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P. T. K. Saunders, P. M. Summers, and J. P. Hearn

Summary. A biologically active gonadotrophin has been partly purified from the media of long-term cultures of trophoblast cells of the common marmoset monkey by a combination of precipitation and chromatography. Marmoset chorionic gonadotrophin (CG) is a glycoprotein which binds Concanavalin A and wheat germ agglutinin. The protein purified from culture media exists as several isoelectric species with pI in the range pH 3·5–4·5. On gel filtration it eluted with an apparent molecular weight of 68–72 000 but on PAGE migrated as if it was 58–65 000. A glycoprotein with similar characteristics has been recovered from plasma samples of pregnant marmosets. Biological activity of partly purified CG from media, as determined by a mouse testicular cell bioassay, was 1–3 i.u./mg protein.

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P. M. Summers, C. T. Taylor, and M. W. Miller

The role of the inner cell mass in the induction of chorionic gonadotrophin synthesis and secretion by the trophoblast of the peri-implantation primate blastocyst was studied in common marmoset monkeys. An in vitro system for the culture of blastocysts commencing with blastocysts collected 8 days after conception was developed. Chorionic gonadotrophin measured in the spent culture fluid was first detected in most blastocysts after 3 or 4 days (day 11 or 12) of culture at a time equivalent to implantation in vitro. Initial secretion of chorionic gonadotrophin coincided with development of parietal endoderm and histological appearance of syncytiotrophoblast in the polar trophoblast. Little chorionic gonadotrophin was secreted by blastocysts with a poorly developed, or absent, inner cell mass. Mural trophoblast removed from blastocysts after 2 days of culture (day 10) grew in vitro as a unilaminar vesicle but failed to secrete significant amounts of chorionic gonadotrophin. However, mural trophoblast from older blastocysts (days 13 and 14) after chorionic gonadotrophin secretion had commenced continued to secrete chorionic gonadotrophin, with trophoblast from day 14 blastocysts secreting significantly more than that from day 13. It was concluded from these studies that while mural trophoblast from marmoset blastocysts will proliferate in vitro in the absence of an inner cell mass, efficient induction of chorionic gonadotrophin secretion requires the presence of the inner cell mass or its derivatives. Once chorionic gonadotrophin secretion has commenced, secretion will continue in the absence of the inner cell mass.

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P. M. Summers, Caroline J. Wennink, and J. K. Hodges

Summary. A single intramuscular injection of 0·5 μg cloprostenol was not luteolytic on Day 6 or 7 of the ovarian cycle (N = 3), but was luteolytic in some animals (3/5) on Day 8 and 9 and luteolytic in all 23 animals treated between Days 10 and 17 of the ovarian cycle, and in 7 animals treated between Days 19 and 43 of pregnancy. Luteal function was monitored by measurement of progesterone in peripheral blood using a simple and rapid non-extraction assay. There was a dramatic fall in peripheral blood progesterone to <10 ng/ml within 24 h of cloprostenol injection; progesterone remained at this low level until the day after post-treatment ovulation. The interval from cloprostenol injection to ovulation in animals treated between Days 8 and 17 was 10·7 ± 0·3 days. A similar interval was found in pregnant animals. Embryos recovered from the uterus after cloprostenol treatment were morphologically normal (23/24).

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G. E. Webley, M. C. Richardson, P. M. Summers, A. Given, and J. P. Hearn

Summary. Dispersed marmoset luteal cells were incubated for 2 h and progesterone production measured after exposure to hCG, cloprostenol, dibutyryl cAMP, PGF-2α, PGF-2, adrenaline or melatonin. The cells were studied on Days 6, 14 and 20 after ovulation in conception and non-conception cycles. Luteal cells from Day 14 non-pregnant marmosets were compared with human luteal cells taken in the mid-luteal phase. All the treatments stimulated progesterone production including cloprostenol, which is luteolytic when administered to the marmoset in vivo, but the degree of response varied with the stage of the cycle or pregnancy and between marmoset and human luteal cells. In the marmoset, overall analysis of the effect of the treatments showed that, on Day 6 after ovulation, there was no significant effect of any of the treatments in cells from pregnant or non-pregnant animals. In contrast, luteal cells from non-pregnant animals on Day 14 showed a significant response to the treatments (F (8,41) = 2·79, P < 0·0145) whereas cells from pregnant Day-14 animals were responsive; in cells from pregnant animals, the control production of progesterone was high and already equivalent to the levels stimulated by the treatments. By Day 20, cells from pregnant animals produced lower control concentrations of progesterone than did those on Day 14 and there was a significant overall effect of the treatments (F (8,33) = 3·78, P < 0·003). These results show that the marmoset CL gains responsiveness to treatment between Days 6 and 14 after ovulation in the non-pregnant cycle. In pregnancy, on Day 14, 2 days after attachment of the embryo, the high control concentrations of progesterone and absence of response to treatment suggest that an embryo message may have affected the CL, providing an endogenous stimulus.

Keywords: marmoset monkey; human; luteal cells; luteotrophic and luteolytic agents; early pregnancy

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P. M. Summers, A. M. Shephard, C. T. Taylor, and J. P. Hearn

Summary. Embryos were collected at the 4–10-cell stage from the oviducts (Day 4; Day 1 = ovulation) or as morulae (Day 7) from the uterus of marmosets and frozen in 1·5 m-DMSO (Days 4 and 7) or 1·0 m-glycerol (Day 4 only), using a slow freezing and thawing technique. Of 22 Day-4 embryos frozen in DMSO, 18 were recovered and 16 of these were transferred to 10 synchronized recipients; 7 recipients became pregnant compared with all 7 control recipients receiving 10 unfrozen embryos. Fifteen frozen– thawed morulae were transferred to 9 Day-6 recipients; the pregnancy rate (55·6%) was lower than for control embryos (85·7%). Embryos frozen in glycerol suffered severe osmotic stress during glycerol addition and removal. Of 8 recipients, 3 (37·5%) became pregnant but only one fetus was carried to term.

These results on embryo collection, freezing and transfer in the marmoset have important implications for developing improved methods for freezing human embryos and the breeding of endangered primates.

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P. M. Summers, A. M. Shephard, J. K. Hodges, J. Kydd, M. S. Boyle, and W. R. Allen

Summary. Blastocysts were collected non-surgically from 2 Przewalski's horse and 2 Grant's zebra mares and transferred extra-specifically to domestic horse and donkey recipients. Nine Przewalski's horse embryos were transferred surgically, and 2 nonsurgically, to domestic Welsh-type pony mares. After surgical transfer, 7 (77·8%) pregnancies were established and 4 foals were born. Twelve Grant's zebra embryos were transferred surgically to 5 pony and 7 domestic donkey recipients respectively and 1 non-surgically to a donkey; 3 (60%) zebra-in-horse pregnancies were established and 2 went to term. Only 2 (28·6%) zebra-in-donkey pregnancies were established but neither went to term, although one zebra foal was aborted alive at Day 292 but failed to survive. No pregnancies resulted from the non-surgical transfers. Measurement of chorionic gonadotrophin concentrations and parental-specific lymphocytotoxic antibodies in the serum of the recipient animals indicated a pronounced maternal immunological response to the extra-specific embryo, but this could not be correlated with success or failure of pregnancy.

The results indicate that extra-specific embryo transfer may be a useful aid to breeding exotic equids in captivity.