The measurement of plasma progesterone levels in the normal mare and its application to some equine breeding problems : a thesis submitted in partial fulfilment of the requirements for the degree of Master of Veterinary Science at Massey University
A radioimmunoassay system was developed to measure plasma concentrations of endogenously produced and exogenously administered progesterone in non-pregnant and pregnant mares in normal and some abnormal reproductive states. Assay sensitivity was 0.5 ng/ml, with a between assay coefficient of variation of 16.8% for a high progesterone sample, estimated over 24 assays, and 8.5% for a low progesterone sample, estimated over 15 assays; the within assay coefficient of variation was 7.3, 10.1 and 6.9% respectively for six replicates of one sample, estimated in three separate assays. Plasma progesterone concentrations of six normal non-pregnant cycling mares followed regular cyclic changes, with levels less than 0.5 ng/ml during oestrus and ranging between 8-22 ng/ml at peak values in dioestrus. The first oestrus following the winter non-breeding period was longer than the following oestrus and the period from ovulation to progesterone decline tended to be less variable than the rest of the cycle. There was a large between-mare variation in plasma progesterone concentrations in mares at both early and late stages of gestation with levels varying from 4.9 to 15 ng/ml in the former and 5.2 to 16.9 ng/ml in the latter group. No significant effect was noted between stage of gestation and progesterone concentration. A group of five mares all had plasma progesterone concentrations greater than 9.5 ng/ml within 24 hours prior to parturition; two of these mares sampled within eight hours prior to parturition had plasma progesterone concentrations of 4.3 and 3.9 ng/ml. The first post partum sample was taken within 24 hours of foaling; by this time plasma progesterone concentrations had fallen to less than 0.5 ng/ml and remained low until sampling ceased at the first post partum oestrus. Prostaglandin F2α (THAM salt) was effective in causing luteolysis in 13 mares with active corpora lutea before treatment. By three days post-injection 12 of the 13 mares had plasma progesterone concentrations of less than 0.5 ng/ml and by five days post-injection 12 of the 13 mares were exhibiting oestrus. Of the ten mares bred at the induced oestrus, seven became pregnant to that mating. Plasma progesterone concentrations were measured on 16 non-pregnant mares in anoestrus. Six of eight mares sampled early in the breeding season (September) had plasma progesterone concentrations of less than 0.5 ng/ml, the other two mares had plasma progesterone concentrations of 0.6 ng/ml. Eight of eight mares sampled later in the breeding season (November and December) had plasma progesterone concentrations greater than 0.5 ng/ml, the levels ranging from 6.2 to 13.1 ng/ml. Concentrations of plasma progesterone in normal dioestrous mares were measured half and one hourly (three mares) for 24 hours and four hourly (two mares) for 120 hours. There were large apparently random variations, with more than 100% differences being recorded between a number of consecutive samples. Plasma progesterone concentrations varied from 7.8-23.0, 3.2-21.9 and 4.2-12.9 ng/ml for the three mares sampled half hourly and hourly, and from 6.8-24.6 and 0.8-11.0 ng/ml for the two mares sampled four hourly. Radioactive progesterone, administered by venepuncture to a mare with no detectable endogenous plasma progesterone, disappeared from the plasma within 40 minutes; 85% of the injected steroid had left the plasma by 2.5 minutes post-injection. Two and 25 mg of progesterone in 16% alcohol in saline was administered by venepuncture to mares with plasma progesterone concentrations of less than 0.5 ng/ml. For the mare given 2 mg, the plasma half life of injected progesterone was 1.75 minutes for the initial "fast" component, and for the mare given 25 mg the plasma half life was 2.75 minutes. There was a second peak of plasma progesterone at from 8 to 19.5 minutes for the former and from 9.5 to 17 minutes for the latter mare. A third much smaller peak was recorded at about 50 minutes post-injection for the mare given 2 mg progesterone. A mare with no detectable endogenous plasma progesterone was administered by intramusuclar injection a total of 600 mg progesterone in arachis oil over a period of seven days. Plasma concentrations of the steroid reached a maximum of 4.3 ng/ml at one day post-treatment and were maintained at this level for only a maximum of 24 hours. A second mare, again with no detectable plasma progesterone, was administered by intramuscular injection a total dose of 2 g of hydroxyprogesterone capronate in castor oil over a period of ten days. Maximum plasma progesterone concentrations of 1.2 ng/ml, maintained for less than 24 hours, were reached nine days after the first injection. Wide variation in plasma progesterone levels within and between mares over relatively short time periods suggest that there are many difficulties in identifying "progesterone insufficiency" as a cause of embryonic absorption or abortion in this species. Moreover the short half life of this steroid in the plasma of the mare, together with the sustained high dose levels that would be required to elevate plasma concentrations of progesterone to a level equivalent to that produced by normal secretory corpora lutea, indicate that current levels of administration of this drug are likely to have little effect in overcoming such a breeding problem unless the progesterone is acting at a local level. A definitive answer in respect to this vexed question concerning the existence or not of "progesterone insufficiency" as a cause of prenatal loss in the mare, together with an appropriate method of treatment, still remains to be found.