J. Dairy Sci. 86:1194–1200
American Dairy Science Association, 2003.
Disruption of Bovine Oocytes and Preimplantation Embryos
by Urea and Acidic pH1
O. M. Ocon2and P. J. Hansen
Department of Animal Sciences, University of Florida,
Feeding cattle diets high in degradable crude pro-
direct effects of urea and acidic pH during oocyte mat-
uration and embryonic development. For experiment
1, oocytes were matured in medium containing 0, 5,
respectively). Cleavage rate was not reduced by any
concentration of urea. However, the proportion of oo-
cytes developing to the blastocyst stage at d 8 after
insemination was reduced by 7.5 mM urea. In addi-
tion, the proportion of cleaved oocytes becoming blas-
tocysts was decreased by 5 and 7.5 mM urea. For ex-
periment 2, putative zygotes were collected ∼9 h after
plex Optimized Medium (KSOM). Urea did not reduce
the proportion of oocytes developing to the blastocyst
stage, although 10 mM urea reduced cleavage rate
slightly. For experiment 3, dimethadione (DMD), a
weak nonmetabolizableacid, was used todecrease cul-
ture medium pH. Putative zygotes were cultured in
modified KSOM containing 0, 10, 15, or 20 mM DMD
for 8 d. DMD reduced cleavage rate at 15 and 20 mM
and development to the blastocyst stage at all concen-
trations. Results support the idea that feeding diets
rich in highly degradable CP compromises fertility
through direct actions of urea on the oocyte and
through diet-induced alterations in uterine pH.
(Key words: urea, pH, preimplantation embryo,
Abbreviation key: COC = cumulus-oocyte com-
plexes, DMD = dimethadione, KSOM = Potassium
Simplex Optimized Medium.
Received April 5, 2002.
Accepted August 6, 2002.
Corresponding author: P. J. Hansen; e-mail: Hansen@animal.
Experiment Station. Research was supported in part by USDA-
CSREES grant # 2001-52101-11318.
vania State University, University Park, 16802-3503.
Feeding dairy cows large amounts of degradable
protein in excess of requirements can reduce preg-
nancy rates per insemination (Canfield et al., 1990;
Elrod and Butler, 1993). Diets high in protein content
elevate urea nitrogen concentrations in plasma and
uterine secretions (Jordan et al., 1983; Canfield et al.,
1990; Elrod and Butler, 1993; Roseler et al., 1993),
and elevations in blood or milk urea nitrogen concen-
trations have also been associated with low fertility
(Butler et al., 1996; Larson et al., 1996; Rajala-Schultz
et al., 2001).
The mechanism by which feeding large amounts of
protein affects fertility is not completely known. One
possibility is that high concentrations of urea associ-
ated with excess feeding of CP could disrupt oocyte
growth or maturation, fertilization, or development.
Feeding a diet generating large amounts of urea and
ammonia increased growth of the second-wave domi-
nant follicle, decreased capacity of oocytes from small
(1 to 4 mm) and medium (>4 to 8 mm) follicles to cleave
when fertilized in culture, and decreased ability of
cleaved embryos formed from oocytes obtained from
tocyst stage (Sinclair et al., 2000). Moreover, exposure
of oocytes to 6 mM of urea during maturation in vitro
impaired meiosis and fertilization rate (De Wit et
Feeding large amounts of protein can also alter the
uterine environment by reducing concentrations of
magnesium, potassium, and phosphorus in uterine se-
cretions (Jordan et al., 1983) and by reducing uterine
pH (Elrod et al., 1993; Elrod and Butler, 1993). Effects
ofprotein diet on potassium,phosphorus, andpH were
only observed during the luteal phase. Although con-
onic function are not known, a reduction in pH from
mise embryonic development in mice (Edwards et al.,
1998). In contrast, development of hamster preim-
plantation embryos was unaffected by a range of pH
from 6.5 to 7.4 (Bavister et al., 1983; Carney and
OCON AND HANSEN
and metabolism of preimplantation sheep embryos and may
affect fetal growth among survivors. Anim. Reprod. Sci.
Parrish, J. J., J. L. Susko-Parrish, M. L. Liebfried-Rutledge, E. S.
Critser, W. H. Eyestone, and N. L. First. 1986. Bovine in vitro
2001. Association between milk urea nitrogen and fertility in
Ohio dairy cows. J. Dairy Sci. 84:482–289.
Rivera, R. M., and P. J. Hansen. 2001. Development of cultured
bovine embryos after exposure to increased temperatures in the
physiological range. Reproduction 121:107–115.
Journal of Dairy Science Vol. 86, No. 4, 2003
Roseler, D. K., J. D. Ferguson, C. J. Sniffen, and J. Herrema. 1993.
Dietary protein degradability effects on plasma and milk urea
nitrogen and milk nonprotein nitrogen in Holstein cows. J.
Dairy. Sci. 76:525–534.
SAS User’s Guide, Version 5.4 Edition. 1989. SAS Inst., Inc.,
Sinclair, K. D., M. Kuran, F. E. Gebbie, R. Webb, and T. G. McEvoy.
2000. Nitrogen metabolism and fertility in cattle. II. Develop-
ment of oocytes recovered from heifers offered diets differing
in their rate of nitrogen release in the rumen. J. Anim. Sci.
Verkman, A. S., J. A. Dix, and J. L. Seifter. 1985. Water and urea
transport in renal microvillus membrane vesicles. Am. J. Phys-