the continuity of nuclear remodeling from the final steps of spermiogenesis. The area accounts for 20% of the nuclear shape variations along the epididymal segment. These events characterize the adaptation of nuclear membranes and chromatin structure surrounded by epididymal environment and comprise a part of the maturation process. Additionally, morphometric variations are associated with defects in the nuclear structures and can be used to determine the conditions of spermatogenesis and sperm maturation based on the evaluation of ejaculated semen.

We thank CNPq/PROPP and Fundect for financial support.