Relationship between colour flow Doppler sonographic assessment of corpus luteum activity and progesterone concentrations in mares after embryo transfer.

Colour-flow Doppler sonography has been described as a means of assessing corpus luteum (CL) function rapidly, because area of luteal blood vessels correlates well with circulating progesterone (P4) concentrations [P4] in oestrous cycling mares. The aim of this study was to assess the relationships between CL size and vascularity, and circulating [P4] during early pregnancy in mares, and to determine whether luteal blood flow was a useful aid for selecting an embryo transfer recipient. Equine embryos (n=48) were recovered 8 days after ovulation and were transferred to available recipient mares as part of a commercial program with the degree of synchrony in timing of recipient ovulation ranging from 1 day before to 4 days after the donor. Immediately prior to embryo transfer (ET), maximum CL cross-section and blood vessel areas were assessed sonographically, and jugular blood was collected to measure plasma [P4]. Sonographic measurements and jugular blood collection were repeated at day 4 after ET for all mares, and again at days 11, 18 and 25 after ET in mares that were pregnant. The number of grey-scale and colour pixels within the CL was subsequently quantified using ImageJ software. The CL blood flow correlated significantly but weakly with plasma [P4] on the day of transfer and on day 4 after ET in all mares, and on days 11 and 25 after ET in pregnant mares (r=0.30-0.36). The CL area and plasma [P4] were also correlated on each day until day 11 after ET (r=0.49-0.60). The CL colour pixel area decreased significantly after day 18, whereas CL area was already decreasing by day 4 after ET. The CL area, area of blood flow, or [P4] was predictive of pregnancy. Findings in the present study suggest that both CL area and blood flow are correlated with circulating [P4] at the time of transfer and in early pregnancy. Evaluation of the CL using B-mode or CF sonography, although practical, provides no improvement in the selection of recipients or prediction of pregnancy outcomes than methods employed currently.

Liquid storage of equine semen: Assessing the effect of d-penicillamine on longevity of ejaculated and epididymal stallion sperm.

Short-term storage of equine sperm at 5°C in an extender containing milk and/or egg yolk components is common practice in the equine breeding industry. Sperm motility, viability, DNA integrity and, consequently, fertilizing ability decline over time, partly due to reactive oxygen species (ROS) generation. We investigated whether adding the anti-oxidant d-penicillamine to a commercial milk/egg yolk extender delayed the decrease in semen quality. Semen was recovered on four consecutive days from eight 3-year old Warmblood stallions. On day 5, seven of the stallions were castrated and sperm recovered from the caudae epididymides. Ejaculated samples were split, and one portion was centrifuged and re-suspended to reduce seminal plasma content. All samples were diluted to 50millionsperm/ml and divided into two portions, one of which was supplemented with 0.5mM d-penicillamine. After 48h, 96h, 144h and 192h storage, sperm motility was assessed by computer-assisted semen analysis (CASA), viability by SYBR14/PI staining, and DNA integrity using the sperm chromatin structure assay (SCSA). d-Penicillamine had no effect on motility of ejaculated sperm (P>0.05) but reduced total and progressive motility of epididymal sperm. Sperm chromatin integrity was not influenced by storage time, seminal plasma or d-penicillamine. In short, adding d-penicillamine to a commercial semen extender was neither beneficial nor detrimental to the maintenance of quality in ejaculated semen stored at 5°C. The negative effect on motility of epididymal sperm may reflect differences in (membrane) physiology of spermatozoa that have not been exposed to seminal plasma.