D-cloprostenol and dinoprost tormetamine at the time of artificial insemination in Nelore heifers: luteal dynamics and pregnancy rate

D-cloprostenol and dinoprost tormetamine at the time of artificial insemination in Nelore heifers: luteal dynamics and pregnancy rate | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article D-cloprostenol and dinoprost tormetamine at the time of artificial insemination in Nelore heifers: luteal dynamics and pregnancy rate Betânia B. Santos, Francisco L. S. Franco, Ingrid R. N. Martins, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6508312/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The aim of this study was to evaluate the effect of combining Dinoprost tromethamine and D-cloprostenol in a fixed-time artificial insemination (FTAI) protocol on ovulation, fertility and pregnancy rate in Nelore heifers. A total of 28 Nelore heifers were evaluated. On D0, they received an intravaginal progesterone (P4) device and an intramuscular (IM) injection of 2 mg of oestradiol benzoate (BE). On D7, an application of 2mL of dinoprost tromethamine, on D9 the P4 device was removed and 0.3 mL of oestradiol cypionate (eCP) and 1.5 mL of equine chorionic gonadotrophin (eCG) were administered, on D11 all the females were inseminated and randomly divided into three groups: T1, T2 and T3. The control (T1; n = 8), in which the animals received 2 mL of saline IM; T2 (n = 11), received 2 mL of dinoprost tromethamine IM; and T3 (n = 9), received 2 mL of D-Cloprostenol IM. Follicle diameter was assessed on D0, D7, D9, D11 and corpus luteum (CL) diameter on D12. Gestation diagnosis (GD) was carried out 30 days after artificial insemination (AI). Non-parametric variables were analyzed using the Friedman test, with a significance level of 5%. The pregnancy rate was different between treatments (p < 0.05), being higher in T3 when compared to T1 (55.55%) vs. (12.5%), as well as T2 (55.55%) vs. (27.27%). There was no interaction between treatments in terms of follicle diameter at D11 and CL formation. It was concluded that the use of two doses of PGF2α (dinoprost tromethamine and D-cloprostenol) was efficient in synchronizing small follicles (< 10mm) and increasing pregnancy rates. Assisted reproduction Bovine females Corpus luteum Pregnancy Prostaglandin Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 INTRODUCTION The demand for food of animal origin with a known origin has made the market more demanding in terms of animal production and its effects (Junior et al. 2020). From 1990 to 2020, meat production in Brazil grew by 122%, with a 159% increase in productivity, and the area of pasture used decreased by 13.6%, due to investments in breeding technology and biotechnology (ABIEC, 2021). Despite advances in reproductive biotechnology, beef heifers face reproductive efficiency challenges, affecting economic viability (Baruselli et al. 2022 ). Early gestation of heifers in the first breeding season is crucial to ensure quality calves and a longer productive life (Rodrigues 2016 ), with the use of fixed-time estrous synchronization (Marques et al. 2019). Research aims to improve fixed-time artificial insemination (FTAI) protocols for heifers, with emphasis on the use of prostaglandins to optimize results (Macmillan et al. 2020 ), some of which have shown positive effects of prostaglandins in ovulation induction, although their mechanism of action is not yet fully understood (Berisha et al. 2019 ). Prostaglandins, derived from arachidonic acid through enzymatic reactions, are essential in synchronization protocols. Analogues used include Dinoprost tromethamine, similar to dinoprost tromethamine and natural D-cloprostenol (PGF2α), with a short half-life, cloprostenol sodium, with a longer half-life, and D-cloprostenol, which has a significantly prolonged half-life, making it more potent than natural PGF2α due to its resistance to enzymatic degradation (Bourne 1981 ; McCracken et al. 1999 ). The aim of this study was to investigate the effect of combining Dinoprost tromethamine and D-cloprostenol in a fixed-time artificial insemination (FTAI) protocol on ovulation and pregnancy rate in Nelore heifers. MATERIALS AND METHODS The work was conducted in accordance with ethical standards and received approval from the institution's Ethics and Biosafety Committee. The animal research was carried out in accordance with the guidelines of the institutional animal use committee, under case number 02/2020. This study was conducted in the municipality of Salinas, Minas Gerais, Brazil, (Latitude: -16.1434S, Longitude: -42.3031O), at an altitude of 513 meters, and the experimental period was from June to August. All the animals used in the experiment were of the Nelore breed ( Bos taurus indicus ) and were kept on pasture made up of Brachiaria brizanthacv. Marandu with water and mineral salt supplied ad libitum . 28 nulliparous Nelore heifers were used, with an average age of 24 to 30 months and a body weight of over 330 kg. Only cyclic females were selected (confirmed by ultrasound evaluations with the presence of corpus luteum (CL) at the beginning of the experiment, with a body condition score (BCS) ≥ 2.75 (on a scale of 1 to 5 where 1 is very thin and 5 is obese, according to Espasandin et al. ( 2023 ). The animals underwent a gynecological examination using rectal palpation and transrectal ultrasound (MIDRAY DP 2200®, São Paulo, SP, Brazil, 6 MHz linear transducer) to assess cyclicity and suitability for inclusion in the experiment. The design was entirely randomized, with the heifers (n = 28) randomly divided into three treatments (T) with homogeneous live weight, body score and age / T1 (n = 8), T2 (n = 11) and T3 (n = 9). The drugs used in the experiment were administered intramuscularly using a 40 x 1.2 mm needle (Solidor®, Barueri, Brazil) and 3.0 and 5.0 mL syringes (SR®, Pedro Juan Caballero, Paraguay). On day zero of the experiment (D0), all the animals were given 2 mL of estradiol benzoate (Gonadiol®, Zoetis, São Paulo, Brazil) and a 1.9 g progesterone intravaginal device (CIDR®, Zoetis, São Paulo, Brazil) was inserted. On day seven (D7), 2.5 mL of Dinoprost tromethamine (Lutalyse®, Zoetis, São Paulo, Brazil) was administered, and on day nine (D9) all the females had the intravaginal device removed, they were given 0.3 mL of oestradiol cypionate (ECP®, Zoetis, São Paulo, Brazil) and 1.5 mL, equivalent to 300 IU of equine chorionic gonadotrophin (eCG) (Novormon®, Zoetis, São Paulo-SP, Brazil). On day eleven (D11) all the animals were inseminated with doses of semen from the bull (REM screaming, Select Sires Beef) and divided into batches, where the animals belonging to T1 received 2 mL of placebo solution (Saline 0. 9%), T2 received 2.0 mL of D-cloprost tromethamine (Lutalyse®, Zoetis) and T3 2.0 mL of D-cloprostenine (Lutalyse®, Zoetis), 9%), T2 received 2.5 mL of Dinoprost tromethamine (Lutalyse®, Zoetis, São Paulo, Brazil) and T3 2.0 mL of D-cloprostenol (Prolise®, Agener União, Embu-Guaçu-SP, Brazil). Experimental scheme adopted (Fig. 1 ). The Jamovi program was used to compare the means. Non-parametric variables were evaluated using the Friedman test, with a significance level of 5%. All variables passed the Shapiro-Wilk normality tests. The animals in each experimental group underwent ultrasound evaluation on days D0, D7, D9 and D11 of the fixed-time artificial insemination (FTAI) protocol, as well as on days D30 after insemination. The ovaries were scanned using a Mindray ultrasound machine, model DP 2200VET, equipped with a 7.5 MHz transrectal linear transducer, to determine ovulation, follicular development (follicular diameter) and luteal development (luteal diameter) and the animal's uterine conditions. Diagnosis was carried out 30 days after insemination. RESULTS In Group T1 - Control, six animals showed follicular growth from day 7 (D7) of the protocol, the daily growth rate was 5.46 mm from the removal of the intravaginal progesterone device (P4), and the growth from the removal on day 9 (D9) to day 11 (D11) was 7.246 mm and from ovulation to the formation of the corpus luteum (CL) was 3.63 mm. On D7, the animals in the control group had an average follicular diameter (FOLD) > 7.35 mm, of which only one had a FOLD < 5 mm, which reached its largest diameter of 6.9 mm during the protocol, and this animal did not ovulate. The overall average FOLD of ovulating females at D11 was 10.83 mm, while the average CL was 14.47 mm. With this treatment, two animals showed follicular regression, with a higher FOLD at D7 and a lower FOLD at D11 (9.75 -6.25mm) and (8.2-6.55mm) respectively. With this treatment, four animals ovulated, with the lowest FOLD being 10.2mm and only one animal achieved pregnancy. For the females that ovulated, the average FOLD on day D11 for this group was 10.83mm, with a daily growth of 3.63mm, forming a CL with an average of 14.47mm, so 4 females ovulated out of a total of 8, and only 01 became pregnant (Figs. 2 and 3 ). In Group T2 - Dinoprost tromethamine, on D7, three animals had FOLsD averaging 6.9mm, while the rest of the batch had FOLsD 5mm, with daily growth of 7.24mm, and from the moment of withdrawal to ovulation, growth was 1.93mm. The average dominant follicle (FD) at D11 was 10.82mm. With this treatment, five (05) of the 11 females ovulated, of which two ovulated with follicles < 10mm and one became pregnant. Of the 5 females that ovulated, 3 (three) became pregnant, with a mean follicular diameter on D11 of 10.82mm, with daily growth of 3.8mm, forming an average CL of 14.62mm (Figs. 4 and 5 ). In Group T3 - D-cloprostenol, on D7, five animals out of a total of nine had FOLsD > 5mm, on D9 the average FOLD was 8.17mm, with an average daily growth of 0.97mm. On D11, 02 females ovulated with FOLsD < 10mm, with this characteristic, 01 female ovulated with a FOLD of 8.3 mm and obtained a positive pregnancy diagnosis. Of the five females that ovulated, all became pregnant. In this study, only 50% (14/28) of the heifers ovulated within 72 hours of removing the progesterone device, showing 50% (4/8), 45% (5/11) and 55% (5/9) for groups T1, T2 and T3 respectively. Of the heifers that ovulated, 64.28% (9/14) became pregnant. The other groups T1, T2 and T3 had pregnancy rates for females that ovulated of (25%, 60% and 100%) respectively. The animals in treatment T3 and T2 had higher reproductive efficiency (p 0.05), but there was a significant difference in the pregnancy rate compared to the control group. The heifers ovulated up to three days after removal of the controlled internal drug release device (CIDR). There was a difference in follicular divergence between the T1, T2 and T3 treatment groups. The T2 and T3 treatment groups had greater follicular divergence on D9, when the progesterone device was removed. However, the heifers in the T1 control group ovulated follicles with average diameters of 10.83mm, and the other groups had diameters of 10.82mm and 10.47mm for T2 and T3 respectively, forming a CL with diameters of 14.47, 14.62 and 14.04mm (Fig. 6 ). In this study, the addition of a second dose of PGF2α on the day of artificial insemination had similar effects to the E2/P4 ovulation synchronization protocol on the pregnancy rate in Nelore heifers (Fig. 7 ). Below (Table 1 ) shows the differences found, as well as their respective treatments. Table 1 Effect of follicular recruitment, growth, ovulation, and corpus luteum (CL) formation in Nelore heifers treated with dinoprost tromethamine and D-cloprostenol (PGF2α) for ovulation synchronization. Treatment group Average Ø FOLD on D7 (mm) Average Ø FOLD on D9 (mm) Average Ø FOLD on D11 (mm) Growth rate between D9 and OV (mm/day) Average Ø CL (mm) Growth rate between FOLD and CL (mm/day) T1 (Control) 7,65 9,3 10,83 0,765 14,47 3,64 T2 (Dinoprost trometamine) 6,9 8,7 10,82 1,06 14,62 3,8 T3 (D-cloprostenol) 8,35 8,63 10,47 1,785 14,04 3,57 FOLD: follicular diameter; CL: corpus luteum; OV: ovulation; D: day. DISCUSSION The use of D-cloprostenol on day 11 (D11) of the synchronization protocol increased the pregnancy rate and promoted ovulation of follicles smaller than 10mm. This is in line with Noronha et al. ( 2020 ), who concluded in their study that the positive effect of using an additional dose of dinoprost tromethamine in fixed-time artificial insemination (FTAI) protocols in Nelore cows provided ovulation in follicles smaller than 13.1mm (Table 2 ). Table 2 Synchronization rates of the dominant follicle (number of animals with follicles > 5 mm at D9 in relation to the total treated), ovulation and pregnancy in Nelore heifers submitted to estrus synchronization and ovulation induction with PGF₂α. Treatments Ovulation (%) Pregnancy (%) T1 (Control) 4/8 (50) 1/8 (12,5) T2 (Dinoprost trometamine) 5/11 (45,45) 3/11 (27,27) T3 (D-cloprostenol) 5/9 (55,55) 5/9 (55,55) Nascimento ( 2019 ), evaluated the luteolytic efficiency of different doses of cloprostenol sodium and dinoprost tromethamine, concluding that the main difference observed after the administration of cloprostenol sodium and dinoprost tromethamine occurred in the effect exerted by the active ingredients on large and small luteal cells. In this study, administration on D11 (diestrus) was effective in inducing complete luteolysis, causing a reduction in progesterone concentration, luteal dimensions (ultrasound and morphometry) and vascularization, as well as inducing caspase-3 activity. The luteolytic capacity of using 50% of the dose of cloprostenol sodium and dinoprost tromethamine, when administered during diestrus (D11), was also demonstrated. In addition to its luteolytic activity, prostaglandin can increase the effects of exogenous progesterone on the hypothalamus after withdrawal of the progesterone source, inducing an increase in the pituitary response to gonadotropin-releasing hormone (GnRH) (Murdoch et al. 1993 ; Weems et al. 2006 ). A study conducted by Ramírez et al. ( 2018 ), using Dinoproste and D- cloprostenol in the cyclonization of ewes, concluded that D-coprostenol was more efficient in responding to the presentation of oestrus, but the two treatments obtained equal pregnancy rates. Carlson et al. ( 1973 ), in their study with sheep, reported the action of dinoprost tromethamine and D-cloprostenol (PGF2α) on the release of luteinizing hormone (LH), which could be a possible mechanism of action of prostaglandins, since ovulation in cows and heifers is associated with the LH peak. Albuquerque et al. ( 2014 ) evaluated the effect of using an additional dose of prostaglandin during a fixed-time artificial insemination (FTAI) protocol based on oestradiol and progesterone on the fertility of lactating Holstein cows in anestrus, concluding that treatment with a second dose of PGF2 improves pregnancy during artificial insemination (AI) in normothermic anovulatory cows due to increased ovulation and improved characteristics of the pre-ovulatory follicle. Given the experimental conditions, it can be concluded that the use of two doses of PGF2α (dinoprost tromethamine and D-cloprostenol) was efficient in synchronizing small follicles (< 10mm) during the FTAI protocol, as well as increasing the pregnancy rate. Declarations Acknowledgements The authors did not receive any specific funding or support for this study. Statement of animal ethics All captures and handling of the heifers were approved by the Ethics Committee for Animal Experiments, in accordance with local laws and regulations. The work was carried out in accordance with ethical standards and received the approval of the institution's Ethics and Biosafety Committee. The animal research was carried out in accordance with the guidelines of the institutional animal use committee, under process number 02/2020. This study was carried out in the municipality of Salinas, Minas Gerais, Brazil. Conflict of interest statement The authors declare no conflict of interest. Funding The authors declare that no funds, grants, or other support were received during the preparation of this manuscript. Competing Interests The authors have no relevant financial or non-financial interests to disclose. Author Contributions Betania B. Santos: conceptualization, development of the methodology, project management, investigation, practical implementation in heifer management, data visualization, writing of the primary manuscript and revision of the manuscript. Francisco L. S. Franco, Glenner Z. Silva: research, practical implementation of heifer management and revision of the manuscript. Ingrid R. N. Martins, Israel P. Linhares: visualization and organization of the data, standardization according to the journal's norms, writing, reviewing and editing the manuscript. Wolff C. M. Filho: research supervision, technical guidance, methodology validation, resource management and critical review of the manuscript. All the authors revised the manuscript. Data Availability The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request. Ethics approval This study was conducted in accordance with the guidelines established by the institutional committee for the use and care of animals. All procedures involving animals were approved by the Animal Use Ethics Committee of the Instituto Federal do Norte de Minas Gerais - Campus Salinas, under protocol number 02/2020. The research was carried out in the municipality of Salinas, Minas Gerais, Brazil, and followed all national and international standards of ethics in animal experimentation, with the aim of ensuring the welfare of the animals and minimizing any form of discomfort. Consent to participate Informed consent was obtained from all individual participants included in the study. References Associação Brasileira das Indústrias Exportadoras de Carne (ABIEC). http://abiec.com.br/publicacoes/mais-tecnologia-mas-sustentabilidade/. Acessado em 29 de dezembro de 2024. Albuquerque JP, Vasconcelos MLJ, Dias PH, et al (2014) Evaluation of two types of prostaglandin (Dinoprot vs Cloprostenol) during an E2 abd P4 based estrus synchronization protocol. Animal reproduction. https://www2.zoetis.com.br/content/_assets/[MGR]-Bovinos/[MGR]-Gerar/[MGR]-pdf/[MGR]-Compromisso-technico/Informativo-T%C3%A9cnico-10-Dinoprost-X-Cloprostenol-(Leite).pdf. Acessado em 29 de dezembro de 2024. Baruselli SP, Santos FFG, Crepaldi AG, at al (2022). FTAI in numbens: evolution and future projection. Revista Brasileira de Reprodução Animal 46:76-83. doi: https://doi.org/10.21451/1809- 3000.RBRA2022.007 Berisha B, Rodler D, Schams D, et al (2019) Prostaglandins in Superovulation Induced Bovine Follicles During the Preovulatory Period and Early Corpus Luteum. Frontiers in Endocrinology 10: 1-8. doi: https://doi.org/10.3389/fendo.2019.00467 Bourne GR (1981) A review of metabolism and clearance studies with 14C-cloprostenol in the cow. Acta Vet Scand Suppl 77:5-9. Carlson JC, Barcikowski B, McCracken JA (1973) PGF2α and the release of LH in 1190 sheep. Journal of Reproduction and Fertility 34:357-361. Espasandin AC, Cardoso SA, Lópes N (2023) Guided classification measurement of body condition in beef cows. Agrociencia Uruguay 27:1-9. doi: https://doi.org/10.31285/AGRO.27.1165 Júnior NPA, Perazzi FP, Nogueira HC, et al (2020) Inclusion of nutraceutecs in the diet of lactation cows and their effects on animal performance. Brazilian Journal of Animal and Environmental Research 3:502-508. doi: https://doi.org/10.34188/bjaerv3n2-009 Macmillan K, Gobikrushanth M, Plastow G, et al (2020) Performance and optimization of an ear tag automated activity monitor for estrus prediction in dairy heifers. Theriogenology 155: 197-204. doi: https://doi.org/10.1016/j.theriogenology.2020.06.018 Marques NMN (2019) Effect of reusing CIDR and PRID on the reproduction of Holstein-Friesian heifers. Dissertation, Escola Superior Agrária de Castelo Branco. McCracken JA, Custer EE, Lamsa JC (1999) Luteolysis: a neuroendocrine-mediated event. Physiol Rev 79:263-323. doi: https://doi.org/10.1152/physrev.1999.79.2.263 Murdoch, W. J., Hansen, T. R., & McPherson, L. A. (1993). A review — Role of eicosanoids in vertebrate ovulation. Prostaglandins, 46:85–115. doi: https://doi.org/10.1016/0090-6980(93)90037-8 Nascimento GAM (2019) Evaluation of the luteolytic efficiency of different doses of cloprostenol sodium and dinoprost tromethamine administered on days 4 and 11 of the estrous cycle in beef females. Dissertation, Universidade Estadual Paulista (UNESP), Faculdade de Ciências Agrárias e Veterinária. Noronha MI, Cooke FR, Martins FGC, et al (2020) Administering an additional prostaglandin F2a injection to Bos indicus beef cows during a treattment regimen for fixed- time artificial insemination. Animal Reproduction Science 219:106-535. doi: https://doi.org/10.1016/j.anireprosci.2020.106535 Ramírez, AA; Villalvazo, MMV; Arredondo, SE; Ramírez, HA e Sevilla, MFH 2018. d- Cloprostenol enchances estrus synchronization in tropical hair sheep. Tropical Animal Health and Production. V. 50, p. 991-996. doi: https://doi.org/10.1007/S11250-018-1522-X Rodrigues ADP (2016) Reproductive performance in Bos indicus and Bos taurus X Bos indicus heifers submitted to ovulation synchronization protocols. Dissertation, Universidade de São Paulo, Faculdade de Medicina Veterinária e Zootecnia. Weems CW, Weems YS, Randel RD (2006) Prostaglandins and reproduction in female farm animals. The Veterinary Journal 171:206-228. doi: https://doi.org/10.1016/j.tvjl.2004.11.014. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6508312","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":446638007,"identity":"91215416-132e-4cc7-95d7-0cffb538f1b6","order_by":0,"name":"Betânia B. 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S.","lastName":"Franco","suffix":""},{"id":446638009,"identity":"6ddd9c88-cc9e-4c4b-82fe-826db7a03ac5","order_by":2,"name":"Ingrid R. N. Martins","email":"","orcid":"","institution":"Universidade Federal de Viçosa","correspondingAuthor":false,"prefix":"","firstName":"Ingrid","middleName":"R. N.","lastName":"Martins","suffix":""},{"id":446638010,"identity":"d9360217-2d26-41bd-a825-aaf175e41082","order_by":3,"name":"Israel P. Linhares","email":"","orcid":"","institution":"Universidade Federal de Viçosa","correspondingAuthor":false,"prefix":"","firstName":"Israel","middleName":"P.","lastName":"Linhares","suffix":""},{"id":446638011,"identity":"2ef02a7c-ff71-4f7d-a15f-43f8e0a8e697","order_by":4,"name":"Glenner Z. Silva","email":"","orcid":"","institution":"Instituto Federal do Norte de Minas Gerais","correspondingAuthor":false,"prefix":"","firstName":"Glenner","middleName":"Z.","lastName":"Silva","suffix":""},{"id":446638012,"identity":"ea340c87-c64a-425a-90c8-47d0042045a5","order_by":5,"name":"Wolff C. M. Filho","email":"","orcid":"","institution":"Instituto Federal Goiano","correspondingAuthor":false,"prefix":"","firstName":"Wolff","middleName":"C. M.","lastName":"Filho","suffix":""}],"badges":[],"createdAt":"2025-04-23 03:08:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6508312/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6508312/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":81631980,"identity":"3a8058a1-7160-4705-ad48-74525191fdf0","added_by":"auto","created_at":"2025-04-29 11:39:49","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":18410,"visible":true,"origin":"","legend":"\u003cp\u003eDiagram of the experiment's activities. On D0, application of 2 mL of oestradiol benzoate and insertion of the progesterone device for 9 days, on D7, application of 2.5 mL of dinoprost tromethamine, D9 removal of the P4 implant, application of 0.3 mL of oestradiol cypionate, 1.5 mL of eCG, and on D11, insemination in all the animals, with separation of the treatment batches, T1 application of 2 mL of a placebo solution, T2 application of 2.5 mL of dinoprost tromethamine and T3 2 mL of D-cloprostenol.\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/1f376fd72da7640411dcf42b.png"},{"id":81632660,"identity":"e1bb5281-8174-4556-8b6c-10e8b77a0072","added_by":"auto","created_at":"2025-04-29 11:47:49","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":38063,"visible":true,"origin":"","legend":"\u003cp\u003eFollicular dynamics of a heifer from Treatment T1 (Control), diagnosed as non-pregnant. Note the persistence of the corpus luteum (CL), with regression in the size of the dominant follicle (DF) and absence of ovulation. There was also regression of the persistent follicle over the days following FTAI (Fixed-Term Artificial Insemination).\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/e3a61a2c504b9327a604978e.png"},{"id":81631983,"identity":"f9014bfe-dbe4-46b8-8ad3-953ed01a0cf3","added_by":"auto","created_at":"2025-04-29 11:39:49","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":25868,"visible":true,"origin":"","legend":"\u003cp\u003eFollicular dynamics of a heifer from Treatment T1 (Control), with a positive pregnancy diagnosis. Follicular diameter (DFOL) growth can be seen from D7, with divergence on D9 and ovulation between D11 and D12, evidenced by the disappearance of the follicle and the formation of the corpus luteum (CL).\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/b01d18528ad93bbf3648f820.png"},{"id":81631982,"identity":"72082cfe-934e-422b-bb55-b309cdfa069e","added_by":"auto","created_at":"2025-04-29 11:39:49","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":40565,"visible":true,"origin":"","legend":"\u003cp\u003eFollicular dynamics of a heifer in group T2 (Dinoprost tromethamine), showing synchronization of the follicular wave, regression of the corpus luteum (CL), follicular recruitment on D9, day of Fixed-Term Artificial Insemination (FTAI), presence of a subordinate follicle (SF), divergence and persistence of a dominant follicle (DF), followed by ovulation of the persistent DF and a positive pregnancy diagnosis.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/0ceba5ebdb57a3d3143249e5.png"},{"id":81633777,"identity":"f8e2b06a-c7f4-4e63-9a48-78dc4250ac65","added_by":"auto","created_at":"2025-04-29 11:55:49","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":44647,"visible":true,"origin":"","legend":"\u003cp\u003eFollicular dynamics of a heifer in group T2 (Dinoprost tromethamine), with a positive pregnancy diagnosis, showing follicular recruitment on D9, day of Fixed-Term Artificial Insemination (FTAI), the presence of a subordinate follicle (SF), divergence between follicles (LF1 and LF2), persistence of a dominant follicle (DF) and ovulation of the persistent DF. There is also regression of the corpus luteum (CL).\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/d11071e1d81782791d571994.png"},{"id":81633776,"identity":"d153a264-86ad-454e-a036-e2083920c243","added_by":"auto","created_at":"2025-04-29 11:55:49","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":50673,"visible":true,"origin":"","legend":"\u003cp\u003eFollicular dynamics of a heifer treated with T3 (D-cloprostenol) after FTAI (Fixed-Term Artificial Insemination) during the experimental period. Divergence of dominant follicles (DF), regression of subordinate follicles (SF) and ovulation with corpus luteum formation (CL), with a positive pregnancy diagnosis.\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/79b06fa19735cdabd8ad416c.png"},{"id":81632664,"identity":"b6ef1ad3-ba5b-4f11-9f91-d8832b5ded7b","added_by":"auto","created_at":"2025-04-29 11:47:49","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":91748,"visible":true,"origin":"","legend":"\u003cp\u003eAverages of treatment groups, luteal dynamics, follicular growth on days D7, D9, D11 and D12, ovulation (OV) and corpus luteum formation (CL), in Nelore heifers treated for ovulation synchronization with D-cloprostenol and Dinoprost tromethamine.\u003c/p\u003e","description":"","filename":"floatimage7.png","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/06616d49ba5ae12e8f909767.png"},{"id":81651263,"identity":"62563d3e-8f6e-48c0-8737-838d0d0e4e2b","added_by":"auto","created_at":"2025-04-29 16:16:15","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":746505,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6508312/v1/486c9620-dc45-498a-9dd9-4a6b17aacca7.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"D-cloprostenol and dinoprost tormetamine at the time of artificial insemination in Nelore heifers: luteal dynamics and pregnancy rate","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eThe demand for food of animal origin with a known origin has made the market more demanding in terms of animal production and its effects (Junior et al. 2020).\u003c/p\u003e \u003cp\u003eFrom 1990 to 2020, meat production in Brazil grew by 122%, with a 159% increase in productivity, and the area of pasture used decreased by 13.6%, due to investments in breeding technology and biotechnology (ABIEC, 2021).\u003c/p\u003e \u003cp\u003eDespite advances in reproductive biotechnology, beef heifers face reproductive efficiency challenges, affecting economic viability (Baruselli et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eEarly gestation of heifers in the first breeding season is crucial to ensure quality calves and a longer productive life (Rodrigues \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), with the use of fixed-time estrous synchronization (Marques et al. 2019).\u003c/p\u003e \u003cp\u003eResearch aims to improve fixed-time artificial insemination (FTAI) protocols for heifers, with emphasis on the use of prostaglandins to optimize results (Macmillan et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), some of which have shown positive effects of prostaglandins in ovulation induction, although their mechanism of action is not yet fully understood (Berisha et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eProstaglandins, derived from arachidonic acid through enzymatic reactions, are essential in synchronization protocols. Analogues used include Dinoprost tromethamine, similar to dinoprost tromethamine and natural D-cloprostenol (PGF2α), with a short half-life, cloprostenol sodium, with a longer half-life, and D-cloprostenol, which has a significantly prolonged half-life, making it more potent than natural PGF2α due to its resistance to enzymatic degradation (Bourne \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e1981\u003c/span\u003e; McCracken et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e1999\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe aim of this study was to investigate the effect of combining Dinoprost tromethamine and D-cloprostenol in a fixed-time artificial insemination (FTAI) protocol on ovulation and pregnancy rate in Nelore heifers.\u003c/p\u003e"},{"header":"MATERIALS AND METHODS","content":"\u003cp\u003e The work was conducted in accordance with ethical standards and received approval from the institution's Ethics and Biosafety Committee. The animal research was carried out in accordance with the guidelines of the institutional animal use committee, under case number 02/2020.\u003c/p\u003e \u003cp\u003eThis study was conducted in the municipality of Salinas, Minas Gerais, Brazil, (Latitude: -16.1434S, Longitude: -42.3031O), at an altitude of 513 meters, and the experimental period was from June to August.\u003c/p\u003e \u003cp\u003eAll the animals used in the experiment were of the Nelore breed (\u003cem\u003eBos taurus indicus\u003c/em\u003e) and were kept on pasture made up of Brachiaria brizanthacv. Marandu with water and mineral salt supplied \u003cem\u003ead libitum\u003c/em\u003e.\u003c/p\u003e \u003cp\u003e28 nulliparous Nelore heifers were used, with an average age of 24 to 30 months and a body weight of over 330 kg. Only cyclic females were selected (confirmed by ultrasound evaluations with the presence of corpus luteum (CL) at the beginning of the experiment, with a body condition score (BCS)\u0026thinsp;\u0026ge;\u0026thinsp;2.75 (on a scale of 1 to 5 where 1 is very thin and 5 is obese, according to Espasandin et al. (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2023\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe animals underwent a gynecological examination using rectal palpation and transrectal ultrasound (MIDRAY DP 2200\u0026reg;, S\u0026atilde;o Paulo, SP, Brazil, 6 MHz linear transducer) to assess cyclicity and suitability for inclusion in the experiment.\u003c/p\u003e \u003cp\u003eThe design was entirely randomized, with the heifers (n\u0026thinsp;=\u0026thinsp;28) randomly divided into three treatments (T) with homogeneous live weight, body score and age / T1 (n\u0026thinsp;=\u0026thinsp;8), T2 (n\u0026thinsp;=\u0026thinsp;11) and T3 (n\u0026thinsp;=\u0026thinsp;9). The drugs used in the experiment were administered intramuscularly using a 40 x 1.2 mm needle (Solidor\u0026reg;, Barueri, Brazil) and 3.0 and 5.0 mL syringes (SR\u0026reg;, Pedro Juan Caballero, Paraguay).\u003c/p\u003e \u003cp\u003eOn day zero of the experiment (D0), all the animals were given 2 mL of estradiol benzoate (Gonadiol\u0026reg;, Zoetis, S\u0026atilde;o Paulo, Brazil) and a 1.9 g progesterone intravaginal device (CIDR\u0026reg;, Zoetis, S\u0026atilde;o Paulo, Brazil) was inserted. On day seven (D7), 2.5 mL of Dinoprost tromethamine (Lutalyse\u0026reg;, Zoetis, S\u0026atilde;o Paulo, Brazil) was administered, and on day nine (D9) all the females had the intravaginal device removed, they were given 0.3 mL of oestradiol cypionate (ECP\u0026reg;, Zoetis, S\u0026atilde;o Paulo, Brazil) and 1.5 mL, equivalent to 300 IU of equine chorionic gonadotrophin (eCG) (Novormon\u0026reg;, Zoetis, S\u0026atilde;o Paulo-SP, Brazil). On day eleven (D11) all the animals were inseminated with doses of semen from the bull (REM screaming, Select Sires Beef) and divided into batches, where the animals belonging to T1 received 2 mL of placebo solution (Saline 0. 9%), T2 received 2.0 mL of D-cloprost tromethamine (Lutalyse\u0026reg;, Zoetis) and T3 2.0 mL of D-cloprostenine (Lutalyse\u0026reg;, Zoetis), 9%), T2 received 2.5 mL of Dinoprost tromethamine (Lutalyse\u0026reg;, Zoetis, S\u0026atilde;o Paulo, Brazil) and T3 2.0 mL of D-cloprostenol (Prolise\u0026reg;, Agener Uni\u0026atilde;o, Embu-Gua\u0026ccedil;u-SP, Brazil). Experimental scheme adopted (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe Jamovi program was used to compare the means. Non-parametric variables were evaluated using the Friedman test, with a significance level of 5%. All variables passed the Shapiro-Wilk normality tests.\u003c/p\u003e \u003cp\u003eThe animals in each experimental group underwent ultrasound evaluation on days D0, D7, D9 and D11 of the fixed-time artificial insemination (FTAI) protocol, as well as on days D30 after insemination. The ovaries were scanned using a Mindray ultrasound machine, model DP 2200VET, equipped with a 7.5 MHz transrectal linear transducer, to determine ovulation, follicular development (follicular diameter) and luteal development (luteal diameter) and the animal's uterine conditions. Diagnosis was carried out 30 days after insemination.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003eIn Group T1 - Control, six animals showed follicular growth from day 7 (D7) of the protocol, the daily growth rate was 5.46 mm from the removal of the intravaginal progesterone device (P4), and the growth from the removal on day 9 (D9) to day 11 (D11) was 7.246 mm and from ovulation to the formation of the corpus luteum (CL) was 3.63 mm.\u003c/p\u003e \u003cp\u003eOn D7, the animals in the control group had an average follicular diameter (FOLD)\u0026thinsp;\u0026gt;\u0026thinsp;7.35 mm, of which only one had a FOLD\u0026thinsp;\u0026lt;\u0026thinsp;5 mm, which reached its largest diameter of 6.9 mm during the protocol, and this animal did not ovulate. The overall average FOLD of ovulating females at D11 was 10.83 mm, while the average CL was 14.47 mm. With this treatment, two animals showed follicular regression, with a higher FOLD at D7 and a lower FOLD at D11 (9.75 -6.25mm) and (8.2-6.55mm) respectively. With this treatment, four animals ovulated, with the lowest FOLD being 10.2mm and only one animal achieved pregnancy.\u003c/p\u003e \u003cp\u003eFor the females that ovulated, the average FOLD on day D11 for this group was 10.83mm, with a daily growth of 3.63mm, forming a CL with an average of 14.47mm, so 4 females ovulated out of a total of 8, and only 01 became pregnant (Figs.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn Group T2 - Dinoprost tromethamine, on D7, three animals had FOLsD averaging 6.9mm, while the rest of the batch had FOLsD\u0026thinsp;\u0026lt;\u0026thinsp;3mm. On D9 of treatment, all the animals had FOLsD\u0026thinsp;\u0026gt;\u0026thinsp;5mm, with daily growth of 7.24mm, and from the moment of withdrawal to ovulation, growth was 1.93mm. The average dominant follicle (FD) at D11 was 10.82mm.\u003c/p\u003e \u003cp\u003eWith this treatment, five (05) of the 11 females ovulated, of which two ovulated with follicles\u0026thinsp;\u0026lt;\u0026thinsp;10mm and one became pregnant. Of the 5 females that ovulated, 3 (three) became pregnant, with a mean follicular diameter on D11 of 10.82mm, with daily growth of 3.8mm, forming an average CL of 14.62mm (Figs.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn Group T3 - D-cloprostenol, on D7, five animals out of a total of nine had FOLsD\u0026thinsp;\u0026gt;\u0026thinsp;5mm, on D9 the average FOLD was 8.17mm, with an average daily growth of 0.97mm. On D11, 02 females ovulated with FOLsD\u0026thinsp;\u0026lt;\u0026thinsp;10mm, with this characteristic, 01 female ovulated with a FOLD of 8.3 mm and obtained a positive pregnancy diagnosis. Of the five females that ovulated, all became pregnant.\u003c/p\u003e \u003cp\u003eIn this study, only 50% (14/28) of the heifers ovulated within 72 hours of removing the progesterone device, showing 50% (4/8), 45% (5/11) and 55% (5/9) for groups T1, T2 and T3 respectively. Of the heifers that ovulated, 64.28% (9/14) became pregnant.\u003c/p\u003e \u003cp\u003eThe other groups T1, T2 and T3 had pregnancy rates for females that ovulated of (25%, 60% and 100%) respectively. The animals in treatment T3 and T2 had higher reproductive efficiency (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) when comparing the variable with group T1.\u003c/p\u003e \u003cp\u003eThe use of dinoprost tromethamine and D-cloprostenol (PGF2α) as an ovulation inducer did not interfere with ovulation rates (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05), but there was a significant difference in the pregnancy rate compared to the control group.\u003c/p\u003e \u003cp\u003eThe heifers ovulated up to three days after removal of the controlled internal drug release device (CIDR). There was a difference in follicular divergence between the T1, T2 and T3 treatment groups. The T2 and T3 treatment groups had greater follicular divergence on D9, when the progesterone device was removed. However, the heifers in the T1 control group ovulated follicles with average diameters of 10.83mm, and the other groups had diameters of 10.82mm and 10.47mm for T2 and T3 respectively, forming a CL with diameters of 14.47, 14.62 and 14.04mm (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn this study, the addition of a second dose of PGF2α on the day of artificial insemination had similar effects to the E2/P4 ovulation synchronization protocol on the pregnancy rate in Nelore heifers (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eBelow (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) shows the differences found, as well as their respective treatments.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eEffect of follicular recruitment, growth, ovulation, and corpus luteum (CL) formation in Nelore heifers treated with dinoprost tromethamine and D-cloprostenol (PGF2α) for ovulation synchronization.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTreatment group\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAverage \u0026Oslash; FOLD on D7 (mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAverage \u0026Oslash; FOLD on D9 (mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAverage \u0026Oslash; FOLD on D11 (mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eGrowth rate between D9 and OV (mm/day)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eAverage \u0026Oslash; CL\u003c/p\u003e \u003cp\u003e(mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eGrowth rate between FOLD and CL (mm/day)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT1 (Control)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e7,65\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e9,3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10,83\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0,765\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e14,47\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e3,64\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT2 (Dinoprost trometamine)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e6,9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e8,7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10,82\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1,06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e14,62\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e3,8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT3 (D-cloprostenol)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e8,35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e8,63\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10,47\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1,785\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e14,04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e3,57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eFOLD: follicular diameter; CL: corpus luteum; OV: ovulation; D: day.\u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eThe use of D-cloprostenol on day 11 (D11) of the synchronization protocol increased the pregnancy rate and promoted ovulation of follicles smaller than 10mm. This is in line with Noronha et al. (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), who concluded in their study that the positive effect of using an additional dose of dinoprost tromethamine in fixed-time artificial insemination (FTAI) protocols in Nelore cows provided ovulation in follicles smaller than 13.1mm (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSynchronization rates of the dominant follicle (number of animals with follicles\u0026thinsp;\u0026gt;\u0026thinsp;5 mm at D9 in relation to the total treated), ovulation and pregnancy in Nelore heifers submitted to estrus synchronization and ovulation induction with PGF₂α.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTreatments\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eOvulation (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePregnancy (%)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT1 (Control)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4/8 (50)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1/8 (12,5)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT2 (Dinoprost trometamine)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5/11 (45,45)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3/11 (27,27)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT3 (D-cloprostenol)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5/9 (55,55)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5/9 (55,55)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eNascimento (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), evaluated the luteolytic efficiency of different doses of cloprostenol sodium and dinoprost tromethamine, concluding that the main difference observed after the administration of cloprostenol sodium and dinoprost tromethamine occurred in the effect exerted by the active ingredients on large and small luteal cells. In this study, administration on D11 (diestrus) was effective in inducing complete luteolysis, causing a reduction in progesterone concentration, luteal dimensions (ultrasound and morphometry) and vascularization, as well as inducing caspase-3 activity. The luteolytic capacity of using 50% of the dose of cloprostenol sodium and dinoprost tromethamine, when administered during diestrus (D11), was also demonstrated.\u003c/p\u003e \u003cp\u003eIn addition to its luteolytic activity, prostaglandin can increase the effects of exogenous progesterone on the hypothalamus after withdrawal of the progesterone source, inducing an increase in the pituitary response to gonadotropin-releasing hormone (GnRH) (Murdoch et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1993\u003c/span\u003e; Weems et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2006\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eA study conducted by Ram\u0026iacute;rez et al. (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), using Dinoproste and D- cloprostenol in the cyclonization of ewes, concluded that D-coprostenol was more efficient in responding to the presentation of oestrus, but the two treatments obtained equal pregnancy rates.\u003c/p\u003e \u003cp\u003eCarlson et al. (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e1973\u003c/span\u003e), in their study with sheep, reported the action of dinoprost tromethamine and D-cloprostenol (PGF2α) on the release of luteinizing hormone (LH), which could be a possible mechanism of action of prostaglandins, since ovulation in cows and heifers is associated with the LH peak.\u003c/p\u003e \u003cp\u003eAlbuquerque et al. (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) evaluated the effect of using an additional dose of prostaglandin during a fixed-time artificial insemination (FTAI) protocol based on oestradiol and progesterone on the fertility of lactating Holstein cows in anestrus, concluding that treatment with a second dose of PGF2 improves pregnancy during artificial insemination (AI) in normothermic anovulatory cows due to increased ovulation and improved characteristics of the pre-ovulatory follicle.\u003c/p\u003e \u003cp\u003eGiven the experimental conditions, it can be concluded that the use of two doses of PGF2α (dinoprost tromethamine and D-cloprostenol) was efficient in synchronizing small follicles (\u0026lt;\u0026thinsp;10mm) during the FTAI protocol, as well as increasing the pregnancy rate.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors did not receive any specific funding or support for this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatement of animal ethics\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll captures and handling of the heifers were approved by the Ethics Committee for Animal Experiments, in accordance with local laws and regulations. The work was carried out in accordance with ethical standards and received the approval of the institution\u0026apos;s Ethics and Biosafety Committee. The animal research was carried out in accordance with the guidelines of the institutional animal use committee, under process number 02/2020. This study was carried out in the municipality of Salinas, Minas Gerais, Brazil.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no conflict of interest.\u003c/p\u003e\n\u003cp\u003eFunding\u003c/p\u003e\n\u003cp\u003eThe authors declare that no funds, grants, or other support were received during the preparation of this manuscript.\u003c/p\u003e\n\u003cp\u003eCompeting Interests\u003c/p\u003e\n\u003cp\u003eThe authors have no relevant financial or non-financial interests to disclose.\u003c/p\u003e\n\u003cp\u003eAuthor Contributions\u003c/p\u003e\n\u003cp\u003eBetania B. Santos: conceptualization, development of the methodology, project management, investigation, practical implementation in heifer management, data visualization, writing of the primary manuscript and revision of the manuscript.\u003c/p\u003e\n\u003cp\u003eFrancisco L. S. Franco, Glenner Z. Silva: research, practical implementation of heifer management and revision of the manuscript.\u003c/p\u003e\n\u003cp\u003eIngrid R. N. Martins, Israel P. Linhares: visualization and organization of the data, standardization according to the journal\u0026apos;s norms, writing, reviewing and editing the manuscript.\u003c/p\u003e\n\u003cp\u003eWolff C. M. Filho: research supervision, technical guidance, methodology validation, resource management and critical review of the manuscript.\u003c/p\u003e\n\u003cp\u003eAll the authors revised the manuscript.\u003c/p\u003e\n\u003cp\u003eData Availability\u003c/p\u003e\n\u003cp\u003eThe datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003eEthics approval\u003c/p\u003e\n\u003cp\u003eThis study was conducted in accordance with the guidelines established by the institutional committee for the use and care of animals. All procedures involving animals were approved by the Animal Use Ethics Committee of the Instituto Federal do Norte de Minas Gerais - Campus Salinas, under protocol number 02/2020. The research was carried out in the municipality of Salinas, Minas Gerais, Brazil, and followed all national and international standards of ethics in animal experimentation, with the aim of ensuring the welfare of the animals and minimizing any form of discomfort.\u003c/p\u003e\n\u003cp\u003eConsent to participate\u003c/p\u003e\n\u003cp\u003eInformed consent was obtained from all individual participants included in the study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAssocia\u0026ccedil;\u0026atilde;o Brasileira das Ind\u0026uacute;strias Exportadoras de Carne (ABIEC). http://abiec.com.br/publicacoes/mais-tecnologia-mas-sustentabilidade/. Acessado em 29 de dezembro de 2024.\u003c/li\u003e\n\u003cli\u003eAlbuquerque JP, Vasconcelos MLJ, Dias PH, et al (2014) Evaluation of two types of prostaglandin (Dinoprot vs Cloprostenol) during an E2 abd P4 based estrus synchronization protocol. Animal reproduction. https://www2.zoetis.com.br/content/_assets/[MGR]-Bovinos/[MGR]-Gerar/[MGR]-pdf/[MGR]-Compromisso-technico/Informativo-T%C3%A9cnico-10-Dinoprost-X-Cloprostenol-(Leite).pdf. Acessado em 29 de dezembro de 2024.\u003c/li\u003e\n\u003cli\u003eBaruselli SP, Santos FFG, Crepaldi AG, at al (2022). FTAI in numbens: evolution and future projection. Revista Brasileira de Reprodu\u0026ccedil;\u0026atilde;o Animal 46:76-83. doi: https://doi.org/10.21451/1809- 3000.RBRA2022.007\u003c/li\u003e\n\u003cli\u003eBerisha B, Rodler D, Schams D, et al (2019) Prostaglandins in Superovulation Induced Bovine Follicles During the Preovulatory Period and Early Corpus Luteum. Frontiers in Endocrinology 10: 1-8. doi: https://doi.org/10.3389/fendo.2019.00467\u003c/li\u003e\n\u003cli\u003eBourne GR (1981) A review of metabolism and clearance studies with 14C-cloprostenol in the cow. Acta Vet Scand Suppl 77:5-9.\u003c/li\u003e\n\u003cli\u003eCarlson JC, Barcikowski B, McCracken JA (1973) PGF2\u0026alpha; and the release of LH in 1190 sheep. Journal of Reproduction and Fertility 34:357-361.\u003c/li\u003e\n\u003cli\u003eEspasandin AC, Cardoso SA, L\u0026oacute;pes N (2023) Guided classification measurement of body condition in beef cows. Agrociencia Uruguay 27:1-9. doi: https://doi.org/10.31285/AGRO.27.1165\u003c/li\u003e\n\u003cli\u003eJ\u0026uacute;nior NPA, Perazzi FP, Nogueira HC, et al (2020) Inclusion of nutraceutecs in the diet of lactation cows and their effects on animal performance. Brazilian Journal of Animal and Environmental Research 3:502-508. doi: https://doi.org/10.34188/bjaerv3n2-009 \u003c/li\u003e\n\u003cli\u003eMacmillan K, Gobikrushanth M, Plastow G, et al (2020) Performance and optimization of an ear tag automated activity monitor for estrus prediction in dairy heifers. Theriogenology 155: 197-204. doi: https://doi.org/10.1016/j.theriogenology.2020.06.018 \u003c/li\u003e\n\u003cli\u003eMarques NMN (2019) Effect of reusing CIDR and PRID on the reproduction of Holstein-Friesian heifers. Dissertation, Escola Superior Agr\u0026aacute;ria de Castelo Branco.\u003c/li\u003e\n\u003cli\u003eMcCracken JA, Custer EE, Lamsa JC (1999) Luteolysis: a neuroendocrine-mediated event. Physiol Rev 79:263-323. doi: https://doi.org/10.1152/physrev.1999.79.2.263\u003c/li\u003e\n\u003cli\u003eMurdoch, W. J., Hansen, T. R., \u0026amp; McPherson, L. A. (1993). A review \u0026mdash; Role of eicosanoids in vertebrate ovulation. Prostaglandins, 46:85\u0026ndash;115. doi: https://doi.org/10.1016/0090-6980(93)90037-8\u003c/li\u003e\n\u003cli\u003eNascimento GAM (2019) Evaluation of the luteolytic efficiency of different doses of cloprostenol sodium and dinoprost tromethamine administered on days 4 and 11 of the estrous cycle in beef females. Dissertation, Universidade Estadual Paulista (UNESP), Faculdade de Ci\u0026ecirc;ncias Agr\u0026aacute;rias e Veterin\u0026aacute;ria.\u003c/li\u003e\n\u003cli\u003eNoronha MI, Cooke FR, Martins FGC, et al (2020) Administering an additional prostaglandin F2a injection to Bos indicus beef cows during a treattment regimen for fixed- time artificial insemination. Animal Reproduction Science 219:106-535. doi: https://doi.org/10.1016/j.anireprosci.2020.106535\u003c/li\u003e\n\u003cli\u003eRam\u0026iacute;rez, AA; Villalvazo, MMV; Arredondo, SE; Ram\u0026iacute;rez, HA e Sevilla, MFH 2018. d- Cloprostenol enchances estrus synchronization in tropical hair sheep. Tropical Animal Health and Production. V. 50, p. 991-996. doi: https://doi.org/10.1007/S11250-018-1522-X \u003c/li\u003e\n\u003cli\u003eRodrigues ADP (2016) Reproductive performance in Bos indicus and Bos taurus X Bos indicus heifers submitted to ovulation synchronization protocols. Dissertation, Universidade de S\u0026atilde;o Paulo, Faculdade de Medicina Veterinária e Zootecnia.\u003c/li\u003e\n\u003cli\u003eWeems CW, Weems YS, Randel RD (2006) Prostaglandins and reproduction in female farm animals. The Veterinary Journal 171:206-228. doi: https://doi.org/10.1016/j.tvjl.2004.11.014.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Assisted reproduction, Bovine females, Corpus luteum, Pregnancy, Prostaglandin","lastPublishedDoi":"10.21203/rs.3.rs-6508312/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6508312/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe aim of this study was to evaluate the effect of combining Dinoprost tromethamine and D-cloprostenol in a fixed-time artificial insemination (FTAI) protocol on ovulation, fertility and pregnancy rate in Nelore heifers. A total of 28 Nelore heifers were evaluated. On D0, they received an intravaginal progesterone (P4) device and an intramuscular (IM) injection of 2 mg of oestradiol benzoate (BE). On D7, an application of 2mL of dinoprost tromethamine, on D9 the P4 device was removed and 0.3 mL of oestradiol cypionate (eCP) and 1.5 mL of equine chorionic gonadotrophin (eCG) were administered, on D11 all the females were inseminated and randomly divided into three groups: T1, T2 and T3. The control (T1; n\u0026thinsp;=\u0026thinsp;8), in which the animals received 2 mL of saline IM; T2 (n\u0026thinsp;=\u0026thinsp;11), received 2 mL of dinoprost tromethamine IM; and T3 (n\u0026thinsp;=\u0026thinsp;9), received 2 mL of D-Cloprostenol IM. Follicle diameter was assessed on D0, D7, D9, D11 and corpus luteum (CL) diameter on D12. Gestation diagnosis (GD) was carried out 30 days after artificial insemination (AI). Non-parametric variables were analyzed using the Friedman test, with a significance level of 5%. The pregnancy rate was different between treatments (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), being higher in T3 when compared to T1 (55.55%) vs. (12.5%), as well as T2 (55.55%) vs. (27.27%). There was no interaction between treatments in terms of follicle diameter at D11 and CL formation. It was concluded that the use of two doses of PGF2α (dinoprost tromethamine and D-cloprostenol) was efficient in synchronizing small follicles (\u0026lt;\u0026thinsp;10mm) and increasing pregnancy rates.\u003c/p\u003e","manuscriptTitle":"D-cloprostenol and dinoprost tormetamine at the time of artificial insemination in Nelore heifers: luteal dynamics and pregnancy rate","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-29 11:39:44","doi":"10.21203/rs.3.rs-6508312/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"34f9b493-c251-43a2-8b17-2569c181fc14","owner":[],"postedDate":"April 29th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-04-29T16:08:08+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-29 11:39:44","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6508312","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6508312","identity":"rs-6508312","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}