Comparative Veterinary Reproduction and Obstetrics, Lecture 4
Patrick J. Hemming DVM
Some protocols used for estrus synchronization
Progestogen therapy
Progesterone or progestin therapy was originally used for estrus and ovulation suppression and birth control in several species. Progesterone and progestins are still used extensively for these purposes. With regards to estrus synchronization, progestins will suppress estrus until it is desired for the animals to be bred. Progestins have the advantage of inducing estrus cyclicity in pre-pubertal heifers and post-partum cows. These methods tend to be the more complex, but frequently they will result in higher estrus response rates and tighter synchrony than simpler methods based primarily on prostaglandins. All efficient progestogen protocols include the use of other hormone pharmaceuticals, such as prostaglandin F2 alpha, to achieve their goal. Some examples of progestin protocols follow:
Treatment of heifers with Melengestrol acetate (MGA) + Prostaglandin F2 alpha
1. MGA .5mg per head, per day, is added to feed for 14 days. This suppresses estrus and ovulation. It is essential that all animals eat approximately the full dose daily for this protocol to be effective.
2. There may be a transient increase in LH secretion in animals without a CL release, this may support persistent follicular growth an prevent atresia.
3. The follicular waves may be prolonged and the dominant follicle may persist for a longer than normal time. The persistent dominant follicles oocyte is less likely to be fertilized and develop normally, adversely affecting fertility, if the animal is bred at the 1st heat after MGA withdrawal. Cattle should never be bred on the first post MGA heat.
4. Endometrial glandular development occurs during therapy.
5. Removal of MGA from feed after 14 days
6. Estrus occurs at 3 to 7 days after last MGA feeding
7. The heifers are NOT bred on this heat due to oocyte age, poor oocyte quality and low fertility of the MGA heat.
8. Poor fertility if overcome by breeding on the second heat after MGA treatment
9. PGF2 alpha is administered at 17 to 19 days after the last MGA feeding, (Approx. 10 to 14 days after the MGA estrus)
10. The animals are bred as usual after observation of estrus
11. Pregnancy rates are about 65% after breeding at the PGF2 alpha induced heat.
12. This program works best with observation of heifers for heat and breeding each heifer 12 to 24 hours after she is observed in estrus.
Treatment of cattle, sheep, goats and deer with progestogen impregnated sponges or progesterone releasing intravaginal devices such as the CIDR-B device.
1. Natural or synthetic progesterone impregnated sponge containing 30 to 60 mg progesterone analog or the CIDR device containing 1.38 or 1.9 gram of natural progesterone is placed in the vagina
2. Hormonal therapy is used to initiate a new wave of follicular growth. Inducing a new dominant follicle is advantageous to initiating tight synchrony of the induced estrus.
3. An estradiol capsule can be attached to the intravaginal device or an injection of estradiol can be administered at the time of vaginal insert placement. Estradiol will cause a rapid decrease in FSH and LH, causing follicular atresia of the dominant and all subordinate follicles. Once the dominant follicle is removed a new follicular wave will begin in 3 to 4 days
4. In the United States, were estradiol is not approved for use, GnRH is used to initiate a new follicular cycle. GnRH will cause ovulation and / or luteinization of dominant LH responsive follicles. Removal of the dominant follicle and elimination of inhibin hormone production from the dominant follicle will allow a new follicular growth to begin.
5. An FSH rebound will occur after removal of the dominant follicle and its inhibin production. A new follicular wave will begin 2 to 3 days following a GnRH injection in most cattle
6. Administration of a prostaglandin 12 to 24 hours prior to vaginal insert removal (or at the time of insert removal) assures regression of the corpus luteum
7. Removal of device at 7 days if estrogen or GnRH is given at device insertion, (cattle)
8. Removal of device at 8 days (cattle), 14 days (ewes), or 16 days (does) if only progesterone is used
9. In sheep and goats eCG (PMSG) or eCG / hCG (PG600) can be given to induce follicular development and ovulation; this improves fertility and contributes to twinning
10. Bull, buck or ram exposure may also increase fertility
11. Estrus occurs at 36 to 72 hours after implant removal, depending on supplemental treatments
12. Pregnancy rate vary depending on species and season
Treatment of mares with the oral progestogen Altrenogest (Regu-Mate)
1. Label indication is suppression of estrus with a predictable occurrence of estrus following drug withdrawal,
2. Used in transitional mares (mares with good follicular activity - follicles of 25mm or greater), mares with anovulation, prolonged estrus, or weak estrus
3. Can be used to suppress estrus prior to scheduled breeding
4. If breeding during January and February, mares should have been maintained under lights, usually initiated in November
5. Treatment continues for 15 days
6. Estrus behavior will usually begin in 48 to 72 hours after the last dose of progestogen
7. Pregnancy rate vary depending on season, typically very good
GnRH therapy (Cystorelin, Fertagyl, Factrel)
GnRH will cause ovulation or luteinization of dominant or large follicles with LH receptors. It accomplishes this by causing the release of LH from the anterior pituitary gland. GnRH is not used by itself to synchronize estrus, but rather it is used as an adjunct to estrus synchronization. The two primary reasons for using GnRH in an estrus synchronization program are to synchronize follicular development or induce ovulation.
When GnRH is used to synchronize follicular development, it is given 7 days prior to the time of progestogen removal and/or prostaglandin injection. At this time GnRH causes ovulation or luteinization of large follicles. This will causes a new group of follicles to begin development (a new follicular wave) at approximately 2 days after GnRH injection. If large follicles are not present or are atretic, small follicle will continue to grow and be ready to ovulate at the scheduled heat. The goal is to have as many animals as possible with a new dominant follicle that is ready to ovulate at the time of progestogen removal and/or prostaglandin injection. In other words GnRH is not used to synchronize estrus, but it is used to synchronize follicle development, hopefully causing more synchronous ovulation.
GnRH can also be used to induce of ovulation at the time a heifer, cow, or mare is in heat or due to be in heat, further increasing the chance for synchronizing ovulation with insemination.
There are several prostaglandin/GnRH based estrus synchronization programs for cattle, Select-Synch, Ov-Synch, Co-Synch, Modified Co-Synch, CIDR-Synch, and a few others.
Some basic protocols for these are:
1. Day 0: inject 100mcg (2ml) GnRH, inducing a LH surge and luteinization of the dominant follicle(s) and atresia of the subordinate follicles. Progesterone levels rise.
2. A new follicular wave is initiated in about 2 days.
3. Day 7: inject prostaglandin; luteolysis of the original CL and the luteinized follicles allow estrus to occur
4. There are then 4 options
A) Breed 12 to 18 hours after observed heat, estrus usually occurs at about 48 hours after prostaglandin injection (referred to as Select-Synch).
B) Inject a second dose of GnRH 48 hours after the prostaglandin injection and breed 18 hours after the second GnRH injection, no heat detection required (referred to as Ov-Synch).
C) Inject a second dose of GnRH at 60 hours (timing varies by practitioner) after the prostaglandin injection and breed at the same time as the second GnRH injection, no heat detection required (referred to as Co-Synch). (Not recommended by A.R.T., as the pregnancy rate is lower than the other protocols)
D) Breed cows 12 to 18 hours after observed heat, then at 60 hours after the prostaglandin injection and inject all cows not observe in heat with GnRH and breed at the same time (referred to as modified Co-Synch)
5. GnRH / Prostaglandin synchronization methods do have the potential to initiate postpartum cyclicity in cows if there is adequate follicular activity and a dominant or large follicle.
6. GnRH / Prostaglandin synchronization methods are not a good choice in peri-pubertal heifers unless a progestogen is added to the protocol, such as using a CIDR-B insert at the time of the first GnRH injection and pulling the CIDR at the time of the prostaglandin injection (referred to as CIDR-Synch). Other methods that utilize progesterone compounds and do not rely on GnRH to synchronize follicular waves, probably work better in heifers.
Induction of ovulation in animals with a mature dominant follicle is possible with GnRH.
1. In cattle an injectable GnRH is effective in inducing a LH surge that should cause ovulation if the GnRH is administered at the time of expected estrus. As long as progesterone levels are low (baseline < 1 ng/ml), and a healthy dominant follicle is present, pregnancy may be achievable even in animals that are not displaying estrus. GnRH should be given at the very start of estrus for maximum effectiveness in animals displaying heat.
2. In mares injectable GnRH has not been effective in inducing ovulation of large (>=30mm) follicles. Therefore an implant has been developed, Ovuplantthat releases GnRH continually over several days. The resulting increase in circulating LH will cause ovulation in 86% of implanted mares within 48 hours. Ovuplant contains 2.1 mg of GnRH analog that is released over 4 to 5 days.
All of the above methods of estrus induction can be used to synchronize the heat period of large numbers of animals for efficient artificial insemination.
Prostaglandin Therapy, Short Cycling and estrus synchronization with prostaglandin F2 alpha
The normal estrus cycle in most farm animals is 18 to 21 days. By causing early CL regression with prostaglandin F2 alpha (PGF), the length of the estrus cycle can be shortened in mares, cows, ewes and does. There are a few important points to consider when using prostaglandin preparations: The CL of the mare and ruminants is refractory to PGF2 alpha treatment for about 5 days after ovulation. Animals that had a heat only 5 days ago will probably not respond. Also, since prostaglandins exert their effect on a mature and functional CL they are only effective in a cycling animal. Animals that are pre-pubertal, and those in post-partum anestrus or seasonal anestrus will not respond. Due to these factors the response rate to a single PGF injection will vary from 0% to 75% maximum. It is rare, except in a small group, to ever get a response rate that exceeds 75%. Prostaglandins are not very effective in swine since their CL is only responsive to PGF2 alpha very late in the estrus cycle when natural PGF2 alpha is released from the uterus anyway.
After administration of a prostaglandin preparation, CL regression is complete within 48 hours for a responsive CL. If a dominant (estrogen active) follicle, of ovulatory size, is present, the animal will exhibit estrus, typically within 48 to 72 hours after injection. Earlier estrus occurs if CL regression was already occurring, in spite of the PGF injection (0 to 36 hours.). A longer interval to estrus will occur if there is not a dominant follicle present (84 hours to 9 days). If the CL was refractory to PGF treatment, estrus will occur at the normal interval for that animal (10 to 21 days after PGF injection).
Prostaglandin F2 alpha analogs are used extensively as an inexpensive and effective method to synchronize estrus for AI in cycling ruminants. Since simple PGF injection protocols (utilizing only PGF) do nothing to synchronize the follicular cycle, these are not good protocols for timed AI and heat detection is usually employed. Several protocols have been used in cattle including:
1. Single injection methods: The following methods will reduce material and semen costs but requires labor for heat detection.
A) Inject a single dose in all cattle, observe for estrus, and breed 12 hours after standing heat is observed. This method is used when minimization of materials and semen costs is necessary. All animals not bred by day 10 or 11 post-injection can be injected with a second dose of PGF and heat detection and breeding can continue.
B) The best (my opinion) single injection method includes observing the cattle for estrus for 5 or 6 days prior to PGF injection. Breed all cattle at 12 hours after seen in heat. On day 6 or 7 inject PGF, only into cattle not yet observed in heat. Continue to breed at 12 hours after observed heat. This protocol effectively deals with refractory CLs, since all animals that are to be injected with PGF would have had a heat over 6 days prior to injection.
C) Timed AI at 75 to 80 hours post injection, of all animals not yet observed in heat. This technique has been utilized with a single injection protocol. Heat detect the cattle and breed all cattle at 12 hours after seen in heat. At 75 to 80 hours all animals not observed in heat are inseminated. Perhaps a few animals will become pregnant to the Timed AI, but many animals that do not show estrus by 80 hours may be anestrous or not yet in heat and too early to breed. A better way to deal with animals that have not responded is to treat them with a CIDR-B protocol. A Fixed Time Artificial Insemination (FTAI) can be used. A FTAI is defined as a single breeding period scheduled in advance for all of the synchronized cattle. This variation of the protocol may be used for labor saving reasons. Inject a single dose of PGF in all cattle and breed at a fixed time of 75 to 80 hours post injection with minimal or no estrus detection. This method is used when semen cost is of no concern, minimization of labor is necessary and conception rate to the timed AI is not critical. Only expect about 65 to 70% of the animals to actually respond and exhibit estrus. Timing of the AI will be poor for many of the animals bred. Overall conception rates of 35 to 50% are expected. Clean-up bulls should be used to breed the balance of the cattle.
2. Double injection methods:
A) Inject first dose of prostaglandin. Expect about 65% to 70% of the animals to respond and exhibit estrus in 2 to 5 days. The animals are not bred at this time.
B) Inject a second dose of prostaglandin 11 days after the first dose. All animals that had a heat after the first dose should respond and exhibit estrus after the second dose. Animals that did not respond after the first dose due to refractory CLs (too soon after the previous estrus) should also respond to the second dose of prostaglandin and exhibit estrus. The only animals that will not respond to the second dose are anestrous. Heat detect after the second dose of PGF and breed 12 hours after standing heat is observed. Timed AI at 75 to 80 hours post injection (no estrus detection) can be employed if desired.
C) If desired, animals seen in heat after the first dose can be inseminated. Obviously these animals would be excluded from the second injection of prostaglandin.
3. Estrus synchronization using GnRH in combination with Prostaglandin, Ov-Synch , Select-Synch, etc. are discussed above.
4. In mares PGF is used to induce a fertile heat but it will not synchronize ovulation due to the long and variable length of estrus.
5. Prostaglandins have some other uses:
A) Early abortion of mis-mated farm animals. In cattle and horses the PGF treatment should be delayed for 6 or 7 days to account for the CLs refractory period. In swine treatment should be delayed for 14 post-ovulation. Usually a single luteolytic dose of PGF is all that is required. Sometimes heifer calves are treated at weaning time with PGF to assure that there are no pregnancies in early maturing individuals.
B) Early abortion of mis-mated bitches (although with much greater difficulty than in farm animals). Requires twice a day (BID) treatment for at least 4 days. There are many side effects.
C) Synchronization of farrowing in swine
D) Treatment of uterine infections associated with a functional CL
E) Treatment of canine pyometra
Gonadotropin therapy
Although gonadotropins (FSH, LH, eCG or hCG) are not used extensively in estrus synchronization, there are some gonadotropin applications, related to estrus control, that are worth mentioning.
PG600 from Intervet is a commercial preparation of eCG (also called PMSG) in combination with hCG. PG600 is used in postpartum sows, pubertal gilts, and anestrous sheep. The eCG part of this product has very potent FSH activity and will stimulate follicular development if given during diestrus or the proestrus parts of the estrus cycle. Follicular development is also stimulated by eCG during the transition from anestrus to proestrus in non-cycling animals. The hCG part of this product has LH activity and promotes final maturation of large graafian follicles. Using this product may hasten or stimulate an estrus.
PG600 is used in pre-pubertal and pubertal gilts at 6 to 8 months of age to initiate an estrus. This will shorten the time to puberty in later maturing gilts and can be used to synchronize gilts for AI. Response to treatment is usually over 65%. PG600 is given to sows at the time of weaning pigs to induce estrus, reduce the number of anestrus sows post-weaning, and shorten the time interval from weaning to first estrus. Most gilts and sows treated with PG600 will be in heat within 4 days.
During progesterone-based synchronization procedures in small ruminants PG600, given at the time of progesterone removal and prostaglandin administration may induce twins or even triplets, which is particularly desirable in sheep.
FSH and PMSG have been used in embryo recipient cows at sub-superovulation dosage to induce 2 or 3 ovulations, thus boosting progesterone levels and theoretically pregnancy rates. There has been some success using this technique.
FSH or PMSG (not PG600) are used for superovulation in cattle, sheep and goats. PMSG has a longer duration of action (half-life of about 2 days) than FSH (half-life of 30 to 60 minutes). PMSG is effective with a single injection. FSH based superovulation requires several injections, usually twice a day for up to 5 days.
