The subtle balance of insulin and thyroxine on survival and development of in vitro cultured caprine preantral follicles enclosed in ovarian tissue.

Numerous studies have reported the importance of thyroid hormones on the development of later preantral and antral follicles, but their interactions with other hormones and effects in regulating early preantral follicle growth remain unclear. Here we investigated the in vitro effects of thyroxine combined with insulin on caprine preantral follicle survival and development. Sliced ovarian tissues were cultured for 1 or 7 days using 10 ng/mL (low) or 10 µg/mL (high) insulin in the presence of thyroxine at 0, 0.5, 1 or 2 µg/mL. Post-culture, we evaluated the follicular survival and development, assessed the expression of apoptotic-related genes (Bcl2/Bax) and receptors of insulin and thyroid hormones, and quantified the estradiol and reactive oxygen species (ROS) production levels. Follicular survival in low-insulin culture conditions was enhanced by the presence of 0.5 µg/mL thyroxine (P < 0.05) as compared to the thyroxine-free medium but remained similar to non-cultured control in the presence of 2 µg/mL (P > 0.05). Significantly higher ROS production was measured from Day 1 to Day 7 in low-insulin culture media containing 0.5 or 2 µg/mL thyroxine (P < 0.05). When compared to high insulin level, the presence of thyroxine in low insulin culture conditions yielded higher stromal cell density (P < 0.05), increased estradiol production on Day 1, and higher Bcl2/Bax ratio on Day 7. Cultures with high levels of both insulin and thyroxine led to follicles and oocytes with larger diameters (P < 0.05). The RNA transcript levels of insulin and thyroid receptors were reduced in the presence of high insulin cultures when compared to controls (non-cultured). In conclusion, the combination of low concentrations of insulin and thyroxine better maintained follicle survival, while high levels ensured better follicular development.

In vivo and in vitro strategies to support caprine preantral follicle development after ovarian tissue vitrification.

The aim of the present study was to compare fresh and vitrified goat ovarian tissue after autotransplantation and in vitro culture. Adult goats were completely ovariectomised and each ovarian pair was sliced and distributed among six different treatment groups: fresh control, fresh transplant, fresh culture, vitrified control, vitrified transplant and vitrified culture. Follicular morphology, development, growth, density, revascularisation and hormone production were evaluated in all groups. Three antral follicles (two in the fresh transplant and one in the vitrified transplant groups) were observed on the surface of the graft 90 days after transplantation. The percentage of morphologically normal follicles was similar in the fresh control, fresh transplant and vitrified transplant groups. The percentage of developing (transition, primary and secondary) follicles was higher after in vitro culture of fresh or vitrified tissue. Transplantation resulted in a lower follicle density. Serum oestradiol concentrations remained constant during the entire transplantation period. In contrast, progesterone production decreased significantly. Expression of CD31 mRNA was lower in fresh culture. In conclusion, restoration of goat ovarian function can be successfully achieved following transplantation of both fresh and vitrified goat ovarian tissue. However, transplantation induced higher follicle loss than in vitro culture.

Anethole improves the in vitro development of isolated caprine secondary follicles.

The aim of this study was to investigate the effect of three concentrations of anethole (30, 300, and 2000 µg/mL) on survival, antrum formation, follicular diameter, and oocyte maturation in the caprine species. The study also evaluated the effects of anethole on transcripts of ICAM-1, CAV-1, TIMP-2, and PAI-1 genes and levels of reactive oxygen species (ROS) in isolated goat preantral ovarian follicles before and after in vitro culture for 18 days. Preantral follicles were isolated from goat ovaries and individually cultured in alpha minimum essential medium modified (α-MEM+), defined as the control treatment, α-MEM+ supplemented with ascorbic acid at a concentration of 100 µg/mL (AA), or α-MEM+ supplemented with three different concentrations of anethole (30, 300, 2000 µg/mL) for a period of 18 days. Treatments were named as α-MEM+, AA, AN30, AN300, and AN2000, respectively. After culture, the follicles were opened, the cumulus oocytes complex (COCs) were removed and matured in vitro. The walls of the follicles were used for the quantitation of mRNA by quantitative real-time polymerase chain reaction. Finally, the medium collected at the end of culture was used for the measurements of ROS. After 18 days of culture, the AA treatment showed the percentage of intact follicles and follicular diameter significantly higher compared with the other treatments. However, daily growth rate, antrum formation, and also oocyte diameter were similar among the treatments. In addition, compared with AA, the rate of oocytes for in vitro maturation (diameter ≥ 110 µm) and the meiosis resumption rate were significantly higher in the treatments AN30 and AN2000, respectively. When assessing gene related to remodeling of the basement membrane, significant differences in mRNA levels for ICAM-1, CAV-1, TIMP-2, and PAI-1 were observed in comparison with Day 0, i.e., in the noncultured control. In addition, the ROS from Day 12, all treatments with the addition of anethole have significantly lower values of ROS than α-MEM+ and AA. In conclusion, the addition of anethole to the in vitro culture medium was able to improve the development of goat preantral follicles by reducing concentrations of ROS and increasing the percentage of oocytes able to resume meiosis.

Anti-Müllerian hormone reduces growth rate without altering follicular survival in isolated caprine preantral follicles cultured in vitro.

The aim of the present study was to evaluate the effect of anti-Müllerian hormone (AMH), with and without FSH, on the in vitro development of isolated caprine preantral follicles, as well as follicular steroid production and mRNA levels of AMH, hormone receptors (AMH and FSH), CYP19A1 (cytochrome P450, family 19, subfamily A, polypeptide 1), CYP17 (cytochrome P450, family 17, subfamily A, polypeptide 1), HSD3B (3-beta-hydroxysteroid dehydrogenase) and Myc (myelocytomatosis oncogene). Isolated secondary follicles were cultured in minimum essential medium alpha (α-MEM+) alone or supplemented with 50ng mL-1 AMH and/or 100ng mL-1 FSH added sequentially on different days of culture. Follicles were cultured for a total of 18 days, with different media during the first (Days 0-9) and second (Days 10-18) halves of the culture period, resulting in six treatment groups, as follows: α-MEM+/α-MEM+, FSH/FSH, AMH/AMH, AMH+FSH/AMH+FSH, AMH/FSH, and FSH/AMH. Follicle development was evaluated on the basis of follicular growth, oocyte maturation and steroid secretion. There was a decrease in follicular growth rate in the AMH, AMH+FSH and AMH/FSH treatment groups compared with α-MEM+ and FSH treatment groups (P<0.05). However, the different culture conditions had no effect on rates of meiotic resumption and steroid secretion (P>0.05). Moreover, follicles cultured in the presence of FSH had lower levels of AMH receptor type II (AMHRII) mRNA compared with non-cultured control (freshly isolated follicles), and the AMH and AMH/FSH treatment groups. In conclusion, AMH reduces the follicular growth rate of isolated goat preantral follicles in vitro without affecting follicular survival.

Caprine ovarian follicle requirements differ between preantral and early antral stages after IVC in medium supplemented with GH and VEGF alone or in combination.

The aim of the present study was to evaluate the effect of growth hormone (GH) and vascular endothelial growth factor (VEGF) added alone, sequentially or in combination, in the presence of insulin at physiological concentration (10 ng/mL) on the IVC of two different follicular categories: preantral (experiment 1; Exp.1) and early antral (experiment 2; Exp.2). Isolated follicles were individually cultured for 24 (Exp.1) and 18 days (Exp.2) in the following treatments: αMEM+ (Control), or Control medium supplemented with 50 ng/mL GH (GH), 100 ng/mL VEGF (VEGF), the combination of both (GH + VEGF), GH during the first 12 days and VEGF from Day 12 until the end of the culture (GH/VEGF) and vice versa (VEGF/GH). At the end of the culture, cumulus-oocyte complexes from in vitro-grown follicles were recovered and subjected to IVM. The following end points were evaluated: Follicle morphology, growth rates and antrum formation, production of estradiol, progesterone and testosterone, oocyte viability and meiotic stage, as well as relative expression of LHR, Amh, HAS2, PTGS2, CYP17, CYP19A1, and 3ßHSD. A considerable amount of viable fully grown oocytes were recovered after the IVC of early antral follicles in all treatments. Nevertheless, the GH treatment presented the highest percentage of fully grown oocytes (60%), mean oocyte diameter (117.74 ± 2.61 µm), and meiotic resumption (50%). Furthermore, GH treatment produced higher (P < 0.05) rates of metaphase II oocytes than all the other treatments, and similar LHR, Amh, and PTGS2 transcript levels to in vivo. Contrary to early antral follicles, preantral follicles were not affected by medium supplementation. In conclusion, the addition of GH to a culture medium containing physiological concentrations of insulin improves oocyte growth and maturation after the IVC of goat early antral follicles.

Fibrin-alginate hydrogel supports steroidogenesis, in vitro maturation of oocytes and parthenotes production from caprine preantral follicles cultured in group.

This study aimed to establish a culture system that improves the in vitro development of caprine preantral follicles. In a first experiment, follicles were encapsulated as a single unit per bead and cultured singly or in groups or with five follicles in the same alginate (ALG) bead for 18 days. In a subsequent experiment, the "five follicles per bead" design was chosen to culture in ALG, fibrin-alginate (FA) or hyaluronate (HA) for 18 days. In a third experiment, we chose the five follicles per bead in FA to culture for 30 days. The culture set-up of five follicles per ALG bead increased antrum formation and follicle diameter compared to the other culture designs (p < .05). Moreover, under this condition, 44.44% of the oocytes from in vitro cultured preantral follicles reached meiotic resumption. A significant increase of follicle diameter occurred in attachment system and FA (p < .05), but the ALG condition reached the highest among all groups on day 18 (p < .05). Follicles encapsulated in matrix produced more estradiol and progesterone than attachment system (p < .05). The expression of MMP-9 mRNA was higher in FA than in other groups (p < .05) and similar to antral follicles from in vivo control (p > .05). Only FA group resulted in oocytes matured. After 30 days, oocytes from preantral follicles in vitro grown in FA developed to eight-cell parthenotes. In conclusion, a culture system using FA supported the development of caprine preantral follicles cultured in group and included in the same bead of hydrogel, improving the oocyte maturation and producing parthenotes.

Modulation of aquaporins 3 and 9 after exposure of ovine ovarian tissue to cryoprotectants followed by in vitro culture.

Our aim has been to evaluate the effect of cryoprotective agents (CPAs) on the exposure, vitrification (VIT), and in vitro culture (IVC) of ovarian tissue with regard to the expression and immunolocalization of aquaporins (AQPs) 3 and 9 in ovine preantral follicles. Tissues were treated as follows: Experiment I: (1) control (without exposure to CPAs), (2) e-EG (exposure to ethylene glycol), (3) er-EG (exposure to and removal of EG), (4) e-DMSO (exposure to dimethyl sulfoxide), (5) er-DMSO (exposure to and removal of DMSO), (6) e-EG+DMSO (exposure to EG+DMSO), (7) er-EG+DMSO (exposure to and removal of EG+DMSO); Experiment II: (1) control, (2) VIT, (3) IVC, (4) VIT-IVC. In Experiment I, following er-EG or er-DMSO, tissue showed the down-regulation (P < 0.05) of AQP3 mRNA. The mRNA transcript levels were reduced (P < 0.05) for AQP9 in tissue following er-EG+DMSO. Immunolocalization was positive for both proteins (AQP3 and AQP9) on ovine preantral follicles following all treatments, except in the e-EG+DMSO group. In Experiment II, the mRNA levels of AQP3 and AQP9 following VIT treatment were similar (P > 0.05) to that of the control group. Nevertheless, VIT-IVC treatment led to the down-regulation of mRNA of AQP3 and AQP9. Thus, AQP3 and AQP9 act in a mutually dependent way, maintaining the cell homeostasis that is essential for the ovary cryopreservation process. Furthermore, the changes in the expression profiles of mRNA and protein after culture are a strong indicator that in vitro conditions have to be strictly controlled to ensure follicle viability and functionality.

Balance of insulin and FSH concentrations improves the in vitro development of isolated goat preantral follicles in medium containing GH.

The aim of this study was to evaluate the effect of different combinations of insulin and FSH concentrations in culture media containing GH on the in vitro follicle morphology, antrum formation, growth rates, estradiol (E2) production, oocyte viability and maturation as well as gene expression for FSHR, GHR, INSR, CYP19A1, CYP17, 3ßHSD. Secondary follicles were individually cultured for 18 days in a basic medium containing 50ng/mL GH supplemented with low insulin concentration (INS-LW: 10ng/mL) or high insulin concentration (INS-HG: 10µg/mL) alone or with a fixed FSH concentration (FSH100: 100ng/mL) or with increasing FSH concentrations (FSH-SEQ: 100ng/mL, days 0-6; 500ng/mL, days 6-12; 1000ng/mL days 12-18). In the INS-LW treatment was observed a higher (P<0.05) incidence of normal follicles at day 18 of culture. However, overall higher (P<0.05) follicular growth, oocyte diameter and meiotic resumption rates were obtained using INS-HG+FSH 100. The INS-HG and INS-HG+FSH100 treatments showed higher E2 production and mRNA levels for CYP19A1, CYP17, 3ßHSD when compared to INS-LW and INS-LW+FSH100. However, the addition of increasing FSH concentration, regardless of insulin concentration, did not improve the follicular growth, meotic resumption, E2 production or gene expression of steroidogenic enzymes when compared with INS-HG+FSH100. In conclusion, in presence of GH, a basic medium supplemented with 10µg/mL insulin and 100µg/mL FSH throughout the culture period, improves follicular and oocyte growth, oocyte meiotic resumption and E2 production from isolated preantral caprine follicles cultured in vitro.

Steady-state level of messenger RNA and immunolocalization of aquaporins 3, 7, and 9 during in vitro growth of ovine preantral follicles.

Aquaporins (AQPs) are a well-conserved family of small (approximately 30 kDa) membrane channel proteins that facilitate rapid movement of fluids and have a unique tissue-specific pattern of expression. These proteins have been found in the female reproductive systems of humans, rats, and mice. However, the expression and cellular localization of AQPs have not extensively been studied in the female reproductive system of sheep. Therefore, this study aimed to evaluate, by real-time polymerase chain reaction and immunohistochemistry respectively, the levels of messenger RNA and the immunolocalization of AQP3, AQP7, and AQP9 in large isolated ovine secondary follicles over a period of IVC. Our analysis revealed that AQP3 and AQP9 were present predominately in follicles that exhibited antrum formation, suggesting a crucial role of these AQPs in the formation of the antrum. Interestingly, AQP7 was only expressed in follicles that had not formed an antrum by Day 12 of culture. In conclusion, the presence of protein channels (AQP3 and AQP9) seems to be essential for the formation of the antrum in isolated ovine secondary follicles cultured in vitro and thus plays an important role during folliculogenesis in this species.

Differential gene expression and immunolocalization of platelet-derived growth factors and their receptors in caprine ovaries.

This study evaluated the messenger RNA (mRNA) expression and immunolocalization of all members of the platelet-derived growth factor (PDGF) family in caprine ovaries by quantitative PCR and immunohistochemistry, respectively. Detectable levels of PDGF-A mRNA were not observed in primordial follicles. Higher levels of PDGF-B mRNA were observed in primary follicles than in primordial follicles (P < 0.05). PDGF-D mRNA levels were higher in secondary follicles than in the other preantral follicle categories (P < 0.05). PDGF-B mRNA expression was higher than PDGF-C mRNA expression in primary follicles (P < 0.05). In antral follicles, PDGF-A mRNA expression was higher in cumulus-oocyte complexes (COCs) from small antral follicles than in those from large antral follicles and their respective granulosa/theca (GT) cells (P < 0.05). Furthermore, in COCs from small and large antral follicles, PDGF-A mRNA expression was higher than that of the other PDGF isoforms (P < 0.05). The mRNA levels of PDGF-B and PDGF-D and PDGFR-α and PDGFR-ß were higher in GT cells from large antral follicles than in GT cells from small antral follicles and in their respective COCs (P < 0.05). In COCs and GT cells from small antral follicles, the mRNA levels of PDGFR-α were higher than those of PDGFR-ß (P < 0.05). All proteins were observed in the cytoplasm of oocytes from all follicular categories. In granulosa cells, all PDGFs and PDGFR-ß were detected from starting at the secondary stage, and in theca cells, all proteins, except PDGF-C, were detected starting at the antral stage. In conclusion, PDGF and its receptors are differentially expressed in the oocytes and ovarian cells according to the stage of follicular development, suggesting their role in the regulation of folliculogenesis in goats.

Catalase addition to vitrification solutions maintains goat ovarian preantral follicles stability.

The aim of this study was to verify whether the addition of catalase (20 IU/mL) at different steps of goat ovarian tissue vitrification affects ROS levels, follicular morphology and viability, stromal cell density, apoptosis and the expression of proteins related to DNA-damage signaling (γH2AX) and repair (53BP1). Goat ovarian tissues were analyzed fresh (control) or after vitrification: without catalase (VS-/WS-), with catalase in vitrification solutions (VS+/WS-), with catalase in washing solutions (VS-/WS+) or with catalase in both solutions (VS+/WS+). The vitrification without catalase had higher ROS levels than the control. The catalase, regardless the step of addition, maintained ROS levels similar to the control. There were no difference between treatments regarding follicular viability, stromal cell density and detection of γH2AX and 53BP1. There was no difference in follicular morphology and DNA fragmentation between groups vitrified. In conclusion, catalase addition to vitrification solutions prevents ROS formation in cryopreserved goat ovarian tissues.

Relative mRNA expression and immunolocalization for transforming growth factor-beta (TGF-ß) and their effect on in vitro development of caprine preantral follicles.

This study aimed to evaluate the immunolocalization and messenger RNA (mRNA) expression for transforming growth factor-beta (TGF-ß) and its receptors (TGF-ßRI and RII), as well as mRNA expression for P450 aromatase and FSH receptor in caprine preantral follicles. The effects of TGF-ß, FSH alone, or in association on the in vitro follicular development were also assessed. Immunohistochemical analyses showed the expression of TGF-ß and its receptors in oocytes of all follicle stages and granulosa cells of primary and secondary follicles. mRNA for TGF-ß receptors and for FSH receptor (FSHR) was present in preantral follicles as well as in oocytes and granulosa cells of antral follicles. Isolated secondary follicles were cultured in α-minimum essential medium (MEM) alone or supplemented with either FSH (100 ng/ml), TGF-ß (10 ng/ml), or TGF-ß + FSH for 18 d. TGF-ß increased significantly oocyte diameter when compared to FSH alone and control. After 18 d of culture, all groups showed a significant reduction in P450 aromatase and FSHR mRNA levels in comparison to fresh control. In contrast, treatment with FSH significantly increased the mRNA expression for TGF-ß in comparison to fresh control and other treatments. In conclusion, the findings showed that TGF-ß and its receptors are present in caprine ovarian follicles. Furthermore, they showed a positive effect on oocyte growth in vitro.

Parameters of the reproductive tract, spermatogenesis, daily sperm production and major seminal plasma proteins of tropically adapted morada nova rams.

This study describes the reproductive parameters of Morada Nova rams, a breed of hair sheep from Brazil and with unique adaption to tropical environments. At 42 weeks of age, 15 rams were subjected to semen collection and, 1 week later, animals were slaughtered for collection of testes, epididymis and accessory sex glands. We conducted 2-D electrophoresis of seminal plasma proteins and major spots of stained gels were identified by LC-MS/MS. Total RNA was isolated from testis, epididymis and vesicular glands and subjected to qPCR. At slaughter, scrotal circumference and testicular weight were 27.5 ± 0.5 cm and 109.5 ± 6.0 g, respectively. Seminiferous tubule (ST) diameter was 188.3 ± 4.0 µm and each testis contained 1.9 ± 0.1 Sertoli cells (×10(9) ). Each Sertoli cell supported 0.1 ± 0.01 A spermatogonia, 3.0 ± 0.2 pachytene spermatocytes and 7.7 ± 0.5 round spermatids/tubule cross section. Daily sperm production reached 5.6 × 10(6)  cells/g of testis parenchyma. Testis size appeared as indicative of ST diameter and associated with epididymal measurements, as well as with the population of round spermatids and Sertoli cells/testis. Rams with heavier testes had greater daily sperm production and more Sertoli cells/testis. We detected 90.9 ± 9.6 spots per 2-D gel of seminal plasma. Major seminal proteins were identified as ram seminal vesicle proteins at 14 and 22 kDa, representing 16.2% and 12.8% of the total intensity of valid spots in the gels, respectively. Expression of both genes was greater in the vesicular glands as compared to testis and epididymis. Pixel intensity for those proteins in the 2-D gels was significantly correlated with seminal vesicle weight. This is the first description of the basic reproductive aspects of Morada Nova rams, including protein profiles of their seminal plasma. These findings will allow a better understanding of their reproductive physiology.

Alginate hydrogel matrix stiffness influences the in vitro development of caprine preantral follicles.

This study examined caprine follicular development in different concentrations of alginate matrix to determine the optimal conditions for culture. Caprine preantral follicles were cultured in a two-dimensional system (control) or a three-dimensional encapsulated system in 0.25%, 0.5%, or 1% alginate (ALG 0.25, ALG 0.5, and ALG 1, respectively). A higher percentage of morphologically normal follicles developed in ALG 0.5 and ALG 1 than in ALG 0.25 or the control (P < 0.05). The rate of antrum formation, however, was higher in ALG 0.25 than in ALG 0.5 and ALG 1 conditions (P < 0.05), but similar to the control. Follicles cultured in ALG 0.25 had higher growth rates and meiotic resumption than those cultured in ALG 0.5, ALG 1, or the control (P < 0.05). Moreover, follicles cultured in ALG 0.25 had higher levels of estradiol and progesterone than those cultured in ALG 0.5, ALG 1, or the control, as well as higher levels of CYP19A1 and HSD3B mRNA. In conclusion, a three-dimensional system that uses ALG 0.25 fosters the in vitro development of caprine preantral follicles and increases the rate of meiotic resumption.

Structure, function, and localization of aquaporins: their possible implications on gamete cryopreservation.

The discovery of water channels (aquaporins, AQPs) was a landmark event for the clarification of water transport through the plasma membrane. AQPs belong to a family of intrinsic membrane proteins that act as selective channels for water and for solutes such as glycerol and urea. AQPs were found in different tissues and organs, including male and female reproductive systems. In the swine female reproductive system, the AQPs were localized in the uterus, oviduct, and ovary, as well as in the granulosa cells from primordial follicles. Knowing the involvement of AQPs with the male and female germ cells, as well as their acknowledged role in transporting water through the plasma membrane, the research of these proteins in cryopreservation processes becomes essential. Thus, this review aims to describe the structure and function of AQPs in membranes, highlighting their role in the reproductive system (male and female). We also discuss the involvement of AQPs in cryopreservation, focusing on the effect and importance of these proteins on the rates of vitrification protocols for preantral follicles present in the ovarian tissue of domestic mammals.

The effects of epidermal growth factor (EGF) on the in vitro development of isolated goat secondary follicles and the relative mRNA expression of EGF, EGF-R, FSH-R and P450 aromatase in cultured follicles.

The effects of varying concentrations of EGF were evaluated in terms of in vitro follicular development and the mRNA expression levels of EGF, EGF-R, FSH-R and P450 aromatase. After 6 days, the addition of 50 ng/mL of EGF to the culture medium increased the antrum formation rates in comparison to cultured control and after 18 days of culture produced oocytes with higher rates of meiosis resumption when compared to the other treatments (P<0.05). The daily follicular growth rates in presence of EGF (50 or 100) were increased in comparison to the cultured control (P<0.05). Treatment with EGF 50 stimulated the expression of EGF mRNA but reduced EGF-R mRNA expression and estradiol secretion as compared to the cultured control (P<0.05). After 18 days of culture, the mRNA levels for FSH-R and P450 aromatase were greater than those of the non-cultured controls (P<0.05). In conclusion, the effects of EGF treatment on the mRNA levels for EGF, EGF-R, FSH-R, and P450 aromatase varied according to the stage of follicle development.

Caste development and reproduction: a genome-wide analysis of hallmarks of insect eusociality.

The honey bee queen and worker castes are a model system for developmental plasticity. We used established expressed sequence tag information for a Gene Ontology based annotation of genes that are differentially expressed during caste development. Metabolic regulation emerged as a major theme, with a caste-specific difference in the expression of oxidoreductases vs. hydrolases. Motif searches in upstream regions revealed group-specific motifs, providing an entry point to cis-regulatory network studies on caste genes. For genes putatively involved in reproduction, meiosis-associated factors came out as highly conserved, whereas some determinants of embryonic axes either do not have clear orthologs (bag of marbles, gurken, torso), or appear to be lacking (trunk) in the bee genome. Our results are the outcome of a first genome-based initiative to provide an annotated framework for trends in gene regulation during female caste differentiation (representing developmental plasticity) and reproduction.

The effect of alternative economic and genetic covariation structures on the relative economic gain from selection using stayability traits.

Three alternative cost ratios and the ratio of annual fixed cost for cows to net replacement cost for three populations that differed in genetic (co)variances were investigated. We studied their effect on the value of longevity records to supplement production records to achieve genetic gains for reducing the nonfeed cost of producing a unit of milk. Data files were constructed from stochastically generated production records and culling variant records; the later records represented the relative value for cow survival of all nonproduction traits. The use of one stayability trait instead of none increased gain by 1 to 9% across the populations and cost ratios. Only 2% additional gain occurred with two stayability traits instead of one. Populations that differed in genetic correlation between production and the survival value of nonproduction traits and in survival heritabilities contributed 2 to 5% toward the greater gains. The different cost ratios contributed 1 to 3%. The effect of increasing the length of herd life independent of production is inherently secondary to the effect of increasing the production during lactation. If genetic antagonism between production and the survival of nonproduction traits is found and if the net replacement costs are high, the use of production and longevity records for selection can enhance gain by up to 10%.