Basic guidelines to select the right synchronization protocol for Timed Artificial Insemination in cattle

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This short review will cover basic aspects to be taken into account when selecting synchronization programs for beef or dairy herds in countries where estrogens are not allowed. This manuscript obviously does not intend to cover the almost infinite variety of Timed Artifical Insemination protocols used in cattle, but rather will indicate key successful hormonal treatments to synchronize ovulation that may be adapted according to unique management situations with the assistance of the herd’s veterinarian.

Alexandre Souza, DVM PhDUniversity of CaliforniaCooperative Extension Service

Alessio Valenza, DVM PhDInternational Technical Manager for Ruminants Corporate MarketingCeva Sante Animale

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The use of synchronization programs for Timed Artificial Insemination (TAI) allows farmers to breed their cattle without relying on estrus detection. In some cases due to prolonged postpartum anoestrus or in herds with high milk production levels, it can be very difficult to detect cows in heat due to different physiological limitations such as lack of Corpus Luteum for 7 to 10 days in the ovaries due to negative energy balance and/or calf presence (in case of beef cows postpartum) or due to high rates of steroid metabolism by the liver (for high producing dairy cows) that may suppress oestrus behavior. Therefore, strategic use of adequate synchronization programs can represent an important tool that helps producers to improve service rates and to treat postpartum anoestrus.

However, relying 100% on the TAI programs to breed cows may not be the best option. In most cases, the ideal approach is to combine good heat detection systems with an effective synchronization program, particularly for second and following postpartum breedings. The goal is to breed nearly 100% of the cows for the first time until 100 days in milk (DIM) for mature cows or within 30 days after reproductive age is achieved in case of virgin heifers. A second objective of a good reproductive schedule must be to keep the average interval between breedings less than 35 days in high producing dairy cows and roughly less than 30 days in heifers (dairy and beef) and mature beef cows.

1. Important aspects to be considered before choosing the synchronization programUnderstanding the concept of “Timed Artificial Insemination”

Reference: Souza AH et al, 2013

Thus, some important features of synchronization programs are associated with enhanced heat detection performance and to treatment of anovular/ cows, bringing them back to cyclicity. The number of cows that get pregnant after the

Percentage of AI performed with synch protocols

Maximum service rate = estrus detection + synchronization protocols

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Hopefully you’re convinced of the importance of computer programs that will help with the daily routine when using synchronization and TAI. Now, which one should I buy in case I don’t have any software? Well, I’d pick the one that can handle what I want to do in terms of management routines. It may not be the cheapest one, but as you know, good things are normally not cheap! Before purchasing any software, recheck whether appropriate support is available while setting up the program on your herd - this is probably an important aspect to be considered when choosing your herd management software.

Herds willing to handle larger volumes of breedings through a sequence of hormonal shots, which in some cases can be quite complex, will need to adopt the use of an herd management software. Unfortunately, more than often we come across large herds with sometimes over 2,000 cows in milk that still try to track shots and breeding routines manually. Tracking cows and hormones is unbearable overtime, and decision needs to be made towards start using a consistent herd management software…no matter how unfamiliar you are with computers, get adapted to it or you may be out of business in a few years. Most computer programs are not very expensive at all, and they will save enormous amounts of time and headache to producers – stop the excuses, stop the hand notes, get started now!

The importance of good record keeping systems

Tracking reproductive performance

use of synchronization programs is normally similar or higher to fertility after natural breedings ranging around 30% to 40% in dairy cows and 45 to 70% in beef cows and heifers (dairy or beef). The decision on “how much” hormones for synchronization needs to be used in the herd depends largely on the producer’s goals in terms of reproduction performance, cattle breed, percentage of anovular/anoestrus cows, price of hormones, labor costs for heat detection (and pregnancy diagnosis), adoption of software to organize the reports of hormones that need to be used, level of milk production, implementation of adequate facilities to handle animals, and most importantly, trained personnel.

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Briefly, frequency of pregnancy exams are key part of finding non-pregnant or “open” cows when relying on TAI protocols. Thus, the correct frequency and type of pregnancy diagnosis approach needs to be well thought during the process of selecting the right synchronization program. Obviously, larger herds will require more frequent and most likely weekly pregnancy exams. Interestingly, in recent years, blood pregnancy tests based on PSPB or PAGs* detection allow smaller producers to re-assign open cows to

Another important point when buying computer programs for herd management is to ask about the features in the software that will allow you and your consultant to track reproduction with modern statistical analysis and ease-to-read graphic tools. For example, try to find out if you can measure the percentage of cows that are bred within 100 DIM, or if you can see the calving distribution from last year’s calving season. With this type of tool you and your consulting team will be able to evaluate current status of the herd and make informed management decisions based on accurate numbers. For instance, if less than 90% of the cows are bred before 100 DIM, an action plan needs to be designed in order to fix the problem, and herd management programs need to provide this type of tool to help farmers and consultants to decide to invest and/or modify the current way of managing reproduction.

How about herds that may be too small to jump into a “wild adventure” of buying a software for herd management? This is a rather common picture and the solution is probably to make sure that producers are using well designed paper-cow cards for record keeping. For instance, in these cases heat detection performance can easily be calculated by using the “Woods” formula. The Woods’s calculation simply uses the assumed cow’s cycle length divided by the average interval between breedings to estimate heat detection performance.

Type and frequency of pregnancy diagnosis according to herd size

Example of “Woods” calculation of heat detection rate of a herd:Herd with two cows (n=2)

• Interval between first and second AI for cow1 (C11-2AI)= 46 days• Interval between first and second AI for cow2 (C21-2AI)= 25 days• Standard cow´s cycle length (CCL)= 21 days

Calculations:Average interval between breedings (IBB) = (C11-2AI) + (C21-2AI) / n = (46 + 25) / 2 = 35.5 days “Woods” calculation of heat detection rate = CCL / IBB

= 21 / 35.5 = 0.59 x 100 = 59% average heat detection rate

*PSPB: pregnancy-specific protein B / PAGs: pregnancy-associated glycoproteins.

Having a solid vision on the strengths and potential weaknesses of the herd is essential for genetic advance. Thus, get in touch with your semen provider to come up with a strong genetic plan for your herd based on your market situation. This plan needs to include matings that will reduce inbreeding to less than 6%, use calving-ease sires for virgin heifers, etc while producing animals that are suitable to your management constraints. As far as semen inventory, you may need to plan ahead before start using TAI protocols! This is particularly truth for beef herds – make sure the semen provider is ready to keep up with the increased amount of breedings (with sexed or non-sexed semen) after adopting synchronization programs.

Sire selection – genetic plan for the herd

100 or less 100 to 500 500 to 100

Number of lactating cows in the herd

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synchronization programs in a weekly basis. In addition to herd size, the importance of frequent pregnancy diagnosis increases a lot as the herd’s heat detection efficiency decreases. Therefore, herds using more efficient heat detection strategies such as daily professional tail chalk service, well managed utilization of automated heat detection devices, and/or use of well managed clean up bulls (particularly in beef herds) may in some situations afford having less frequent pregnancy diagnosis. Thus, here is a suggestion of MINIMUM frequency of pregnancy exams according to herd size for dairy herds:

Minimum number of required veterinary visits according to herd size.(This frequency may need to be increased in herds with poor estrus detection).

Photo’s courtesy of J. Ruiz del Olmo

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Before jumping in to the TAI programs, here is a short list of main hormones, its main purpose when used in TAI protocols, and designated short-name that will be used interchangeably throughout this manuscript:

Standard Operating Procedure or S.O.P. is a simple document that explains the weekly routine for any cow-related activities in the herd. Thus, the S.O.P for reproduction can be a written file or just a simple chart that summarizes by day of the week what happens in terms of treatments to postpartum cows, detailed heat detection routine, actions that need to be performed when cows are detected open, people in charge of each task, etc. Sticking to the S.O.P can simplify the overall reproduction management.

2. Reproductive routine: the concept of “Standard Operating Procedure”– S.O.P.

Progesterone Releasing Intravaginal Device

Prostaglandin F2α

Gonadotropin Releasing Hormone

Equine Gonadotropin

• Priming of progesterone to induce cyclicity in anovular/anoestrus cows• Avoid premature estrus behavior during the synchronization protocols

• Induce CL regression in mature CLs (older than 5d)

• Induce LH surge and ovulation of dominant follicles

• Hormone with FSH/LH-like function used to enhance follicle development at final stages during synchronization protocols• Proven to improve ovulation rates and fertility when used in anovular/anoestrus cows

PRID, P4, P4 device

PGF, PGF2α

GnRH

eCG (formerly known as PMSG)

Hormone/Device Short-name, will be used interchangeably throughout the manuscript

Main function when used in synchronization programs

3. TAI protocols

This short P4-based protocol has been developed somewhat recently and most reports are showing very promising results. It seems to work well for mature cows and heifers (beef or dairy). The main advantage of this protocol is its simplicity and length – only 3 managements within only 7 days! In addition, this protocol does not use any type of estrogen on day 0 (day of progesterone device insertion), so it can be initiated in pregnant cows with no risk of CL regression, which may save a couple of days when trying to Re-synchronize cows or heifers. This protocol may include a GnRH treatment on Day 0 together with P4 device insertion, and a second PGF treatment given 24h after the first PGF at device removal, mandatory when using GnRH at beginning of the 5d-protocol, otherwise CL will not regress in a good portion of cows (particularly dairy cows) and fertility following TAI will definitely be compromised. Last, more studies are needed to confirm whether eCG can be successfully used at device removal without undesirable increase in double ovulation rates. Previous variations of this protocol included longer periods of P4 device insert also showing fairly good success rates.

5d-Protocol

5d-Protocol: recommended for dairy or beef cows and heifers

PGF

D08 AM

D58 AM

GnRH + AI

D74PM

PRID for 5 days

*For best results minimum voluntary waiting period of 60 or more days to AI is strongly recommended HD=strategic heat detection to catch non-pregnant cows returning in heat after 1st postpartum AI.

Pregnancy Diagnosis at 28 days after AI (at the time of P4 insertion) or at 33 days after AI (at the time of P4 removal).

Weekly schedule:

SU MO TU WE TH

HD+AIHD+AI

HD+AIHD+AI

HD+AIHD+AI

8AM: Insert PRID8AM: Remove PRID + PGF

HD+AI

SAFR

HD+AI HD+AI

8AM: Remove PRID + PGF

4PM: 56h after PRID removal - GnRH+AI*

8AM: Insert PRID4PM: 56h after PRID removal - GnRH +AI

HD+AI

56h to 72h

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Pre-

sync

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ch®

1st AI

GG

PG fo

r 2nd

+ AIs

This protocol has been developed more than one decade ago and it is probably the most common TAI protocol used in large dairy herds world-wide. The intervals between PGF and Ovsynch® of 14 and 10 days respectively seems to produce somewhat improved results compared to the traditional 14-14d intervals between PGF and Ovsynch®. Some disadvantages of this protocol are mainly related to its inability to treat anovular/anestrus cows and its length and complexity, making it very difficult to use in herds without advanced Standard Operating Procedures (S.O.P.) and computer programs. Therefore, Pre-synch strategies relying exclusively on PGF treatments should be avoided in herds with postpartum cyclicity issues. However, it can be a good option in confined dairy herds that are not undergoing problems related to anovular/anestrus cows. Heat detection and AI after the 2nd PGF are normally encouraged, but as a rule of thumb only if conception results, out of these estrus-breedings, are >30%. The weekly routine presented below by days of the week shows the sequence of treatment for a Pre-synch Ovsynch® for 1st postpartum AI and the use of the GGPG* protocol (good option when Re-synchronizing open cows for 2nd and following AIs) for Re-synchronizing lactating dairy cows.

*For best results minimum voluntary waiting period of 60 days to AI in high producing cows is strongly recommended HD=strategic heat detection to catch non-pregnant cows returning in heat after 1st postpartum AI.Pregnancy Diagnosis at 32 days after AI (at the time of the second GnRH).

SU MO TU WE TH

HD+AI HD+AI

8AM: PGF

HD+AI

HD+AI

8AM: GnRHHD+AI

SAFR

HD+AI

HD+AI

HD+AI

HD+AI*

4PM: GnRH

8AM: PGF8AM: GnRH to open cows

8AM: GnRHHD+AI

4PM: GnRH

HD+AI

8AM: AI

HD+AI

8AM: AI

HD+AI

8AM: PGF

8AM: PGF

HD+AI

{{

Heat detection and AI in cows showing estrus behavior{TAIGnRHPGF2α PGF2α PGF2αGnRH

16h56h14d 10d 7d

D33 4PM

D34 8AM

D08AM

D14 8AM

D24 8AM

D31 8AM

Adding progesterone devices in Ovsynch®-like programs between the 1st GnRH of Ovsynch® until PGF injection increases conception rate of 5 to 8% in both cycling and anovular dairy cows

*GGPG (GnRH before Ovsynch®)

Weekly schedule:

Pre-synch Ovsynch® (2PgF + Ovsynch®)

HD+AI

This is another GnRH-PGF based protocol but with the advantage of having a GnRH treatment 7 days before the initiation of the Ovsynch® protocol. This GnRH treatment should help with cows presenting anovular-large follicle condition postpartum. Can also be a good option for high producing dairy cows. Notice that some of these protocols are quite complex and follow up of treatments need to be through a computer software to avoid mistakes. If computer programs are lacking in the herd you may want to pick a more simple approach to synchronize cows. The weekly schedule below shows the sequence of treatments for the G-6-G protocol that has been slightly modified to facilitate the weekly routine, followed by the GGPG protocol for 2nd and following breedings.

G-6-G protocol (modified to facilitate weekly routine)

Modified G-6-G protocol: recommended for the 1st AI postpartum in confined high producing dairy cows

*For best results minimum voluntary waiting period of 70 or more days to AI in high producing cows is strongly recommended HD=strategic heat detection to catch non-pregnant cows returning in heat after 1st postpartum AI.

Pregnancy Diagnosis at 32 days after AI (at the time of the second GnRH).

D08AM

56h 3d 7d 7d 16h

D3 8AM

D10 8AM

D17 8AM

D19 4 PM

D20 8AM

PGF GnRH AIGnRHPGFGnRH

Adding progesterone devices in Ovsynch®-like programs between the 1st GnRH of Ovsynch® until PGF injection increases conception rate of 5 to 8% in both cycling and anovular dairy cows

SU MO TU WE TH

8AM: GnRH

SAFR

HD+AIHD+AI

4PM: GnRH

8AM: PGF8AM: GnRH to open cows

8AM: GnRHHD+AI

4PM: GnRH

HD+AI

8AM: AI

8AM: AI

HD+AI

8AM: PGF

HD+AI

{

Mod

ified

G6G

1st AI

GG

PG fo

r 2nd

+ AI

s {

8AM: PGF8AM: GnRH

HD+AIHD+AI

TECHNICAL PUBLICATIONS

*For best results minimum voluntary waiting period of 70 or more days to AI in high producing cows is strongly recommended HD=strategic heat detection to catch non-pregnant cows returning in heat after 1st postpartum AI.

Pregnancy Diagnosis at 32 days after AI (at the time of the second GnRH).

This GnRH-PGF protocol has been designed to synchronize 1st postpartum AI in lactating cows in very high producing dairy herds. It is a fairly long and complex protocol. Thus, it makes the use of computer programs to track cows and treatments mandatory. It is very effective to treat anovular high producing dairy cows that usually present follicles with ovulatory capacity or with diameter > 10mm, but are not cycling (condition now known as “large follicle anovular syndrome”). It is not effective in cows undergoing the standard “small follicle-postpartum anoestrus”, and is not recommended for virgin heifers either. Main advantage is related to high circulating P4 after D17 or at the time of the first GnRH of the “breeding Ovsynch®” (see picture below), which in turn tends to produce oocytes with very high quality even in high producing dairy cows. Seem to reduce double-ovulation rates in lactating cows. Because of its length is normally not recommended during Re-synchronization programs for 2nd and later AIs.

Double-Ovsynch® protocol

Double-Ovsynch® protocol: recommended for the 1st AI postpartum in confined-high producing dairy cows

GnRH PGF AIPGFGnRHGnRH

D08AM

D78AM

D108AM

D178AM

D248AM

D264PM

GnRH

D278AM

7d 3d 7d 7d 16h56h

Pre-synch Ovsynch® Breeding Ovsynch®

8AM: GnRH8AM: GnRH

Doub

le-O

vsyn

ch®

1st A

ISU MO TU WE TH SAFR8AM: GnRH {

HD+AIHD+AI

8AM: PGF8AM: GnRH to open cows

8AM: GnRHHD+AI

4PM: GnRH

HD+AI

8AM: AI

HD+AI HD+AI

GG

PG fo

r 2nd

+ AI

s{

8AM: PGF

HD+AIHD+AI

8AM: PGF 4PM: GnRH 8AM: AI

Because dairy herds normally have relatively lower conception results after 1st AI (~30 to 40%), wide adoption of artificial insemination, and most importantly poor estrus detection efficiency, most herds need to have a precise plan for 2nd and further breedings in their S.O.P. for reproduction management. Below are a few examples of Re-Synchronization strategies that can be successfully used in dairy herds. Other good option for Re-synchronization in dairy herds may include the 5d-protocol, or even a simple PGF injection to open cows followed by heat detection and Ovsynch® to cows not detected in heat 10 days later.

A few further considerations on Re-synchronization strategies

GGPG protocol: recommended for the 2nd+ AIs (Re-synch) postpartum in high producing dairy cows

GnRH AIPGF2αGnRH

7d

D21 after AI8AM

D288AM

D358AM

D374PM

GnRH

D388AM

7d 16h56h

Pregnancy diagnosis

5d-Protocol: recommended for dairy or beef cows and heifers

PGF

D08AM

D58AM

GnRH + AI

D74PM

PRID for 5 days56h to 72h

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4. Final thoughtsThe flow chart below summarizes main synchronization strategies that can be used in different cattle breeds and animal category.

Important:- Choose adequate Voluntary Waiting Period (VWP) due to risk of low conception if cows are inseminated too early;- If clean-up bull is necessary: complete breeding soundness exam at least 2x/year and correct bull management including adequate resting time for the bull (particularly dairy herds), correct proportion cows per bull, etc is mandatory.

Mature beef or dairy cows?Beef or dairy Heifers?

lactating Dairy cows

Beef cowsBeef heifersDairy heifers Cows housed in confined systems or pasture/mixed?

Pasture based Confinement

If herd is more than 50 heads:buy herd management software now!

If herd is more than 50 heads:buy herd management software now!

Is good heat detection system in place?Greater than 60% of estrus detection rates?

Sure?

1st AI: Pre-synch + Ovsynch® and AIs in cows showing estrus after the 2nd PgF injection Resynch: simple Ovsynch® to non-pregnant cows may work, but adding progesterone device during Ovsynch® increases conception results from 5 to 8% in most cases

1st AI: 5d protocol, Double-Ovsynch®, G-6-G protocol Resynch: must have weekly preg checks + GGPG protocolor 5d-protocol

Yes

5d protocol + eCG or calf removal if mature beef cows have low BCS or less than 100 days in milk

No

It is important to highlight that synchronization programs can unquestionably help producers to treat anovular/anoestrus cows and increase service rates or heat detection efficiency. Conception results may occasionally be improved with the use of specific protocols in relation to natural estrus breedings, particularly in high producing cows. However, in herds in which cows are inseminated but not getting pregnant very easily, producers and herd consultants need to review other important areas of management such as the transition period, Cow Comfort and nutrition requirements besides trying to solve their fertility issues solely through a fancy TAI protocol. In addition, reproductive performance can be improved through tracking key performance indicators and help producers and consultants to perform management changes based on real evidence.

Bisinotto, R. S., E. S. Ribeiro, L. T. Martins, R. S. Marsola, L. F. Greco, M. G. Favoreto, C. A. Risco, W. W. Thatcher, and J. E. P. Santos. 2010. Effect of interval between induction of ovulation and artificial insemination (AI) and supplemental progesterone for resynchronization on fertility of dairy cows subjected to a 5-d timed AI program. Journal of Dairy Science 93(12):5798-5808.

Bisinotto, R. S. and J. E. P. Santos. 2011. The use of endocrine treatments to improve pregnancy rates in cattle. Reproduction, Fertility and Development 24(1):258-266.

Bloch, A., Y. Folman, M. Kaim, Z. Roth, R. Braw-Tal, and D. Wolfenson. 2006. Endocrine Alterations Associated with Extended Time Interval Between Estrus and Ovulation in High-Yield Dairy Cows. Journal of Dairy Science 89(12):4694-4702.

Brusveen, D. J., A. P. Cunha, C. D. Silva, P. M. Cunha, R. A. Sterry, E. P. B. Silva, J. N. Guenther, and M. C. Wiltbank. 2008. Altering the Time of the Second Gonadotropin-Releasing Hormone Injection and Artificial Insemination (AI) During Ovsynch Affects Pregnancies per AI in Lactating Dairy Cows. Journal of Dairy Science 91(3):1044-1052.

Brusveen, D. J., A. H. Souza, and M. C. Wiltbank. 2009. Effects of additional prostaglandin F-2 alpha and estradiol-17 beta during Ovsynch in lactating dairy cows. Journal of Dairy Science 92(4):1412-1422.

Chebel, R. C., M. J. Al-Hassan, P. M. Fricke, J. E. P. Santos, J. R. Lima, C. A. Martel, J. S. Stevenson, R. Garcia, and R. L. Ax. 2010. Supplementation of progesterone via controlled internal drug release inserts during ovulation synchronization protocols in lactating dairy cows. Journal of Dairy Science 93(3):922-931.

Colazo, M. G., A. Dourey, R. Rajamahendran, and D. J. Ambrose. 2013. Progesterone supplementation before timed AI increased ovulation synchrony and pregnancy per AI, and supplementation after timed AI reduced pregnancy losses in lactating dairy cows. Theriogenology 79(5):833-841.

Hernández-Cerón, J., L. Zarco, and V. Lima-Tamayo. 1993. Incidence of delayed ovulation in Holstein heifers and its effects on fertility and early luteal function. Theriogenology 40(5):1073-1081.

Kaim, M., A. Bloch, D. Wolfenson, R. Braw-Tal, M. Rosenberg, H. Voet, and Y. Folman. 2003. Effects of GnRH Administered to Cows at the Onset of Estrus on Timing of Ovulation, Endocrine Responses, and Conception. Journal of Dairy Science 86(6):2012-2021.

Lima, F. S., H. Ayres, M. G. Favoreto, R. S. Bisinotto, L. F. Greco, E. S. Ribeiro, P. S. Baruselli, C. A. Risco, W. W. Thatcher, and J. E. P. Santos. 2011. Effects of gonadotropin-releasing hormone at initiation of the 5-d timed artificial insemination (AI) program and timing of induction of ovulation relative to AI on ovarian dynamics and fertility of dairy heifers. Journal of Dairy Science 94(10):4997-5004.

Moreira, F., C. Orlandi, C. A. Risco, R. Mattos, F. Lopes, and W. W. Thatcher. 2001. Effects of Presynchronization and Bovine Somatotropin on Pregnancy Rates to a Timed Artificial Insemination Protocol in Lactating Dairy Cows. Journal of Dairy Science 84(7):1646-1659.

References:

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Ask your Ceva representative for ReprodAction advice

reprodActionReproductive management in Action

Pursley, J. R., M. O. Mee, and M. C. Wiltbank. 1995. Synchronization of ovulation in dairy cows using PGF2α and GnRH. Theriogenology 44(7):915-923.

Ribeiro, E. S., R. S. Bisinotto, M. G. Favoreto, L. T. Martins, R. L. A. Cerri, F. T. Silvestre, L. F. Greco, W. W. Thatcher, and J. E. P. Santos. 2012. Fertility in dairy cows following presynchronization and administering twice the luteolytic dose of prostaglandin F2α as one or two injections in the 5-day timed artificial insemination protocol. Theriogenology 78(2):273-284.

Santos, J. E. P., C. D. Narciso, F. Rivera, W. W. Thatcher, and R. C. Chebel. 2010. Effect of reducing the period of follicle dominance in a timed artificial insemination protocol on reproduction of dairy cows. Journal of Dairy Science 93(7):2976-2988.

Sartori, R., J. M. Haughian, R. D. Shaver, G. J. M. Rosa, and M. C. Wiltbank. 2004. Comparison of Ovarian Function and Circulating Steroids in Estrous Cycles of Holstein Heifers and Lactating Cows. Journal of Dairy Science 87(4):905-920.

Souza, A. H., H. Ayres, R. M. Ferreira, and M. C. Wiltbank. 2008. A new presynchronization system (Double-Ovsynch) increases fertility at first postpartum timed AI in lactating dairy cows. Theriogenology 70(2):208-215.

Souza, A. H., A. Gumen, E. P. B. Silva, A. P. Cunha, J. N. Guenther, C. M. Peto, D. Z. Caraviello, and M. C. Wiltbank. 2007. Supplementation with estradiol-17 beta before the last gonadotropin-releasing hormone injection of the ovsynch protocol in lactating dairy cows. Journal of Dairy Science 90(10):4623-4634.

Valenza, A., J. O. Giordano, G. Lopes Jr, L. Vincenti, M. C. Amundson, and P. M. Fricke. 2012. Assessment of an accelerometer system for detection of estrus and treatment with gonadotropin-releasing hormone at the time of insemination in lactating dairy cows. Journal of Dairy Science 95(12):7115-7127.

Walker, W. L., R. L. Nebel, and M. L. McGilliard. 1996. Time of Ovulation Relative to Mounting Activity in Dairy Cattle. Journal of Dairy Science 79(9):1555-1561.

Walsh, R. B., S. J. LeBlanc, T. F. Duffield, D. F. Kelton, J. S. Walton, and K. E. Leslie. 2007. The effect of a progesterone releasing intravaginal device (PRID) on pregnancy risk to fixed-time insemination following diagnosis of non-pregnancy in dairy cows. Theriogenology 67(5):948-956.

Wiltbank, M. C., P. D. Carvalho, K. Abdulkadir, K. S. Hackbart, M. A. Meschiatti, M. R. Bastos, J. N. Guenther, A. B. Nascimento, M. M. Herlihy, M. C. Amundson, and A. H. Souza. 2011. Effect of Progesterone Concentration During Follicle Development on Subsequent Ovulation, Fertilization, and Early Embryo Development in Lactating Dairy Cows. SSR, 161 (Abstract).

Wiltbank, M. C., A. H. Souza, J. O. Giordano, A. B. Nascimento, J. L. M. Vasconcelos, M. H. C. Pereira, P. M. Fricke, R. S. Surjus, F. C. S. Zinsly, P. D. Carvalho, R. W. Bender, and R. Sartori. 2012. Positive and negative effects of progesterone during timed AI protocols in lactating dairy cattle. Page 10 in Animal Reproduction. Vol. 9.

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