Progress 09/15/01 to 09/30/04
Outputs
We have shown that cows Days 22, 25, 30 and 40 of pregnancy respond to oxytocin (OT) by secretion of prostaglandin F2 alpha, which is very surprising since the effects of interferon-tau secreted by the conceptus were believed to prevail until around Day 30. In an attempt to understand this enigma the target for OT action at this stage of pregnancy was determined by ascertaining which tissues contain the oxytocin receptors (OTR) and the prostanoid generating enzymes prostaglandin H synthase-2 (PGHS-2), prostaglandin E synthase (PGES) and prostaglandin F synthase (PGFS). Respective mRNA was measured by Real Time PCR in caruncular and intercarcuncular endometrium in cows Days 7, 14, 18, 22, 30, 40 and 50 of pregnancy and Days 0, 3, 7, 14, 18 and 21 of the estrous cycle. As expected OTR mRNA was high in cows at proestrus and estrus and low in pregnant cows at comparable times. Post-implantation, relative abundance of OTR mRNA increased starting Day 30 paralleling increased
responsiveness to OT. Earlier we had shown that OT induces expression of PGHS-2 mRNA. However, in this study when OT mRNA was low, PGHS-2 was highly expressed starting on Day 14 of pregnancy. Expression of endometrial PGHS-2 mRNA was higher in pregnant cows than on comparable days of the estrous cycle, i.e., Days 14, 18 and 21. Expression of PGES mRNA was high at estrus, low during luteal phase and decreased during the peri-implantation period. Whereas, PGFS mRNA was expressed highly on Days 7 and 18 of the estrous cycle and comparable days of pregnancy, but was down-regulated later with advancing pregnancy toward Day 50. The presence of immunoreactive OTR, PGHS-2, PGES, and PGFS was detected in the uterine luminal and glandular epithelium that paralleled their respective mRNA. Increased expression of PGES at estrus may be involved in ovulation and stimulation of increased blood flow to the uterus. Increased PGHS-2 and decreased expression of PGFS during the peri-implantation period
may enhance maintenance of pregnancy. Decline in endometrial PGES mRNA, however, was unexpected when considering that PGE is a luteotropic factor. These expression patterns may reflect an increasing ratio of PGE/PGF2 alpha and this changing ratio may be favorable to pregnancy. Ligand that induces elevated PGHS-2 mRNA during early pregnancy may not be OT since there is low OTR mRNA during this time. Nonetheless, the OT/PG system is functional during the peri-implantation period of pregnancy. Additional hypothesis is that between Days 20 and 30 of pregnancy a source of OT to drive the secretion of PG is the corpus luteum. Luteal OT mRNA was present at least through Day 22 of pregnancy and Day 18 of the cycle. No detection of luteal OT mRNA was observed at estrus (corpus albicans) or Days 30, 40 and 50 of pregnancy. However, another luteal peptide, relaxin-like factor (RLF)/insulin-like factor (INSL-3) mRNA was observed on all days studied except Day 3. The signal increased with
advancing pregnancy. Although OT was observed in steroidogenic large luteal cells the RLF was localized to nonsteroidogenic cells yet to be identified. Role of highly expressed RLF in peri-implantation cow is unknown.
Impacts
Embryonic loss following the period of early maternal recognition signaling is estimated to range from 6 to 46%. The well managed beef cow shows the lowest losses whereas the high producing dairy cow with marginal management has shown the highest losses. The etiologies of these losses can be quite varied. Identification of factors that influence the prostanoid-generating system as well as the time of their action might be relevant to early embryonic losses providing valuable information on the underlying causes. Perhaps this information could also provide an opportunity to mitigate the system by appropriate management protocols such as in the form of feed additives, forage composition, and milking schedules, to name a few. A fine tuned oxytocin prostanoid generating system may aid in conceptus implantation and its ultimate survival. However, an over stimulated system could be detrimental to conceptus survival. This could be the case when the system is too robust for
marginally developed conceptuses that involve either inherent developmental factors or environmental factors such as a toxin in feed that stimulates the system. At the time of embryo death a corpus luteum may still be functional. The presence of an oxytocin prostanoid system during this critical window of implantation could result in clearing the fragmented conceptus from the uterus, regression of the corpus luteum and finally a return to cyclicity. This would provide a renewed opportunity for pregnancy to ensue.
Publications
- Nicole N., H. Binta, P.A. Fields, M. Drost, S-M. Chang, R. Ivell, M.J. Fields. 2005. Immunohistochemical Localization of Relaxin-Like Factor/Insulin-Like Peptide-3 in the Bovine Corpus Luteum. Ann. N.Y. Acad. Sci. 1041:506-509
- Fields, M.J., M. Shemesh. 2004. Extragonadal LH Receptors in the Reproductive Tract of Domestic Animals. Biol. Reprod. 71:1412-1418.
- Binta H., A-R. Fuchs, M. Drost, W.W. Thatcher, N. Denslow, N. Nichols, S-M. Chang, E. Johnson, M. J. Fields. 2003. Expression of Oxytocin Receptor, Prostaglandin (PG) H Synthase (S)-2, PGE-S and PGF-S in Endometrium During the Estrous Cycle and Peri-Implantation in Cattle. Biol. Reprod. 62(Suppl 1):204.
- Nichols N., H.P. Binta, P.A. Fields, M. Drost, M. Campbell-Thompson, R. Ivell, S-M. Chang, E. Johnson, M.J. Fields. 2003. Expression and Localization of Oxytocin and Relaxin-Like Factor During the Peri-Implantation Period of Bovine Pregnancy. Biol. Reprod. 62(Suppl 1):542.
- Fields, M,J., M. Shemesh. 2003. Expression and Functionality of Uterine LH Receptors During the Estrous Cycle of Domestic Animals. Biol. Reprod. 62(Suppl 1):98.
- Shemesh, M., D. Mizrahci., M. Gurevich, L.S. Shore, J. Reed, S-M. Chang, W.W. Thatcher, M.J. Fields. 2002. Expression of Functional Luteinizing Hormone (LH) Receptor and Its Messenger Ribonucleic Acid in Bovine Endometrium: LH Augmentation of cAMP and Inositol Phosphate In Vitro and Human Chorionic Gonadotropin (hCG) Augmentation of Peripheral Prostaglandin In Vivo. Reprod. Biol. 1:13-32.
- Fuchs, A.R., L.G. Graddy, A.A. Kowalski, M.J. Fields. 2002. Oxytocin Induces PGE2 Release From Bovine Cervical Mucosa In Vivo. Prostaglandin. 70:119-129.
- Landaeta-Hernandez, A. J., J.V. Yelich, M.J. Fields, P.J. Chenoweth. 2002. Environmental, Genetic and Social Factors Affecting the Expression of Estrus. Theriogenology 57:1357-1370.
- Fields, M.J., J.V. Yelich, R.S. Sand. 2002. (Editors) Factors Affecting Calf Crop: Biotechnology of Reproduction. CRC Press, Inc., Boca Raton, FL.
- Fuchs, A.R., R. Ivell, N. Ganz, M.J. Fields. 2001. Secretion of Oxytocin in Pregnant and Parturient Cows: Corpus Luteum May Contribute to Plasma Oxytocin at Term. Biol. Reprod. 65:1135 -1141.
- Shemesh M., D. Mizrachi, M. Gurevich, Y. Stram, L.S. Shore, M.J. Fields. 2001. Functional Importance of Bovine Myometrial and Vascular LH Receptors and Cervical FSH Receptors. Seminars Reprod. Med. 19:87-96.
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Progress 10/01/02 to 10/01/03
Outputs
We have shown that cows on Days 22, 25, 30 and 40 of pregnancy respond to oxytocin by secretion of prostaglandin F2-alpha, which is very surprising since the effects of interferon-tau secreted by the conceptus were believed to prevail until around Day 30. One of our initial objectives was to determine the tissue target for oxytocin action as this stage of pregnancy by ascertaining which tissue oxytocin receptors and the prostanoid generating enzymes prostaglandin H synthase-2 (PGHS-2), prostaglandin E synthase (PGES) and prostaglandin F synthase (PGFS) respective mRNA were measured by Real Time PCR in the caruncular and intercaruncular endometrium in cows days 7, 14, 18, 22, 30, 40, and 50 of pregnancy and days 0, 3, 7, 14, 18 and 21 of the estrous cycle. As expected oxytocin receptor mRNA was high in cows at proestrus and estrus and low in pregnancy cows at comparable times. Post implantation, the relative abundance of oxytocin receptor mRNA increased starting day 30
paralleling the increased responsiveness to oxytocin. Earlier we had shown that oxytocin induces expression of PGHS-2 mRNA. However, in this study when oxytocin mRNA was low, PGHS-2 was highly expressed starting on day 14 of pregnancy. Relative expression of endometrial PGHS-2 mRNA was higher in the pregnant cow than on comparable days of the estrous cycle, i.e., days 14, 18 and 21. Expression of PGES mRNA was high at estrus, low during the luteal phase, and decreased during the peri-implantation period. Whereas, PGFS mRNA was expressed highly on days 7 and 18 of the estrous cycle and comparable days of pregnancy, but was down-regulated later with advancing pregnancy towards day 50. The presence of respective protein of oxytocin receptor, PGHS-2, PGES, and PGFS was detected in the uterine luminal and glandular epithelium that paralleled the respective mRNA. It is concluded that increased expression of PGES at estrus may be involved in ovulation and stimulation of increased blood flow
to the uterus. The increased PGHS-2 and decreased expression of PGFS during the peri-implantation period may enhance maintenance of pregnancy. The corpus luteum remained a source of oxytocin through day 22 of pregnancy when the uterine oxytocin receptors were low. Conversely depletion of luteal oxytocin and its message declined as uterine oxytocin receptors increased.
Impacts
Identification of the factors that influence the prostanoid-generating system as well as the time of their action might be relevant for early embryonic loss and could thus provide valuable information on the underlying causes; perhaps also provide an opportunity to mitigate this system by appropriate treatments
Publications
- Nichols N. 2004. Oxytocin, Relaxin-Like Factor, and the Search for Bovine Relaxin During the Peri-Implantation Period of Pregnancy. Thesis. University of Florida.
- Binta H. 2004. Expression of Genes Involved in Prostanoid Synthesis in the Endometrium and Chorioallantois of the Cow. Dissertation. University of Florida.
- Binta H., A-R. Fuchs, M. Drost, W.W. Thatcher, N. Denslow, N. Nichols, S-M. Chang, E. Johnson, M.J. Fields. 2003. Expression of Oxytocin Receptor, Prostaglandin (PG) H Synthase (S)-2, PGE-S and PGF-S in Endometrium During the Estrous Cycle and Peri-Implantation in Cattle. Biol. Reprod. 62(Suppl 1):204.
- Nichols N., H.P. Binta, P.A. Fields, M. Drost, M. Campbell-Thompson, R. Ivell, S-M. Chang, E. Johnson, M.J. Fields. 2003. Expression and Localization of Oxytocin and Relaxin-Like Factor During the Peri-Implantation Period of Bovine Pregnancy. Biol. Reprod. 62(Suppl 1):542.
- Fields, M.J., M. Shemesh. 2003. Expression and Functionality of Uterine LH Receptors During the Estrous Cycle of Domestic Animals. Biol. Reprod. 62(Suppl 1):98.
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Progress 10/01/01 to 10/01/02
Outputs
We have shown that cows on Days 22, 25, 30, and 40 of pregnancy respond to oxytocin by secretion of prostaglandin F2 alpha, which is very surprising since the effects of interferon-tau secreted by the conceptus were believed to prevail until around Day 30. One of our initial objectives was to determine the tissue target for oxytocin action at this stage of pregnancy by ascertaining which tissue oxytocin receptors and the oxytocin induced expression of cyclooxygenase-2 reside. The oxytocin and cyclooxygenase-2 mRNA was measured by Real Time PCR in the caruncular and intercarcuncular endometrium and when available chorioallantois in cows days 7, 14, 18, 22, 30, 40, and 50 of pregnancy and days 0, 3, 7, 14, 18, and 21 of the estrous cycle. As expected oxytocin receptor mRNA was high in cows at proestrus and estrus and low in pregnant cows at comparable times. Late implantation the relative abundance of oxytocin receptor mRNA increased starting day 30 paralleling the
increased responsiveness to oxytocin. There was no difference in the caruncular versus intercarcuncular expression of oxytocin mRNA. Earlier we had shown that oxytocin induces expression of cyclooxygenase-2 mRNA. However, in this study when oxytocin mRNA was low, cyclooxygenase-2 was highly expressed starting on day 14 of pregnancy. Relative expression of endometrial cyclooxygenase-2 mRNA was higher in the pregnant cow than on comparable days of the estrous cycle, i.e., days 14, 18, and 21. In addition, the conceptus membranes from day 30 to 50 of pregnancy had relative cyclooxygenase-2 mRNA that were four fold higher than detected in the endometrium. The ligand inducing the elevated cyclooxygenase-2 mRNA during early pregnancy may not be oxytocin since there is low oxytocin receptor mRNA during this time. However, the oxytocin/prostaglandin system is functional during the peri-implantation period of pregnancy. Additional hypothesis is that between days 20 and 30 of pregnancy a source
of oxytocin to drive the secretion of prostaglandins is the corpus luteum. Luteal oxytocin mRNA measured by Northern blot was present at least through day 22 of pregnancy and day 18 of the cycle. No detection of luteal oxytocin mRNA was observed on estrus or days 30, 40 and 50 of pregnancy. Luteal relaxin-like factor mRNA measured by Northern blot was observed on all days studied except day 3 of the estrous cycle. The signal increased with advancing pregnancy. Role of the relaxin-like factor in the peri-implantation cow is yet to be determined, i.e., is the relaxin-like factor the native relaxin of the cow or has the cow lost it to a pseudogene as has been shown in the ewe. Several attempts this past year to PCR the relaxin gene/pseudogene in the cow has not been successful.
Impacts
Identification of the factors that influence the prostanoid-generating system as well as the time of their action might be relevant for early embryonic loss and could thus provide valuable information on the underlying causes; perhaps also provide an opportunity to mitigate this system by appropriate treatments.
Publications
- Fuchs, A. R., Graddy, L. G., Kowalski, A. A., Fields, M. J. 2002. Oxytocin induces PGE2 release from bovine cervical mucosa in vivo. Prostaglandin. 70:119-129.
- Shemesh, M., Mizrahci, D., Gurevich, M., Shore, L.S., Reed, J. Chang, S-M. T., Thatcher, W.W., Fields, M.J. 2002. Expression of functional luteinizing hormone (LH) receptor and its messenger ribonucleic acid in bovine endometrium: LH augmentation of cAMP and inositol phosphate in vitro and human chorionic gonadotropin (hCG) augmentation of peripheral prostaglandin in vivo. Reprod. Biol. 1:13-32.
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