Increased estrogen causes an increased turnover rate of vaginal epithelial cells, resulting in the progressive cornification
seen on vaginal cytology, and thickening of the vaginal wall in preparation for breeding. Also seen is progressive edema of
the vaginal mucosa, which can be visualized with endoscopic examination.
Estrogen assays are performed by RIA by many commercial laboratories. However, the information given is of little value
for ovulation timing, since peak estrogen levels are variable from bitch to bitch, and even relative changes do not correlate
to ovulation or the fertile period. Estrogen is best assessed by serial vaginal cytologies and vaginoscopy.
Estrogen levels do not indicate the fertile period since ovulation is triggered by the LH surge, not an estrogen peak.
Examination of the cells on the surface of the vaginal epithelium will give information about the stage of the estrous cycle.
Proper technique is important so that the cells obtained are representative of the hormonal changes occurring. The sample
should be collected from the cranial vagina, since cells from the clitoral fossa, vestibule and/or caudal vagina are not as
indicative of the stage of the cycle. Under the influence of rising estrogen levels, the number of layers making up the vaginal
epithelium increases dramatically, presumably to provide protection to the mucosa during copulation. Therefore, as estrogen
rises during proestrus, the maturation rate of the epithelial cells increases, as does the number of keratinized, cornified
epithelial cells seen on a vaginal smear. Full cornification continues throughout estrus, until the "diestral shift" that
occurs 7 to 10 days after the LH surge, signifying the first day of diestrus.
The vaginal smear then changes abruptly from full cornification to 40-60% immature (parabasal and intermediate) cells
over a 24-36 hour period. If vaginal cytology is performed until the diestral shift is observed, a retrospective analysis
of the LH surge, ovulation and the fertile period can be obtained.
Luteinizing Hormone (LH)
At the end of the follicular phase of the estrous cycle, a marked increase in LH over usual baseline values occurs over a
24-48 hour period, followed by a return to baseline values. This surge in LH is thought to take place in response to the decline
in the estrogen: progesterone ratio that occurs as estrogen levels decrease and progesterone rises. The LH surge triggers
ovulation and thus makes it the central endocrinological event in the reproductive cycle of the bitch, with all events following
being consistent between bitches. Therefore, daily serial measurement of LH to identify the exact date of the LH surge is
the most accurate diagnostic tool for timing breedings. Affordable semiquantitative in house kits are available for measuring
serum LH levels in the dog, identifying the pre-ovulatory LH surge and thus, the time of ovulation and the true fertile period.
This testing is the most accurate means of ovulation timing, and thus should be considered a "gold standard". Samples must
be drawn daily, at about the same time, for LH testing, since the LH surge may have a duration of only 24 hours in many bitches,
and could be missed if one day was skipped. The commercially available LH kits can be subject to variable operator interpretation,
thus the same person should run the tests if possible.