Equitable infrastructure: Achieving resilient systems and restorative justice through policy and research innovation.

Recent major investments in infrastructure in the United States and globally present a crucial opportunity to embed equity within the heart of resilient infrastructure decision-making. Yet there is a notable absence of frameworks within the engineering and scientific fields for integrating equity into planning, design, and maintenance of infrastructure. Additionally, whole-of-government approaches to infrastructure, including the Justice40 Initiative, mimic elements of process management that support exploitative rather than exploratory innovation. These and other policies risk creating innovation traps that limit analytical and engineering advances necessary to prioritize equity in decision-making, identification and disruption of mechanisms that cause or contribute to inequities, and remediation of historic harms. Here, we propose a three-tiered framework toward equitable and resilient infrastructure through restorative justice, incremental policy innovation, and exploratory research innovation. This framework aims to ensure equitable access and benefits of infrastructure, minimize risk disparities, and embrace restorative justice to repair historical and systemic inequities. We outline incremental policy innovation and exploratory research action items to address and mitigate risk disparities, emphasizing the need for community-engaged research and the development of equity metrics. Among other action items, we recommend a certification system-referred to as Social, Environmental, and Economic Development (SEED)-to train infrastructure engineers and planners and ensure attentiveness to gaps that exist within and dynamically interact across each tier of the proposed framework. Through the framework and proposed actions, we advocate for a transformative vision for equitable infrastructure that emphasizes the interconnectedness of social, environmental, and technical dimensions in infrastructure planning, design, and maintenance.

Effect of maternal age on ATP content and distribution of mitochondria in bovine oocytes.

Our objective was to understand how maternal age influences the mitochondrial population and ATP content of in vivo matured bovine oocytes. We hypothesized that in vivo matured oocytes from older cows would have altered mitochondrial number and distribution patterns and lower cytoplasmic ATP content compared to the oocytes obtained from younger cows. Follicles ≥5mm were ablated in old cows (13 to 22 yrs, Old Group, n = 7) and their younger daughters (4 to 10 years old, Young Group; n = 7) to induce the emergence of a new follicular wave. Cows were treated twice daily with eight doses of FSH starting 24 hr after ablation (Day 0, day of wave emergence). Prostaglandin F2alpha (PGF) was given on Days 3 and 3.5, LH on Day 4.5, and cumulus-oocyte-complexes were collected 18-20 hours post-LH by ultrasound-guided follicular aspiration. Oocytes were either processed for staining with MitoTracker Deep Red FM or for ATP assay. Stained oocytes were imaged with a Zeiss LSM 710 confocal microscope, and mitochondria were segmented in the oocyte volume sets using Imaris Pro 7.4. In vivo matured oocytes obtained from old cows were similar in morphological grades to those from young cows. However, the oocytes of COC from older cows had 23% less intracellular ATP (27.4±1.9 vs 35.7±2.2 pmol per oocyte, P = 0.01) than those of young cows. Furthermore, the average volume of individual mitochondria, indicated by the number of image voxels, was greater (P<0.05) in oocytes from older cows than in those from younger cows. Oocytes from older cows also tended to have a greater number of mitochondrial clusters (P = 0.06) and an increased number of clusters in the central region of the oocytes (P = 0.04) compared to those from younger cows. In conclusion, our study demonstrated that maternal age was associated with a decrease in the cytoplasmic ATP content of in vivo mature oocytes and an altered distribution of mitochondrial structures. These findings suggest that maternal age may negatively influence the developmental competence of oocytes from older cows.

An efficient synthesis and characterization of La@MOF-808: A promising strategy for effective arsenic ion removal from water.

Addressing serious waterborne arsenic issues, for the first time, lanthanum-doped MOF-808 (La@MOF-808) has been developed to remove total arsenic (Total As) and arsenite [As(III)] from water. This study involves the solvothermal synthesis of La@MOF-808, its characterization via FTIR, XRD, TGA, and SEM, in which distinct physicochemical attributes were identified, and the adsorption capacity of arsenic ions. The saturated adsorption capacity of La@MOF-808 for Total As and As(III) reached 282.9 mg g-1 and 283.5 mg g-1, as compared to 229.7 mg g-1 and 239.1 mg g-1 for pristine MOF-808, respectively. XRD and ATR-FTIR analyses underscored the central roles of electrostatic interactions and hydroxyl groups in the pollutant adsorption process. The impact of temperature, concentration, pH, and exposure duration times on adsorption performance was thoroughly investigated. The Langmuir model showed the maximum adsorption capacities (qmax) of La@MOF-808 was 307.7 mg g-1 for Total As and 325.7 mg g-1 for As(III), surpassing those of MOF-808 adsorbent, which suggests that monolayer adsorption occurred. Optimal adsorption was observed in a pH range of 2.0-7.0, and thermodynamic studies classified the process as spontaneous and endothermic. The adsorbent retains high capacity across repeated cycles, outperforming many standard adsorbents. Lanthanum doping markedly enhances MOF-808's arsenic removal, underscoring its potential for water treatment.

Building a Better Forecast: Reformulating the Ensemble Kalman Filter for Improved Applications to Volcano Deformation.

As the volume of data collected at monitored volcanoes continues to expand, researchers will need quick, reliable, and automated methods of inverting those data into useful models of the underlying magma systems. Recently adapted from other fields for use in volcanology, the Ensemble Kalman Filter (EnKF) is one such inversion technique that has been used to produce several successful forecasts and hind-casts of volcanic unrest, correlating geodetic deformation with mechanical stresses around the magma reservoir. However, given the similarity in which changes to a reservoir's size and pressure are expressed at the surface, the filter can have trouble fully resolving magmatic conditions. In this study, we therefore test several different published variations of the EnKF workflow to produce an optimal configuration for use in future forecasting efforts. By generating synthetic observations of ground deformation under known conditions and then assimilating them through different implementations of the EnKF, we find that many variants favored in other fields underperform for this specific application. We conclude that correlations between model parameters that develop within the EnKF's Monte Carlo ensemble distort the filter's ability to correctly update the model state, causing the filter to systematically favor changes in some parameters over others and ultimately converge to a partially inaccurate solution. This effect can be somewhat mitigated by interrupting these parameter correlations, and the filter remains sensitive to many aspects of the magma system regardless. However, further research and novel approaches will be needed to truly optimize the EnKF for use in volcanology.

Is seminal nerve growth factor-induced luteinizing hormone release in camelids mediated at the hypothalamus?

In brief: Seminal nerve growth factor induces ovulation in camelids by influencing the secretion of gonadotrophin-releasing hormone (GnRH) into the portal vessels of the pituitary gland. We show that the nerve growth factor-induced release of GnRH is not mediated directly through interaction with hypothalamic neurons. Abstract: Ovulation in camelids is triggered by seminal nerve growth factor (NGF). The mechanism of action of NGF appears to occur via the central nervous system. In this study, we tested the hypothesis that NGF acts in the hypothalamus to induce GnRH release. To determine if NGF-induced ovulation is associated with a rise in NGF concentrations in the cerebrospinal fluid (CSF), llamas were i) mated with an urethrostomized male, ii) mated with intact male, or given intrauterine iii) seminal plasma or i.v.) saline (Experiment 1). To characterize the luteinizing hormone (LH) response after central vs peripheral administration, llamas were treated with saline (negative control) or NGF either by i.v. or intracerebroventricular (ICV) administration (Experiment 2). To determine the role of kisspeptin, the effect of ICV infusion of a kisspeptin receptor antagonist on NGF-induced LH secretion and ovulation was tested in llamas (Experiment 3). In Experiment 1, a surge in circulating concentrations of LH was detected only in llamas mated with an intact male and those given intrauterine seminal plasma, but no changes in CSF concentrations of NGF were detected. In Experiment 2, peripheral administration (i.v.) of NGF induced an LH surge and ovulation, whereas no response was detected after central (ICV) administration. In Experiment 3, the kisspeptin receptor antagonist had no effect on the LH response to NGF. In conclusion, results did not support the hypothesis that NGF-induced ovulation is mediated via a trans-synaptic pathway within the hypothalamus, but rather through a releasing effect on tanycytes at the median eminence.

Granulosa cell tumour in a llama (Lama glama).

Hormonally active tumours are characterized by production and secretion of hormones, irrespective of endogenous feedback mechanisms. An adult llama had exuberant oestrous behaviour, infertility, elevated concentrations of oestradiol and a large ovarian mass. Necropsy revealed the presence of two large abdominal masses, one effacing the right ovary and one in the mesocolon. Considering the clinical and histopathological findings, we conclude that the llama was affected by a granulosa cell tumour. The case suggests that granulosa cell tumours in camelids are hormonally active, and the clinical presentation resembles that of other large animal species. To our knowledge, this is the first case report of an oestrogen-producing, metastatic granulosa cell tumour in a llama.

Mechanism of LH release after peripheral administration of kisspeptin in cattle.

Kisspeptin modulates GnRH secretion in mammals and peripheral administration of 10-amino acid fragment of kisspeptin (Kp10) induces LH release and ovulation in cattle. Experiments were done to determine if iv administration of kisspeptin will activate GnRH neurons (i.e., after crossing the blood-brain barrier) and if pre-treatment with a GnRH receptor blocker will alter kisspeptin-induced LH release (from gonadotrophs) and ovulation. In Experiment 1, cows (n = 3 per group) were given human-Kisspeptin10 (hKp10; 3 x 15 mg iv at 60-min intervals) or normal saline and euthanized 150 min after treatment was initiated. Every 20th free-floating section (50 µm thickness) from the preoptic area to hypothalamus was double immunostained to colocalize GnRH- (DAB) and activated neurons (cFOS; Nickel-DAB). Kisspeptin induced plasma LH release from 15 to 150 min (P = 0.01) but the proportion of activated GnRH neurons did not differ between groups (5.8% and 3.5%, respectively; P = 0.11). Immunogold electron microscopy detected close contacts between kisspeptin fibers and GnRH terminals in the median eminence. In Experiment 2, pubertal heifers (n = 5 per group) were treated with 1) hKp10 iv, 2) Cetrorelix (GnRH antagonist; im) + hKp10 iv or 3) saline on Day 6 of the follicular wave under low-progesterone condition. A rise in plasma LH concentration was detected from 15 to 240 min in the hKp10 group but not in cetrorelix or control group (P<0.001). Ovulations were detected only in the hKp10 group (4/5; P = 0.02). Cetrorelix treatment was associated with regression of the preovulatory dominant follicle and emergence of a new follicular wave 3.4±0.75 days after the treatment in all five heifers. Results support the hypothesis that the effect of peripheral kisspeptin is mediated downstream of GnRH synthesis and does not involve GnRH-independent LH release from gonadotrophs. Peripheral kisspeptin may release pre-synthesized GnRH from the nerve terminals in areas outside the blood-brain barrier.

Strategies for oocyte collection and in vitro embryo production in free-roaming bison herds.

The study was conducted to test the feasibility of protocols for field collection of cumulus-oocyte complexes (COC) for in vitro embryo production (IVP) in wild bison. The study was done with captive wood bison during the anovulatory season. In Experiment 1, the efficiency of transvaginal ultrasound-guided COC collection was compared between bison restrained in a squeeze chute without sedation vs in lateral recumbency after chemical immobilization using a dart gun (n = 8/group). In Experiment 2, a 2 × 2 design was used to examine the effects of superstimulation treatment [single dose of equine chorionic gonodotrophin (eCG) vs multiple doses of follicle stimulating hormone (FSH)] and method of drug administration (manual injection vs field darting) on COC collection and IVP. In Experiment 1, no difference was detected between chute-restrained vs chemically immobilized groups in the time required to complete COC collections, the number of follicles aspirated (11.5 ± 1.9 vs 9.3 ± 1.8; P = 0.4) or the COC recovery rate [COC recovered/follicle aspirated; 58/92 (63%) vs 44/69 (64%); P = 0.9]. In Experiment 2, no differences were detected between superstimulation treatments (eCG vs FSH). The total number of follicles available for aspiration did not differ between manual injection and field darting (23.9 ± 2.7 vs 21.6 ± 1.9; P = 0.4). Compared with the random start unstimulated group, the embryo production rate was higher [18/132 (14%) vs 53/189 (28%); P = 0.04] after wave synchronization and superstimulation. Results suggest that COC collection is equally feasible in a recumbent position after chemical immobilization as those bison restrained in a standing position in a hydraulic chute. Ovarian superstimulation with a single-dose eCG protocol is as effective as a multiple-dose FSH protocol, and field darting is as effective as chute-side administration of superstimulation treatments. The strategies in the present study are ready to be incorporated into field collections in free-roaming bison herds.

Influence of ovarian follicular wave synchronization and single-dose eCG superstimulation on oocyte collection and in vitro embryo production in bison during the ovulatory and anovulatory seasons.

In an effort to develop an effective, minimum-handling protocol for the conservation of wood bison, the present study was designed to determine the effects of ovarian synchronization and superstimulation on cumulus oocyte complex (COC) collection and in vitro embryo production in wood bison during the ovulatory (Exp. 1) and anovulatory seasons (Exp. 2). We tested the hypotheses that COC collection and in vitro embryo production are 1) greater after follicular wave synchronization than at random stages of the follicular wave, 2) repeatable within individuals, 3) greater after ovarian superstimulation with a single dose of eCG than without treatment, and 4) greater during the anovulatory season than the ovulatory season. In Exp. 1, ultrasound-guided COC collection was performed on Day -1 in wood bison to induce follicular wave emergence the following day (Day = 0). Immediately after the COC collection on Day -1, bison were given a single im dose of 2500 IU eCG or saline (n = 6 per group). Subsequent COC collections were on Days 4 and 9. A similar design was used in Exp. 2, with an additional treatment group given 5000 IU eCG (n = 8 per group). In Exp. 1, compared to the saline-treated group, a single dose of 2500 IU eCG resulted in a greater number of ≥8 mm follicles at the time of the Day 4 COC collection (P = 0.03), but not at the Day 9. In Exp. 2, treatment with 5000 IU eCG resulted in a greater number of ≥8 mm follicles than 2500 IU eCG or the saline treatment (37.5 ± 6.9, 17.5 ± 2.0, 16.9 ± 2.0; P = 0.01, respectively). Although the number of embryos produced/COC submitted to IVM was not different among groups (mean = 18.6%), treatment with 5000 IU eCG produced more than twice as many embryos per bison as unstimulated bison (0.8 vs 1.9). In summary, embryo production rates were higher from COC collected subsequent to follicular wave synchronization vs random stages of the wave, and ovarian superstimulation with eCG resulted in a dose-related increase in the number of ≥8 mm follicles, COC collected, and embryos produced. Repeated COC collections after successive wave synchronization resulted in similar follicular counts and embryo production rates within individuals, and the greatest number of follicles aspirated, COC collected, and embryos produced was in the anovulatory season. We conclude that the minimum-handling COC collection protocols in the present study are effective and provide realistic options for embryo production in wild bison.

Development of a letrozole-based synchronization protocol for fixed-time artificial insemination in beef cattle.

Experiments were done to determine ovulation synchrony following a 4-day letrozole treatment (Exp 1), compare the efficacy of a letrozole-based protocol with other commonly used synchronization protocols for FTAI (Exp 2), and test a new intravaginal letrozole-releasing device (LRD) with and without pre-synchronization (Exp 3). In Exp 1, heifers and lactating cows at random stages of the estrous cycle were given an LRD for 4 days, PGF at LRD removal, and GnRH at 48 or 60 h after PGF, or no GnRH. In Exp 2, heifers and lactating cows were assigned to three FTAI groups: i) LRD, ii) estradiol+progesterone, iii) 5-d Co-synch+PRID. In Exp 3, heifers were pre-synchronized with PGF or not (control) 8 days before insertion of either an X-LRD or T-LRD for 3 days; FTAI was done 48 h after device removal. In Exp 1, the variation in interval to ovulation in cows, but not heifers, in the GnRH 48-h group was less than half that in other groups (P < 0.05). In Exp 2, the P/AI was lower (P < 0.001) in the LRD group compared to the other groups. In Experiment 3, the X-LRD increased (P < 0.0001) plasma letrozole concentrations compared to the T-LRD; the pregnancy rate was not affected by pre-synchronization or the type of LRD. Although ovulation synchrony was achieved following LRD treatment, the LRD group had the lowest P/AI compared to other protocols, perhaps because of too short an interval between LRD removal and GnRH/FTAI. Drug delivery was enhanced with the new X-LRD.

Hypothalamic involvement and the role of progesterone in the NGF-induced LH surge pathway.

To elucidate the mechanism by which nerve growth factor (NGF) influences the LH secretory pathway in camelids, a series of experiments were done to determine the involvement of the hypothalamus (Experiment 1), the role of GnRH neurons (Experiment 2), and the effect of progesterone (Experiment 3) on the NGF-induced LH surge and ovulation in llamas. In Experiment 1, the declining phase of the NGF-induced LH surge was used to determine if the decline is a result of pituitary depletion or hypothalamic unresponsiveness. Female llamas were treated with NGF and, 7 h later, assigned to three groups and given a second dose of NGF (n = 5), a dose of GnRH (n = 5), or saline (n = 6). The LH response was attenuated after the second dose of NGF vs GnRH. In Experiment 2, Fos expression (marker of neuronal activation) in GnRH neurons was examined in the hypothalamus of llamas after NGF or saline treatment (n = 3 per group). Despite an LH surge in the NGF group but not in the saline group, no differences were detected between groups in Fos/GnRH co-expression. In Experiment 3, llamas in low-, medium-, and high-plasma progesterone groups (n = 4 per group) were treated with NGF. The NGF-induced LH surge did not differ among treatment groups. Results from the present study show that the induction of a preovulatory LH surge by NGF may be controlled by a novel pathway involving GnRH neuro-terminals downstream of the hypothalamus and is independent of progesterone influence.

Repair Failures Call for New Policies to Tackle Leaky Natural Gas Distribution Systems.

Methane leaks in natural gas systems are low-hanging fruit for near-term, locally driven climate policy. Recent work suggests this emissions source is larger than previously believed and that repairing a small number of high emitters can cost-effectively reduce system-wide leakage. How successful are these repairs on the ground? Here, we assess the effectiveness of repair policies in the Massachusetts distribution system. Our analysis leverages state-wide utility data, on-site empirical measurements, stakeholder interviews, and document and legal analysis. We use these mixed methods to investigate the rate of repair failure, where a gas utility identifies and fixes a leak, but on-site emissions are not eliminated. We find that repair failures are relatively common, yet they are repeatedly neglected in policy. By not accounting for repair failures, policy may overestimate the effectiveness of distribution system repairs in meeting local greenhouse gas reduction targets. These results also underscore the importance of data transparency for monitoring and verifying subnational climate policies.

Antral follicle counts and association with ovarian superstimulatory response to gonadotropins in prepubertal calves.

Antral follicle count (AFC) repeatability at the time of follicular wave emergence and duration of gonadotropin treatment in calves with small and large AFC affects the superstimulatory response of follicles. In Study I, the individual AFC was determined, calves were ranked as having a small, medium or large AFC, and a second count was performed prior to FSH treatments. There was a positive association between the number of follicles ≥1 mm after the first and second counts (r = 0.4; P = 0.003). In Study II, calves with small and large AFC were administered pFSH for 4 or 7 days, pLH 20 h after last pFSH administration and cumulus-oocyte-complexes (COC) were collected. In calves having large as compared with small AFC, number of follicles ≥6 mm were greater (P = 0.01) and COC collected were greater (P = 0.001). The proportion of large-sized follicles (>9 mm) was greater in the 7-day than in the 4-day gonadotropin treatment group (56.4 ±â€¯8.3 and27.8 ±â€¯7.5 %, respectively; P = 0.01). In Study III, there was a positive association between AFC and number of follicles ≥6 mm at the time of COC collection (r = 0.6; P = 0.003). In summary, the number of follicles at the time of follicular wave emergence was associated with the number of follicles recruited during subsequent waves of follicular development and ovarian response following gonadotropin superstimulation. Calves with a large AFC had more COC collected than calves with a small AFC.

Evidence for the LH-releasing pathway of seminal plasma NGF in male camelids.

In the female camelid, systemic administration of NGF induces a preovulatory LH surge that results in ovulation, but the effects of seminal NGF in the male are unknown. In the present study, we tested the hypothesis that the LH-releasing pathway of NGF is present in male camelids. In Experiment 1, male llamas and alpacas were treated with NGF or GnRH (n = 2 llamas and 3 alpacas) and blood samples were collected from 1 h before to 3 h after treatment. Plasma LH concentrations increased after treatment in a surge-like fashion in both GnRH- and NGF-treated groups, but concentrations reached a maximum 2.5 times higher and remained elevated for at least 2 h longer in the NGF-treated group (treatment-by-time interaction, P = 0.01). In Experiment 2, we evaluated the LH and testosterone response to NGF vs saline treatment (n = 3 llamas and 3 alpacas). The LH response to NGF was similar to that in Experiment 1, and plasma testosterone concentrations were higher in the NGF group than in the saline group at 2, 4 and 6 h after treatment (P < 0.05). Results support the hypothesis that the LH-releasing pathway for NGF exists in male South American camelids. The LH response to NGF sustained circulating testosterone concentrations in llamas, suggesting a moderate role of NGF in testosterone secretion.

Cholesterol-cyclodextrin complex as a replacement for egg yolk in bull semen extender: sperm characteristics post-thawing and in vivo fertility.

Egg yolk, a major semen extender constituent, lacks a defined composition, therefore, there are biosecurity concerns with use of egg yolk. Cryopreservation of bull semen without inclusion of animal protein in the semen extender, therefore, is an important consideration. Cholesterol may be delivered and incorporated into the sperm plasma membrane by cyclodextrins to protect sperm during cryopreservation. The aim of this study was to determine suitability of a cholesterol-cyclodextrin semen extender, without inclusion of egg yolk, for cryopreservation of bull semen. Bull semen was collected and cryopreserved in either egg yolk or with inclusions of three different concentrations of cholesterol-cyclodextrin complex (0.5, 1 or 2 mg/mL semen) in Tris-glycerol (TG) extender. Sperm motion characteristics examined using the computer-assisted sperm analysis, and plasma membrane and acrosome integrity examined using flow cytometry, were similar for all extenders. The inclusion of the greatest concentration of cholesterol-cyclodextrin complex (2 mg/mL semen) followed by dilution in TG extender resulted in lesser pregnancy rates (P <  0.05). There was a pregnancy rate of as great as 56 % when sperm cryopreserved in 0.5 mg/mL cholesterol-cyclodextrin Tris-glycerol extender were used for artificial insemination following imposing of a hormonal treatment regimen for synchrony of timing of ovarian functions among cows for conducting fixed-time artificial insemination (FTAI). Results indicate cholesterol-cyclodextrin Tris-glycerol extender, with a chemically defined composition and without inclusion of egg yolk, may be used to cryopreserve bull sperm with there being acceptable pregnancy rates when this semen is used for FTAI.

Neuroanatomical basis of the nerve growth factor ovulation-induction pathway in llamas†.

The objective of the study was to characterize the anatomical framework and sites of action of the nerve growth factor (NGF)-mediated ovulation-inducing system of llamas. The expression patterns of NGF and its receptors in the hypothalamus of llamas (n = 5) were examined using single and double immunohistochemistry/immunofluorescence. We also compare the expression pattern of the P75 receptor in the hypothalamus of llama and a spontaneous ovulator species (sheep, n = 5). Both NGF receptors (TrkA and P75) were highly expressed in the medial septum and diagonal band of Broca, and populations of TrkA cells were observed in the periventricular and dorsal hypothalamus. Unexpectedly, we found NGF immunoreactive cell bodies with widespread distribution in the hypothalamus but not in areas endowed with NGF receptors. The organum vasculosum of the lamina terminalis (OVLT) and the median eminence displayed immunoreactivity for P75. Double immunofluorescence using vimentin, a marker of tanycytes, confirmed that tanycytes were immunoreactive to P75 in the median eminence and in the OVLT. Additionally, tanycytes were in close association with GnRH and kisspeptin in the arcuate nucleus and median eminence of llamas. The choroid plexus of llamas contained TrkA and NGF immunoreactivity but no P75 immunoreactivity. Results of the present study demonstrate sites of action of NGF in the llama hypothalamus, providing support for the hypothesis of a central effect of NGF in the ovulation-inducing mechanism in llamas.

Effect of dose and duration of FSH treatment on ovarian response in prepubertal calves.

The objective of the study was to determine relative effects of dose (200 or 350 mg) and duration (4 or 7 days) of superstimulatory treatment on the ovarian response in prepubertal calves. Calves with similar antral follicular counts at wave emergence (n = 24) were given eight doses of either 25 or 44 mg pFSH every 12 h for 4 days or 14 doses of either 14 or 25 mg pFSH for 7 days beginning at the time of follicular wave emergence and 12.5 mg of pLH im 12 h after the last FSH treatment. On Day 4 of pFSH treatment, calves given 14 mg had fewer follicles ≥3 mm than those given 25 mg (15.1 ± 1.9 and 27.9 ± 3.3, respectively; P = 0.04). At the end of treatment (24 h post-LH), number of follicles ≥9 mm was greater in calves of groups treated with 350 than 200 mg (13.5 ± 1.8 and 8.8 ± 1.3, respectively; P = 0.02) and calves of groups treated for 7 than 4 days (13.3 ± 1.8 and 9.0 ± 1.3, respectively; P = 0.03). The number of spontaneous ovulations was greater in calves of groups treated for 7 than 4 days as was the total number of ovulations (9.7 ± 0.9 and 6.9 ± 1.0, respectively; P ≤ 0.05). In summary, a dose of 25 mg of pFSH per treatment given twice daily for 7 days resulted in a greater ovarian response than other superstimulatory treatments in prepubertal calves.

An attempt to potentiate the ovarian superstimulatory response in cattle by co-treatment with an aromatase inhibitor.

Letrozole is used for the treatment of subfertility in women undergoing ovarian superstimulation, but the mechanism of action has not been investigated critically. The objective was to test the hypothesis that treatment with letrozole will potentiate the superstimulatory response following gonadotropin treatment by increasing the number of follicles present at ovarian follicular wave emergence in cattle. In Experiment 1, ovarian follicular wave emergence was synchronized among beef heifers (n = 8) by transvaginal ultrasound-guided follicle ablation. On Day 0 (wave emergence), a letrozole-releasing device (LRD) was placed intravaginally for 5 days, followed again by transvaginal follicle ablation on Day 5. The number of follicles ≥3 mm was recorded by transrectal ultrasonography on Days 0 and 6.5 (i.e., pre- vs. post-LRD treatment). In Experiment 2, non-lactating dairy cows were assigned randomly to one of two groups (n = 15/gp) after follicle ablation-induced synchronization of wave emergence (Day 0), and given either an LRD or sham device for 5 days. Superstimulatory treatment was initiated on Day 0, consisting of 8 doses of 50 mg of porcine FSH im at 12 h intervals, and luteolytic doses of prostaglandin on Days 3 and 3.5. The LRD/sham devices were removed on Day 3.5, GnRH was given im on Day 5, estrus response was determined on Days 5 and 6, and the ovarian response was recorded by ultrasonography on Days 0, 3.5, 5, 6.5, and 12. In Experiment 1, no difference was detected in the number of antral follicles at wave emergence pre- vs. post-LRD treatment (23.2 ± 3.2 vs. 23.5 ± 3.8 follicles; P = 0.67; mean ± SEM). In Experiment 2, the interval from prostaglandin treatment to estrus was longer (50.3 ± 1.1 vs. 40.7 ± 2.0 h; P < 0.001) and less variable (residuals: 3.1 ± 0.5 vs. 6.7 ± 0.9 h; P < 0.01) in the LRD vs. sham group. The proportion of ovulations (number of CL on Day 12 over the number of follicles ≥3 mm on Day 0) did not differ (0.65 ± 0.02 vs. 0.70 ± 0.02; P = 0.15) nor did the number of CL on Day 12 (15.9 ± 2.5 vs. 19.0 ± 2.0; P = 0.32) between the LRD and sham groups. In summary, treatment with letrozole did not increase the number of antral follicles at wave emergence or the superstimulatory response, but increased precision in the interval to estrus and may be useful for artificial insemination at a fixed time in superstimulatory protocols.

Cryopreservation of bison semen without exogenous protein in extender and its fertility potential in vitro and in vivo following fixed-time artificial insemination.

Successful cryopreservation of bison semen is fundamental for restoration of genetic diversity in Canada's wood bison. Conventional bovine semen extenders contain animal products, such as egg yolk and milk, which are undesirable because of biosecurity risks and undefined composition. In this study, we examined the efficacy of an exogenous protein-free extender containing cholesterol-cyclodextrin complex (CC) to cryopreserve bison semen. The study also provided an opportunity to determine the effectiveness of different ovulation synchronization protocols for fixed-time artificial insemination in bison. Semen was collected from wood bison bulls via electroejaculation and cryopreserved in either Tris-egg yolk-glycerol (called 'TEYG') extender or pretreated with CC (2 mg/mL semen) and diluted in Tris-glycerol (collectively called 'CC-TG') extender. Post-thaw sperm motion characteristics and in vitro fertilization of cattle oocytes confirmed that CC alone without egg yolk protected bison sperm during cryopreservation process. In the first fertility trial, however, no pregnancy was obtained following fixed-time artificial insemination of bison cows with CC-TG extender. In a follow-up trial, low concentration of CC (1 mg/mL semen) resulted in better post-thaw sperm motion characteristics, fertility rate, and birth of live calves following fixed-time artificial insemination. Results showed that 1 mg CC/mL semen completely replaced egg yolk in bison semen extender. In addition, both follicular ablation and steroid treatment protocols provided ovulation synchrony to permit successful application of fixed-time artificial insemination in bison. This is the first report on the birth of live bison calves following fixed-time artificial insemination using semen cryopreserved in a defined extender.