226 EFFECT OF AMMONIA IN BOVINE FOLLICULAR FLUID AND OOCYTE MORPHOLOGY DERIVED FROM ABATTOIR OVARIES

Abstract

In cattle, high intake of crude protein elevates rumen ammonia and blood urea nitrogen and has been associated with decreased fertility. The aim of this study was to analyze the concentration of ammonia in bovine follicular fluid derived from abattoir ovaries and to examine the relation between ammonium concentration and morphology of oocytes from antral follicles. In the first experiment, follicular fluid was collected within 1 h of slaughter from 3919 small (2-5 mm in diameter), 366 medium (6-9 mm), and 159 large (10-20 mm) follicles from 203 ovaries. Ammonium concentration was analyzed in 6 batches pooled from follicular fluids of each size (derived from 15, 39, 7, 36, 81, and 25 ovaries, respectively). Ammonium concentration was measured by the microdiffusion method using an Ami Check Meter kit (Kyoto Daiichi Kagaku Co., Ltd., Japan). In the second experiment, follicular fluids were aspirated from small (2-5 mm) follicles of 38 cows to evaluate the relationship between ammonium concentration and oocyte morphology. Oocytes were classified into 4 morphological categories according to the surrounding cumulus cells (Konishi M et al. 1996 Theriogenology 45, 573-581): grade 1 (≥5 layers); grade 2 (3-4 layers); grade 3 (1-2 layers); and grade 4 (denuded, degenerated cumulus cell mass or ooplasm). The oocyte morphology for the low-ammonium group (<330 μmol, 239 oocytes from 19 cows) and high-ammonia group (≥330 μmol, 255 oocytes from 19 cows) was compared. Statistical significance was tested by chi-square test or t-test Ammonium con- centrations of small (281.8 ± 20.0 μM), medium (175.4 ± 15.8 μM), and large (96.3 ± 9.3 μM) follicles were significantly different (P < 0.0001). Ammonium concentrations in follicular fluid of the 38 cows ranged from 180.6 to 802.4 μM. In a comparison of oocyte morphology between the low- (263.8 ± 41.6 μM) and high-ammonium (423.0 ± 108.9 μM) groups, the number of grade 1, 2, 3, and 4 oocytes in the former (80, 68, 18, and 73, respectively) were not significantly different from the numbers in the latter (96, 61, 27, and 71, respectively; P = 0.30-0.99). These results indicate that, in follicles of different sizes derived from abattoir ovaries, the ammonium concentration was higher in small follicles than in the other sizes of follicles. However, there was no evidence that in vivo exposure of bovine oocytes to ammonium badly influenced oocyte morphology. Further studies are needed to examine the relationship between ammonium concentration and the developmental potential of oocytes.