Abstract In the current study, we aimed to assess the expression levels of two circulating microRNAs (miRNA) (oar-miR-485-5p and oar-miR-493-5p) in the ovine plasma during the peri-implantation. After mating, we collected the plasma samples from a total of 8 ewes on day 22 of pregnancy (P22; n = 4) and day 22 of the estrous cycle (C22; n=4). We used mature miRNA sequences for oar-miR-485-5p and oar-miR-493-5p out of one hundred fifty, which were retrieved from our microarray results of previous study. We showed that the miRNA expression of oar-miR-485-5p and oar-miR-493-5p were upregulated in P22 (P<0.05) when compared to C22. Those two miRNAs targeted 311 target genes in the peri-implantation period of pregnancy. Furthermore, we revealed 151 GO/pathway terms in biological process (BP) and 25 GO/pathway terms in molecular function (MF), while we demonstrated 13 GO/pathway terms in cellular component (CC). We revealed three hub genes as interleukin 2 (IL2), interleukin 18 (IL18), and C-X-C Motif Chemokine Ligand 10 (CXCL10). In conclusion, both miR-485-5p and oar-miR-493-5p have the potential to be a biomarker to understand peri-implantation of the ovine pregnancy in the aspect of pregnancy-reflected changes in maternal plasma. study (miRNA oarmiR4855p oarmiRp oar miR 485 5p p oarmiR4935p 493 periimplantation. periimplantation peri implantation. implantation mating P (P22 4 C22 C (C22 n=4. n4 n=4 . n=4) fifty P2 P<0.05 P005 0 05 (P<0.05 31 Furthermore 15 GOpathway GO pathway BP (BP MF, MF , (MF) 1 CC. CC (CC) IL2, IL2 IL (IL2) IL18, IL18 (IL18) CXC X CXCL10. CXCL10 CXCL (CXCL10) conclusion miR4855p miRp pregnancyreflected reflected oarmiR 48 49 (P2 C2 (C2 n= P<0.0 P00 (P<0.0 3 (MF (CC (IL2 IL1 (IL18 CXCL1 (CXCL10 (P (C P<0. P0 (P<0. (IL (IL1 (CXCL1 P<0 (P<0 (CXCL P< (P<

Abstract In vitro cell culture is a well-established technique present in numerous laboratories in diverse areas. In reproduction, gametes, embryos, and reproductive tissues, such as the ovary and endometrium, can be cultured. These cultures are essential for embryo development studies, understanding signaling pathways, developing drugs for reproductive diseases, and in vitro embryo production (IVP). Although many culture systems are successful, they still have limitations to overcome. Three-dimensional (3D) culture systems can be close to physiological conditions, allowing greater interaction between cells and cells with the surrounding environment, maintenance of the cells' natural morphology, and expression of genes and proteins such as in vivo. Additionally, three-dimensional culture systems can stimulated extracellular matrix generating responses due to the mechanical force produced. Different techniques can be used to perform 3D culture systems, such as hydrogel matrix, hanging drop, low attachment surface, scaffold, levitation, liquid marble, and 3D printing. These systems demonstrate satisfactory results in follicle culture, allowing the culture from the pre-antral to antral phase, maintaining the follicular morphology, and increasing the development rates of embryos. Here, we review some of the different techniques of 3D culture systems and their applications to the culture of follicles and embryos, bringing new possibilities to the future of assisted reproduction. wellestablished well established areas reproduction gametes embryos tissues endometrium cultured studies pathways diseases IVP. IVP . (IVP) successful overcome Threedimensional Three dimensional D (3D conditions environment morphology vivo Additionally threedimensional three produced drop surface scaffold levitation marble printing preantral pre phase Here (IVP

Abstract We hypothesized that the hCG modulates the expression of IFNT-pathway and ISGs in bovine endometrium during early pregnancy. The aim of the current study is to evaluate the effect of hCG on IFNT-pathway signals and ISGs expression in endometrial cells. For this, 29 non-lactating cross-bread cows were used in the study and submitted to a 9-day fixed-time artificial insemination (FTAI) protocol. The day of the AI was considered Day 0 (D0), and five days (D5) after the FTAI, the cows were allocated into two groups: Control and hCG group, when a hCG group received a single dose of 2.500UI of hCG. On day 18 after FTAI (D18) cows were slaughtered and endometrial tissue samples were collected. There was no difference between the embryo recovery rate of the cows in C compared to the hCG. The hCG group increased the accessory corpus luteum formation rate. The hCG resulted in greater serum progesterone concentration in the hCG group compared to the C on Day 14. Only the expression of IFNAR2 and STAT1 were upregulated on pregnant cows of the hCG group compared to the C group. The pathway genes (JAK1, STAT2, and IRF9) were not regulated. The mRNA abundance of ISG15, MX1, MX2, and OAS1 was upregulated in pregnant cows for hCG group, compared to C group. The results show that the administration of hCG, 5 days after AI, in addition to increasing the serum progesterone, modulates the expression of IFNT-pathway and ISGs on bovine endometrium on Day 18 of pregnancy. IFNTpathway IFNT pregnancy cells this 2 nonlactating non lactating crossbread cross bread 9day 9 fixedtime fixed time (FTAI protocol D0, D0 D , (D0) D5 (D5 groups 2500UI UI 500UI 1 D18 (D18 collected 14 IFNAR STAT JAK1, JAK1 JAK (JAK1 STAT2 IRF9 IRF regulated ISG15 ISG MX1 MX MX2 OAS (D0 (D D1 (D1 (JAK ISG1

Abstract One of the crucial aspects to be considered for successful in vitro production (IVP) of embryos is the composition of the various media used throughout the stages of this reproductive biotechnology. The cell culture media employed should fulfill the metabolic requirements of both gametes during oocyte maturation and sperm development, as well as the embryo during its initial cell divisions. Most IVP protocols incorporate blood serum into the media composition as a source of hormones, proteins, growth factors, and nutrients. Numerous studies have suggested Platelet-Rich Plasma (PRP) as a substitute for fetal sera in cell culture, particularly for stem cells. Therefore, the objective of this study is to assess the potential use of PRP as a replacement for fetal bovine serum (FBS) during oocyte maturation for in vitro production of bovine embryos. During in vitro maturation (IVM), cumulus-oocyte complexes (COCs) were allocated into the following experimental groups: Group G1 (IVM medium with 5% PRP); Group G2 (MIV medium with 5% PRP and 5% SFB); Group G3 (MIV medium with 5% SFB); and Group G4 (MIV medium without either PRP or SFB). Subsequently, the cumulus-oocyte complexes were fertilized with semen from a single bull, and the resulting zygotes were cultured for seven days. Cleavage and blastocyst formation rates were assessed on days 2 and 7 of embryonic development, respectively. The quality of matured COCs was also evaluated by analyzing the gene expression of HSP70, an important protein associated with cellular stress. The results demonstrated that there were no significant differences among the experimental groups in terms of embryo production rates, both in the initial cleavage stages and blastocyst formation (except for the G4 group, which exhibited a lower blastocyst formation rate on D7, as expected). This indicates that PRP could be a cost-effective alternative to SFB in the IVP of embryos. (IVP biotechnology development divisions hormones proteins factors nutrients PlateletRich Platelet Rich (PRP cells Therefore FBS (FBS IVM, IVM , (IVM) cumulusoocyte cumulus (COCs G 5 PRP) MIV SFB) SFB. . Subsequently bull respectively HSP70 HSP stress except group D7 D expected. expected expected) costeffective cost effective HSP7

Abstract Since the 1970s, maternal corticosteroid therapy has been used successfully to induce labor. This allows for better monitoring of parturients and provision of first aid to neonates, improving neonatal viability, as this treatment induces maturation in a variety of fetal tissues, thereby reducing morbidity and mortality. Although the effects of corticosteroids are well known, few studies have investigated the influence of this therapy in Santa Inês sheep. This study aimed to evaluate the efficacy of dexamethasone at two doses (8 and 16 mg) to induce lambing in Santa Inês ewes at 145 days of gestation and assess its effects on neonatal vitality. For this study, 58 ewes raised in an extensive system were investigated. Pregnancy was confirmed after artificial insemination at a set time or after controlled mounting. Ewes were separated into three groups: an untreated control group (G1: 0 mg) and groups treated with two doses of dexamethasone (G2: 8 mg and G3: 16 mg). In total, 79 lambs were born. Their vitality was assessed based on their Apgar score, weight, temperature, and postnatal behavior. SAS v9.1.3 (SAS Institute, Cary, NC) was used to analyze data, considering a 5% significance level for all analyses. The births in the induced groups occurred 48.4 ± 22.1 h after induction, while the ewes that underwent non-induced labor gave birth 131.96 ± 41.9 h after placebo application (p < 0.05), confirming the efficacy of dexamethasone to induce and synchronize labor. The induced and non-induced neonates had similar Apgar scores, temperatures, weights, and postnatal behavioral parameters (p > 0.05). This study showed that inducing labor in Santa Inês ewes at 145 days of gestation with a full (16 mg) or half dose (8 mg) of dexamethasone is an effective technique and does not compromise neonate vitality. 1970s s viability tissues mortality known sheep ( 1 14 5 mounting G1 G (G1 G2 (G2 G3 mg. . total 7 born score weight temperature behavior v913 v v9 3 v9.1. Institute Cary NC data analyses 484 48 4 48. 221 22 22. induction noninduced non 13196 131 96 131.9 419 41 9 41. p 0.05, 005 0.05 , 05 0.05) scores temperatures weights 0.05. (1 (G v91 v9.1 2 1319 13 131. 00 0.0 v9. 0.

Abstract The subcortical maternal complex, which consists of maternal-effect genes, plays a crucial role in the development of oocytes and preimplantation embryo until the activation of the zygote genome. One such gene, known as peptidyl-arginine deiminase VI (Padi6), is involved in the oocyte maturation, fertilization and embryonic development. However, the precise function of Padi6 gene in buffalo is still unclear and requires further investigation. In this study, the sequence, mRNA and protein expression patterns of Padi6 gene were analyzed in oocytes, preimplantation embryos and somatic tissues of buffalo. The coding sequence of gene was successfully cloned and characterized. Real-time quantitative PCR results indicated an absence of Padi6 transcripts in somatic tissues. Notably, the expression levels of Padi6 in oocytes showed an increased from the germinal vesicle stage to metaphase II stage, followed by a rapid decrease during the morula and blastocyst stages. Immunofluorescence analysis confirmed these findings, revealing a noticeable decline in protein expression levels. Our research provides the initial comprehensive expression profile of Padi6 in buffalo oocytes and preimplantation embryos, serving as a solid foundation for further investigations into the functionality of maternal-effect genes in buffalo. complex maternaleffect effect genome peptidylarginine peptidyl arginine Padi6, Padi , (Padi6) maturation However investigation study characterized Realtime Real time Notably stages findings (Padi6 (Padi

Abstract This study aimed to analyze the characteristics of the HSP70 gene and protein in spermatozoa of Bali bulls of different age groups and to examine its potential as a biomarker determining bull fertility. This study used frozen semen produced from six Bali bulls divided into two groups based on age (≤ 9 years and ≥ 12 years). Parameters of frozen semen quality analyzed included sperm motility and kinetics using computer-assisted semen analysis, sperm morphological defects using Diff-Quick staining, acrosome integrity using FITC-PNA staining, and DNA fragmentation using acridine orange staining. HSP70 gene expression characterization was analyzed using qRT-PCR, and HSP70 protein abundance was analyzed using enzyme immunoassays. Fertility field data were obtained by analyzing the percentage conception rate for each bull based on the artificial insemination service data contained in the Indonesian-integrated system of the National Animal Health Information System (iSIKHNAS). The results showed significant differences (P<0.05) in total and progressive motility, morphological defects of the neck and midpiece, and tail of sperm, and acrosome integrity between the age groups of Bali bulls. HSP70 gene expression and protein abundance showed no significant differences (P>0.05) in different age groups. HSP70 gene expression correlated with fertility rate (P<0.05). Age affected several semen quality parameters but did not affect HSP70 gene expression and protein abundance. The HSP70 gene molecule could be a biomarker that determines the fertility of Bali bulls. HSP HSP7 ≤ ( 1 years. . years) computerassisted computer assisted analysis DiffQuick Diff Quick staining FITCPNA FITC PNA qRTPCR, qRTPCR qRT PCR, PCR qRT-PCR immunoassays Indonesianintegrated Indonesian integrated iSIKHNAS. iSIKHNAS (iSIKHNAS) P<0.05 P005 P 0 05 (P<0.05 midpiece P>0.05 (P>0.05 P<0.05. (iSIKHNAS P<0.0 P00 (P<0.0 P>0.0 (P>0.0 P<0. P0 (P<0. P>0. (P>0. P<0 (P<0 P>0 (P>0 P< (P< P> (P> (P

Abstract The objective of this study was to evaluate the effect of three doses of equine chorionic gonadotropin (eCG) for ovarian superstimulation on ovarian response, follicular development and cumulus-oocyte complexes (COCs) collection in llamas. For this purpose, eighteen multiparous non-lactating adult (4–7 yo) female llamas with an average body condition of 2.8 (BCS 1–5) were submitted to a follicular ablation (FA) to induce a new follicular wave emergence. Two days after FA (Day 0), synchronized llamas were randomly allocated to three treatment groups (n = 6/group) and given 500, 750 and 1000 IU of eCG (Novormon®, Syntex, Buenos Aires, Argentina) per animal respectively to induce ovarian superstimulation. Transrectal ultrasonography were performed on Days 2, 4, and 6; and ovum pick up (OPU) was performed on Day 6. Data was evaluated by one-way analysis of variance (ANOVA), repeated measures ANOVA, and 2-tailed Chi-square. The average size (mm) of follicles was greater (p≤ 0.05) in the 1000 IU group compared to the other groups. There was a greater (p≤ 0.05) number of follicles ≥ 7 mm in the 1000 IU group compared to the 500 IU group. Number of COCs collected on Day 6 and the COC recovery rate were not different among groups. In conclusion, a single dose of 1000 IU of eCG induced the best ovarian response resulting in larger and greater number of follicles at the time of OPU. (eCG cumulusoocyte cumulus oocyte (COCs purpose nonlactating non lactating 4–7 47 4 (4– yo 28 2 8 2. BCS 1–5 15 1 5 (FA emergence 0, 0 , 0) n 6/group 6group 75 100 Novormon®, Novormon (Novormon® Syntex Aires Argentina OPU (OPU oneway one way ANOVA (ANOVA) 2tailed tailed Chisquare. Chisquare Chi square. square Chi-square (mm p≤ p (p 0.05 005 05 50 conclusion 4– (4 1– 10 Novormon® (Novormon (ANOVA 0.0 00 ( 0.

Abstract This study aims to investigate the gene expression of sperm-borne phospholipase C zeta (PLCζ), WW domain-binding protein 2N-Terminal Like (WBP2NL), and Tumor necrosis factor (TNF-α), as a negative control, in spermatozoa and their relationship with fertility and seminal quality in stallions. Ejaculates from 40 Criollo stallions were used, whose fertility was assessed on the basis of their pregnancy rate per cycle in at least two breeding seasons. Pregnancy rates ranged from 20% to 90% and were used to divide the stallions into two groups: High rates (≥ 50%) (n = 25), and Low rates (< 50%) (n = 15). A computer-assisted sperm analysis system - (CASA) analyzed semen after collection. Also were evaluated the physical and functional integrity of the plasmatic membrane and sperm morphology alterations. All stallions expressed PLCζ, WBP2NL, and TNF-α. PLCζ positively correlates with conception rate, total motility (TM), progressive motility (PM), plasmatic membrane functionality, and integrity. A simple linear regression was detected between pregnancy rate and PLCζ expression (P = 0.003), TM (P < 0.001) and PM (P < 0.001). PLCζ gene expression was higher (P = 0,012) in the High rates group than in the Low group. WBP2NL and TNF-α did not correlate with seminal quality and stallion’s fertility. It was concluded that PLCζ gene expression in the spermatozoa might be used as a biomarker of fertility and seminal quality in stallions. Parameters of sperm kinetics also showed, positive correlation between TM, PM and pregnancy rate. spermborne borne , (PLCζ) domainbinding domain binding 2NTerminal NTerminal 2N Terminal N WBPNL WBP NL (WBP2NL) TNFα, TNFα TNF α (TNF-α) control 4 seasons 20 90 groups ≥ ( 50% 50 n 25, 25 25) 15. 15 . 15) computerassisted computer assisted CASA (CASA collection alterations TNFα. α. (TM) PM, (PM) functionality P 0.003, 0003 0.003 0 003 0.003) 0.001 0001 001 0.001. 0,012 0012 012 stallion s showed (PLCζ (WBP2NL (TNF-α 2 9 5 1 (TM (PM 000 0.00 00 0,01 01 0.0 0,0 0. 0,

Abstract The CRISPR/Cas9 system is a simpler and more versatile method compared to other engineered nucleases such as Zinc Finger Nucleases (ZFNs) and Transcription Activator-Like Effector Nucleases (TALENs), and since its discovery, the efficiency of CRISPR-based genome editing has increased to the point that multiple and different types of edits can be made simultaneously. These advances in gene editing have revolutionized biotechnology by enabling precise genome editing with greater simplicity and efficacy than ever before. This tool has been successfully applied to a wide range of animal species, including cattle, pigs, dogs, and other small animals. Engineered nucleases cut the genome at specific target positions, triggering the cell's mechanisms to repair the damage and introduce a mutation to a specific genomic site. This review discusses novel genome-based CRISPR/Cas9 editing tools, methods developed to improve efficiency and specificity, the use of gene-editing on animal models and translational medicine, and the main challenges and limitations of CRISPR-based gene-editing approaches. CRISPRCas9 CRISPRCas CRISPR Cas9 Cas CRISPR/Cas ZFNs (ZFNs ActivatorLike Activator Like TALENs, TALENs , (TALENs) discovery CRISPRbased based simultaneously before species cattle pigs dogs animals positions cells cell s site genomebased tools specificity geneediting medicine approaches (TALENs

Abstract The adnexa fetal tissues are sources of mesenchymal stromal cells (MSCs) due to their noninvasive harvest, with all biological material discarded most of the time. MSCs are a promise regarding to their plasticity, self-renewal, differentiation potentials, immunomodulatory and anti-inflammatory properties, which have made clinical stem cell therapy a reality. The present study aimed to characterize and evaluate the immunomodulation ability of bovine mesenchymal cells collected from bovine amniotic fluid (bAFMSCs) isolated and subjected to sixth consecutive culture passages in vitro. The multilineage properties of the bAFMSCs collections confirmed the ability to undergo adipogenic, chondrogenic and osteogenic differentiation. The mesenchymal gene transcription CD106, CD73, CD29, CD90 and CD166 were detected in bAFMSCs, whereas CD34 and CD45 were not detected. Regarding cytokine mRNA expression, IL2, IL6, INFα, INFβ, INFγ, TNFα and TNFβ were downregulated, while IL10 was highly regulated in all studied passages. The present study demonstrated the immunological properties and multipotency of in vitro bAFMSCs collections, and thus, they can be tested in cattle pathological treatments or multiplication by nuclear transfer cloning. (MSCs harvest time plasticity selfrenewal, selfrenewal self renewal, renewal self-renewal potentials antiinflammatory anti inflammatory reality (bAFMSCs adipogenic CD106 CD CD73 CD29 CD9 CD16 CD3 CD4 expression IL2 IL IL6 INFα INFβ INFγ downregulated IL1 thus cloning CD10 CD7 CD2 CD1

Abstract Steroids and gonadotrophins are essential for the regulation of late stages of preantral development and antral follicular development. Although the luteinizing hormone receptor (LHCGR) has been detected in the preantral follicles of rats, rabbits, and pigs, its expression, in bovine fetal ovary, has not been demonstrated. Based on this, we aimed to investigate the expression of the LHCGR and LHCGR mRNA binding protein (LRBP), as well as, to quantify bta-miR-222 (a regulatory microRNA of the LHCGR gene) during the development of bovine fetal ovary. In summary, LHCGR expression was observed in the preantral follicle in bovine fetal ovary, from oogonias to primordial, primary and secondary stages, and the mRNA abundance was lower on day 150 than day 60. However, the mRNA abundance of LRBP followed the opposite pattern. Similar to LRBP, the abundance of bta-miR-222 was higher on day 150 than day 60 or 90 of gestation. The LHCGR protein was detected in oogonia, primordial, primary, and secondary follicles. Moreover, both oocytes and granulosa cells showed positive immunostaining for LHCGR. In conclusion, we suggest the involvement of LHCGR/LRBP/bta-mir222 with mechanisms related to the development of preantral follicles in cattle. (LHCGR rats rabbits pigs ovary demonstrated this , (LRBP) btamiR222 btamiR bta miR 222 bta-miR-22 a gene summary primordial 15 However pattern 6 9 gestation oogonia Moreover conclusion LHCGR/LRBP/btamir222 LHCGRLRBPbtamir222 LHCGRLRBPbtamir LHCGR/LRBP/bta mir222 mir LHCGR/LRBP/bta-mir22 cattle (LRBP btamiR22 22 bta-miR-2 1 btamir222 btamir LHCGR/LRBP/btamir22 LHCGRLRBPbtamir22 LHCGRLRBPbta mir22 LHCGR/LRBP/bta-mir2 btamiR2 2 bta-miR- btamir22 LHCGR/LRBP/btamir2 LHCGRLRBPbtamir2 mir2 LHCGR/LRBP/bta-mir bta-miR btamir2 LHCGR/LRBP/btamir

Abstract The reproductive efficiency of the herd is correlated with higher productivity in livestock. Reproduction biotechniques, such as ovulation synchronization protocols, are important to optimize production and accelerate genetic profit in beef and dairy herds. The objective of this review is to describe the evolution over the last 40 years of the artificial insemination (AI) and the timed-AI (TAI) protocols in cattle from a Brazilian perspective. TAI protocols are based on synchronizing emergence of the wave of follicular growth, controlling circulating progesterone (P4) concentrations, stimulating the final growth of the follicle and inducing a synchronized ovulation. Hormonal alternatives that optimize the response at the end of the protocol and strategies to induce final follicle growth and ovulation in categories of females with low expression of estrus are described. Furthermore, the potential positive effect of previous exposure to injectable P4 on fertility of Bos indicus and Bos taurus cows is also discussed. livestock biotechniques herds 4 AI (AI timedAI timed (TAI perspective P (P4 concentrations described Furthermore discussed (P

Abstract Assisted reproductive technologies (ART) are fundamental for cattle breeding and sustainable food production. Together with genomic selection, these technologies contribute to reducing the generation interval and accelerating genetic progress. In this paper, we discuss advancements in technologies used in the fertility evaluation of breeding animals, and the collection, processing, and preservation of the gametes. It is of utmost importance for the breeding industry to select dams and sires of the next generation as young as possible, as is the efficient and timely collection of gametes. There is a need for reliable and easily applicable methods to evaluate sexual maturity and fertility. Although gametes processing and preservation have been improved in recent decades, challenges are still encountered. The targeted use of sexed semen and beef semen has obliterated the production of surplus replacement heifers and bull calves from dairy breeds, markedly improving animal welfare and ethical considerations in production practices. Parallel with new technologies, many well-established technologies remain relevant, although with evolving applications. In vitro production (IVP) has become the predominant method of embryo production. Although fundamental improvements in IVP procedures have been established, the quality of IVP embryos remains inferior to their in vivo counterparts. Improvements to facilitate oocyte maturation and development of new culture systems, e.g. microfluidics, are presented in this paper. New non-invasive and objective tools are needed to select embryos for transfer. Cryopreservation of semen and embryos plays a pivotal role in the distribution of genetics, and we discuss the challenges and opportunities in this field. Finally, machine learning (ML) is gaining ground in agriculture and ART. This paper delves into the utilization of emerging technologies in ART, along with the current status, key challenges, and future prospects of ML in both research and practical applications within ART. ART (ART selection progress animals possible decades encountered breeds practices wellestablished well established relevant (IVP counterparts systems eg e g e.g microfluidics noninvasive non invasive transfer genetics field Finally (ML status

Abstract Reproductive maturation is a complex physiological process controlled by the neuroendocrine system and is characterized by an increase in gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) pulsatile secretion. Nutrition during early development is a key factor regulating puberty onset, which is defined as first ovulation in females. In heifers, nutrient restriction after weaning delays puberty, whereas elevated levels of nutrition and energy reserves advance reproductive maturation. Recent studies in cattle and other animal models have shown that the dam’s nutrition during gestation can also program the neuroendocrine system in the developing fetus and has the potential to alter timing of puberty in the offspring. Among the metabolic signals that modulate brain development and control timing of puberty is leptin, a hormone produced primarily by adipocytes that communicates energy status to the brain. Leptin acts within the arcuate nucleus of the hypothalamus to regulate GnRH secretion via an upstream network of neurons that includes neurons that express neuropeptide Y (NPY), an orexigenic peptide with inhibitory effects on GnRH secretion, and alpha melanocyte-stimulating hormone (αMSH), an anorexigenic peptide with excitatory effects on GnRH neurons. Another important population of neurons are KNDy neurons, neurons in the arcuate nucleus that co-express the neuropeptides kisspeptin, neurokinin B, and dynorphin and have strong stimulatory effects on GnRH secretion. Our studies in beef heifers indicate that increased nutrition between 4 to 8 months of age advances puberty by diminishing NPY inhibitory tone and by increasing excitatory inputs of αMSH and kisspeptin, which collectively lead to increased GnRH/LH pulsatility. Our ongoing studies indicate that different planes of nutrition during gestation can alter maternal leptin concentrations and promote changes in the fetal brain. Nonetheless, at least in Bos indicus-influenced heifers, deficits programmed prenatally can be overcome by adequate postnatal nutrition without negatively impacting age at puberty or subsequent fertility. gonadotropinreleasing gonadotropin releasing (GnRH LH (LH onset females dams dam s offspring NPY, , (NPY) melanocytestimulating melanocyte stimulating αMSH, (αMSH) coexpress co kisspeptin B GnRHLH pulsatility Nonetheless indicusinfluenced indicus influenced fertility (NPY (αMSH