Ovulation rate, prolificacy
and pregnancy rate in goats
treated with oral glycerol
Ubaldo Aguilar1*
Joel Hernández Cerón1*
Yesmín Domínguez2
Carlos G. Gutiérrez1
1 Departamento de Reproducción
2 Centro de Enseñanza, Investigación
y Extensión en el Altiplano
Facultad de Medicina Veterinaria y Zootecnia
Universidad Nacional Autónoma de México
Av. Universidad 3000, Ciudad de México
04510, México
*Corresponding author:
Tel: +52 5 5622-5860
Email address: jhc@unam.mx
Received: 2015-10-12
Accepted: 2016-03-16
Published: 2016-03-28
DOI: http://dx.doi.org/10.21753/vmoa.3.1.360
Additional information and declarations here
Copyright 2016 Ubaldo Aguilar et al.
open access
Distributed under Creative Commons CC-BY 4.0
Abstract
This study tested whether the oral administration of glycerol at the time of progestin removal and in the first 6 days of the estrous cycle increased the ovulation rate, prolificacy, and pregnancy rate in goats. Intravaginal sponges impregnated with fluorogestone acetate were inserted into 129 goats for 12 days; upon sponge removal, goats were randomly assigned to one of the two following treatment groups: the glycerol group (n = 65), which received an oral drench of 100 mL of glycerol upon sponge removal and was repeated on days 0, 2, 4, and 6 following estrus (estrus = day 0), and the control group (n = 64), which did not receive glycerol. The goats in estrus were mated, and their ovulation rate was determined by ultrasonography between days 8 and 12 of the estrous cycle. Pregnancy was diagnosed by ultrasonography on day 40, and the prolificacy was determined at birth. In 6 goats treated with glycerol and 5 controls, their insulin concentrations at 0, 2, 4, 8, and 12 h after the glycerol drench were determined by radioimmunoassay. The proportion of goats with multiple ovulations (glycerol = 71 vs control = 64) and the proportion of goats with multiple births (glycerol = 52 vs control = 56) were similar (P > 0.05) between treatments. Likewise, the pregnancy rate was similar (P > 0.05) between treatments (glycerol = 88 vs control = 85%). The insulin concentrations tended to be higher in goats treated with glycerol (P = 0.08). In conclusion, an oral drench of 100 mL of glycerol at the time of progestin withdrawal and in the first 6 days of the estrous cycle did not increase the ovulation rate, prolificacy, or pregnancy rate in goats.
Keywords: Ovulation rate; Prolificacy; Glycerol; Insulin; Goats
Introduction
An increase in the ovulation rate and prolificacy in small ruminants has been achieved in an efficient way through hormone treatments and by increasing the energy content in the diet (flushing). Although the flushing mechanism is not yet fully understood, the evidence indicates that its effects are produced at the ovarian level; furthermore, they are independent of gonadotropin concentrations and are associated with increases in the blood glucose and insulin levels (Downing et al., 1995; Muñoz-Gutiérrez et al., 2004; Dupont et al., 2014).
In sheep, short-term supplementation with lupin grain, a legume with a high metabolizable energy and protein content, can increase ovulation rates. Short-term supplementation with lupin grain stimulates folliculogenesis and increases ovulation rates, without inducing changes in body weight and condition (Scaramuzzi et al., 2006). Glucose and insulin may influence the ovulation rate by stimulating follicle development, resulting in a higher number of follicles capable of responding to gonadotropin stimulus (Gutiérrez et al., 1997; Viñoles et al., 2005). Furthermore, glucose and glucosamine stimulate follicle steroidogenic capacity by increasing insulin-like growth factor type I (IGF-I) activity (Muñoz-Gutiérrez et al., 2004). Studies performed in goats show that an increase in insulin, either induced by diet or by insulin administration, increases the ovulation rate (Suguna et al., 2009; Zabuli et al., 2010).
The administration of glycogenic solutions can cause an increase in the blood glucose and insulin concentrations. In sheep, short-term treatments with glycerol and propylene glycol have been used to increase ovulation rates. Rodriguez Iglesias et al. (1996) were able to increase the ovulation rate in seasonally anovulatory ewes through the oral administration of 100 mL of a glycerol and propylene glycol solution, which were administered immediately before ram exposure. Gutierrez et al. (2011) observed increased serum glucose concentrations, insulin, and ovulation rates with a single oral dose of 300 mL of glycerol during the induction of luteolysis with PGF2α in Pelibuey sheep. In the same study, a similar effect on the ovulation rate with a single oral dose of 100 mL of glycerol was achieved.
Moreover, insulin promotes early embryonic development. In vitro insulin exerts a mitogenic and an antiapoptotic effect on embryos (Byrne et al., 2002; Augustin et al., 2003), resulting in an increased proportion of embryos reaching the blastocyst stage. Additionally, high insulin concentrations negatively affect embryonic development in cattle (Fouladi-Nashta et al., 2006) and sheep (Carrera-Chávez et al., 2014). Furthermore, a positive effect from insulin on luteal function has been observed. In a goat study, insulin was associated with higher concentrations of progesterone during pregnancy (Suguna et al., 2009). High levels of progesterone are associated with accelerated embryonic development (Garret et al., 1988), which could translate into fewer embryonic losses. In dairy cows, the administration of 1 L of oral glycerol during the first 6 days post-insemination increases blood insulin levels and increases the pregnancy rate (Ortega et al., 2010). Therefore, this study tested whether the oral administration of 100 mL of glycerol, at the time of progestin removal and in the first 6 days of the estrous cycle, increased the ovulation rate, prolificacy, and pregnancy rate in goats.
Materials and methods
Location
This study was conducted in an experimental station at the Facultad de Medicina Veterinaria y Zootecnia of Universidad Nacional Autónoma de México (UNAM), located in Tequisquiapan, Querétaro, at a latitude of 20°31’21” N and at 1881 m above sea level. The regional climate is temperate and semi-arid, with an average temperature of 17.5º C and an average annual rainfall of 512 mm (García, 1981). The experiment was carried out from September to November, which corresponds to the reproductive season of goats in Mexico (Arvizu et al., 1995).
Animals
In this experiment, 129 crossbreed (Boer-Alpine French) cycling goats (22 doelings and 107 does varying in parity) with a body condition score between 2 and 3 were included (Russel et al., 1969). The goats were confined and fed a diet based on alfalfa hay, corn silage, and concentrate according to NRC requirements. This experiment was approved by the Consejo Académico del Posgrado en Ciencias de la Producción y de la Salud Animal from UNAM.
Treatments
In all goats, an intravaginal sponge impregnated with 45 mg of fluorogestone acetate (FGA; Chronogest; MSD Animal Health, Mexico; Intervet México, S.A. de C.V., Huixquilucan, Estado de México, México) was inserted and remained in situ for 12 days. When the sponge was removed, a luteolytic dose of sodium cloprostenol (Celosil; MSD Animal Health, Mexico) was administered. Upon sponge withdrawal, the goats were randomly assigned to one of the following treatment groups: the glycerol (n = 65) group, which received an oral drench of 100 mL of glycerol repeated on days 0, 2, 4, and 6 following estrus (estrus = day 0), and the control (n = 64) group, which did not receive glycerol. Males fitted with an apron performed estrus detection. Females in estrus were mated to males of proven fertility.
Between days 8 and 12 post-mating, the ovulation rate was determined via a transrectal ultrasound exam by counting the number of corpora lutea; for this purpose, a 7.5 MHz linear transducer (Aloka, Co., Ltd., Tokyo, Japan) was used, which was adapted to a rigid support to allow for handling (Gutierrez et al., 2011). The pregnancy rate was determined by ultrasonographic pregnancy diagnosis on day 40 after service. Prolificacy was determined at birth (Fig. 1).
Figure 1. A schematic overview of the treatments.
Blood sampling and sample processing
To determine blood levels of progesterone and insulin, 6 goats were treated with oral glycerol and 5 controls were randomly sampled. Blood samples for progesterone quantification were taken daily between days 1 and 17 of the estrous cycle. Two goats in the control group presented with short cycles; therefore, they were not included in the progesterone statistical analysis. On day 4 of the estrous cycle, sampling was conducted in order to determine blood insulin levels induced by the oral administration of glycerol. The first sample was taken before treatment (0 h), and the next samples were taken at 2, 4, 8, and 12 h after the glycerol drench.
Blood samples (10 mL) were obtained by puncture of the jugular vein using Vacutainer® tubes containing 100 µL of sodium citrate. The blood samples were centrifuged at 1500 x g for 10 min to separate the plasma, which was subsequently stored at -20 °C until analysis. The progesterone and insulin concentrations were determined by solid-phase radioimmunoassay (Coat-A-Count®, RIA kit, DPC; USA). In the case of progesterone, the assay sensitivity was 0.1 ng/mL with an intra-assay variation coefficient of 4.2%, while the insulin sensitivity was 0.052 ng/mL, with an intra-assay variation coefficient of 3.3%.
Statistical analysis
A logistic regression model was used to determine the effect of oral glycerol treatment on the percentage of goats with multiple ovulations and multiple births. Likewise, the effect of parity (primiparous and multiparous) and its interaction with treatment was tested by logistic regression [SAS version 9.2 (SAS Institute Inc., Cary, NC)]. The ovulation rate and prolificacy were compared by the Mann-Whitney U test, in which the effect of treatment and its interaction with parity were considered. The differences in the progesterone and insulin concentrations between treatments were tested by ANOVA for repeated measurements; the model included treatment, time, and their interactions. In all cases, P < 0.05 was considered to be significantly different and P ≤ 0.10 was considered to be a tendency.
Results and Discussion
The oral administration of 100 mL of glycerol at the time of progesterone withdrawal did not increase the ovulation rate, and the proportion of goats with multiple ovulations or multiple births was similar between treatments (Tables 1 and 2). The ovulation rate [glycerol= 1.84 ± 0.64 vs control=1.80 ±0.72 (mean ± standard deviation)] and the prolificacy [glycerol= 1.58 ± 0.61 vs control=1.60 ±0.64 (mean ± standard deviation)] were similar between treatments. These results are different from those observed in sheep by Rodriguez Iglesias et al. (1996) and Gutierrez et al. (2011). The reason the treatment did not have a favorable effect on the ovulation rate may be related to the metabolic response to the oral glycerol doses. The 100 mL glycerol dose was determined based on the results obtained in sheep by Gutierrez et al. (2011); in their study, the administration of 100 mL glycerol caused an increase in the ovulation rate similar to the one obtained with 300 mL. For this reason, the administration of 100 mL was chosen. In the present study, serum insulin concentrations were not affected by the treatment (P = 0.11), but there was an interaction between the treatment and time (P = 0.08). Thus, insulin concentrations tended to be higher within 2 and 4 h post-treatment (Fig. 2). Likely, this slight increase in insulin concentrations was not enough to improve the ovulation rate. The cause of the variation in response to the same glycerol dose observed between sheep and goats is unknown. Studies evaluating glycogenic solutions have been performed in sheep and cattle, but not in goats (Rodriguez-Iglesias et al., 1996; Ortega et al., 2010; Gutierrez et al., 2011). Although these 3 species are ruminants, they present differences in their digestive function and their use of forages. Goats have physiological adaptations that allow them to leverage high fiber feeds more efficiently (Silanikove, 2000). Therefore, they may be less sensitive to variations in the quality and quantity of offered feeds. Oral glycerol is used as a substrate in rumen fermentation from which propionate can be obtained. However, it can also be absorbed directly as glycerol through the rumen mucosa, which will be converted later into glucose in the liver (Trabue et al., 2007). Although this metabolic pathway is the same in ruminants, its efficiency in triggering insulin secretion may be different between species; this could explain, in part, the success of the treatment in increasing the ovulation rate in sheep as well as its failure in goats. The results of this study promote the development of future studies in goats to evaluate different glycerol doses, until the dose that triggers an optimal response in serum insulin is determined.
Table 1. Percentage of goats with multiple ovulations that received an oral drench of 100 mL glycerol.
Table 2. Percentage of goats kidding more than one kid that received an oral drench of 100 mL glycerol.
Figure 2. Average insulin concentrations (± standard error) in goats treated with 100 mL of glycerol () and controls (
). Treatment by time interaction: P = 0.08.
Furthermore, this study proposed that the oral administration of glycerol during the first days of embryo development (days 0, 2, 4, and 6 of the estrous cycle) favors embryonic survival, which would increase the proportion of pregnant goats and prolificacy. This hypothesis was based on the expected effect of glycerol on insulin concentrations and the effects of insulin on embryonic development. Studies performed in vitro and in vivo show that insulin improves embryonic development by increasing cell proliferation and decreasing apoptosis (Augustin et al., 2003; Suguna et al., 2009). Meanwhile, low insulin concentrations adversely affect embryonic development and interferon tau production (Thatcher et al., 1995). Furthermore, in dairy cows, an increase in insulin concentrations induced by the oral administration of 1 L of glycerol during the first 6 days of embryonic development (days 0, 2, 4, and 6) has been shown to improve pregnancy rates (Ortega et al., 2010). In the present study, however, the pregnancy rate (glycerol = 88 vs control = 85%) and the proportion of multiple births were similar between treatments (P > 0.05; Table 2). The lack of effect on prolificacy and pregnancy rate observed in this study was probably due to the marginal increase in insulin concentrations after the oral administration of 100 mL of glycerol. Furthermore, the pregnancy rate obtained in both groups was high, which makes it more difficult to identify a positive effect from any treatment aimed at improving fertility. Parity was observed to have an effect on ovulation rate and prolificacy; however, there was no interaction between the treatment and parity (Tables 1 and 2; P > 0.1).
The administration of oral glycerol during the first days of the estrous cycle may favor embryonic development and conception rate by increasing serum progesterone concentrations. This hypothesis is based on studies in cattle in which insulin increases circulating progesterone concentrations (Spicer y Echternkamp, 1995; Cooke et al., 2012). In this study, nonetheless, progesterone concentrations between 1 and 17 days post-mating were similar between treatments, and no interaction was observed between the treatment and time (P > 0.05; Fig. 3).
Figure 3. Average concentrations of progesterone (± standard error) in goats treated with 100 mL of oral glycerol () and controls (
) (P > 0.05).
The good body condition of the goats used in this study, as well as the season in which the treatment was applied, may have influenced the lack of a positive response to glycerol administration. The goats received a diet that met the NRC requirements, and they had a body condition between 2 and 3 at the beginning of the experiment, which is reflective of the conditions associated with a high pregnancy rate and high prolificacy and are similar to those obtained consistently within the flock. Conversely, this study was completed during the height of the reproductive period, which contributed to the excellent response in fertility. Future experiments should be conducted in conditions in which the fertility and prolificacy are diminished, such as in cyclicity induction programs during anestrus or in marginally nourished goats; in these circumstances, the supplementation proposed in this study may improve fertility. Treatments exist that do not increase pregnancy rates globally, but rather do so in subgroups of animals. In a previous study conducted by our group, for example, bovine recombinant somatotropin only increased pregnancy rate in goats in anestrus, as well as only increased prolificacy in primiparous goats, who naturally tend to have a lower pregnancy rate and prolificacy (Hernández y Gutiérrez, 2012).
Conclusions
In conclusion, the oral administration of 100 mL of glycerol at the time of progestin withdrawal and in the first six days of the estrous cycle did not increase the ovulation rate, prolificacy, or pregnancy rate in goats.
Funding
Part of this study was funded by project PAPIIT IN22235 of UNAM, Mexico.
Acknowledgments
The authors are grateful to the workers and technicians of the experimental station for providing the facilities, animals, and constant support during the study.
The authors declare that they have no conflicts of interest.
Author contributions
Ubaldo Aguilar performed the experiment and analyzed the results.
Yesmín Domínguez performed the experiment.
Joel Hernández Cerón and Carlos G. Gutiérrez designed the experiment, analyzed the results and wrote the manuscript.
References
1) Arvizu R, Hernández Cerón J, Alberti A, Porras A, Valencia J. 1995. Inicio de la actividad ovárica posparto y características de la primera ovulación de cabras criollas paridas en dos épocas del año. Avances en Investigación Agropecuaria 4:9-15.
2) Augustin R, Pocar P, Wrenzycki C, Niemann H, Fischer B. 2003. Mitogenic and anti-apoptotic activity of insulin on bovine embryos produced in vitro. Reproduction 126:91-99.
3) Byrne AT, Southgate J, Brison DR, Leese HJ. 2002. Regulation of apoptosis in the bovine blastocyst by insulin and insulin-like growth factor (IGF) superfamily. Molecular Reproduction and Development 62:489-495.
4) Carrera-Chávez JM, Hernández-Cerón J, López-Carlos MA, Lozano-Domínguez RR, Molinar F, Echavarría-Cháirez FG, Bañuelos-Valenzuela R, Aréchiga CF. 2014. Superovulatory response and embryo development in ewes treated with two doses of bovine somatotropin. Animal Reproduction Science 151: 105-111.
5) Cooke RF, Cappellozza BI, Reis MM, Bohnert DW, Vasconcelos JL. 2012. Plasma progesterone concentration in beef heifers receiving exogenous glucose, insulin, or bovine somatotropin. Journal of Animal Science 90:3266–3273.
6) Downing JA, Joss J, Scaramuzzi RJ. 1995. Ovulation rate and the concentrations of gonadotrophins and metabolic hormones in ewes infused with glucose during the late luteal phase of the oestrous cycle. Journal of Endocrinology 146:403-410.
7) Dupont J, Scaramuzzi RJ, Reverchon M. 2014. The effect of nutrition and metabolic status on the development of follicles, oocytes and embryos in ruminants. Animal 8:1031–1044.
8) Fouladi-Nashta AA, Campbell KHS. 2006. Dissociation of oocyte nuclear and cytoplasmic maturation by the addition of insulin in cultured bovine antral follicles. Reproduction 131; 449-460.
9) García E. 1981. Modificaciones al Sistema de Clasificación Climática de Köppen. 5ª edición. México, D. F., México: Instituto de Geografía, UNAM.
10) Garret JE, Geisert RD, Zavy MT, Morgan GL. 1988. Evidence for maternal regulation of early conceptus growth and development in beef cattle. Journal of Reproduction and Fertility 84:437-446.
11) Gutiérrez CG, Oldham J, Bramley TA, Gong JG, Campbell BK, Webb R. 1997. The recruitment of ovarian follicles is enhanced by increased dietary intake in heifers. Journal of Animal Science 75:1876-1884.
12) Gutierrez CG, Ferraro S, Martinez V, Saharrea A, Cortez C, Lassala A, Basurto H, Hernandez Ceron J. 2011. Increasing ovulation quota: more than a matter of energy. Acta Scientiae Veterinariae 39 (Suppl 1):305-316.
13) Hernández-Cerón J, Gutiérrez CG. La somatotropina bovina recombinante y la reproducción en bovinos, ovinos y caprinos. Agrociencia 47: 35-45. 2013.
14) Muñoz-Gutiérrez M, Blache D, Martin GB, Scaramuzzi RJ. 2004. Ovarian follicular expression of mRNA encoding the type I IGF receptor and IGF-binding protein-2 in sheep following five days of nutritional supplementation with glucose, glucosamine or lupins. Reproduction 128:747-756.
15) Ortega LA, Hernández-Cerón J, Gutiérrez CG. 2010. La administración oral de glicerol después de la inseminación incrementa el porcentaje de concepción en vacas Holstein. Técnica Pecuaria en México 48:69-74.
16) Rodríguez-Iglesias RM, Ciccioli NH, Irazoqui H, Giglioli C. 1996. Ovulation rate in ewes after single oral glucogenic dosage during a ram-induced follicular phase. Animal Reproduction Science 44:211-221.
17) Russel AJF, Doney JM, Gunn RG. 1969. Subjective assessment of body fat in live sheep. Journal of Agricultural Science 72:451-454.
18) Scaramuzzi RJ, Campbell BK, Downing JA, Kendall NR, Khalid M, Muñoz-Gutiérrez M, Somchit A. 2006. A review of the effects of supplementary nutrition in the ewe on the concentrations of reproductive and metabolic hormones and the mechanisms that regulate folliculogenesis and ovulation rate. Reproduction Nutrition Development 46:339-354.
19) Silanikove N. 2000. The physiological basis of adaptation in goats to harsh environments. Small Ruminant Research 35:181-193.
20) Spicer, LJ, Echternkamp SE. 1995. The ovarian insulin andinsulin-like growth factor system with an emphasis on domestic animals. Domestic Animal Endocrinology 12:223–245.
21) Suguna K, Mehrotra S, Agarwal SK, Hoque M, Shanker U, Singh SK, Varshney VP. 2009. Effect of exogenous insulin administration on ovarian function, embryo/fetal development during pregnancy in goats. Animal Reproduction Science 111:202-213.
22) Thatcher WW, Meyer MD, Danet-Desnoyers G. 1995. Maternal recognition of pregnancy. Journal of reproduction and Fertility 49 (Suppl.), 15–28.
23) Trabue S, Scoggin K, Tjandrakusuma S, Rasmussen MA, Reilly PJ. 2007. Ruminal Fermentation of Propylene Glycol and Glycerol Journal of Agricultural and Food Chemistry 55:7043–7051.
24) Viñoles C, Forsberg M, Martin GB, Cajarville C, Repetto J, Meikle A. 2005. Short-term nutritional supplementation of ewes in low body condition affects follicle development due to an increase in glucose and metabolic hormones. Reproduction 129:299-309.
25) Zabuli J, Tanaka T, Lu W, Kamomae H. 2010. Intermittent nutritional stimulus by short-term treatment of high-energy diet promotes ovarian performance together with increases in blood levels of glucose and insulin in cyclic goats. Animal Reproduction Science 122:288-293.