Mean LH Concentrations Research Articles

Long-term Glucose Infusions had no Influence on Gonadotropins Hormone Secretion in Ovariectomized Goats

‌Background: It is reported that high nutritional supplementation treatment for seven days stimulated pulsatile LH secretion attended by increased in the plasma glucose and insulin concentrations in goat. The present study tested the hypothesis that whether providing peripheral glucose to goats associated with digestion of high nutrition supplementation increase gonadotropin secretion in ovariectomized goats. Materials and Methods: There was two experimental groups. The treatment animals from both experiences received glucose infusion. In the first experiment (constant glucose infusion; 50% glucose + saline at a rate of 7.5 ml/h for 7 days) and in the second experiment (progressive glucose infusion; 20% glucose + saline by increasing 5% glucose in 24hr intervals for 7 days at a rate of 15 ml/h for 7 days). The control animals from both experiences received saline infusion in the same manner. Findings: In the first experiment, there was no significant diversity in the concentrations of glucose and insulin among the treatment period as compared with mean pre-infusion period or compared to control groups except for Day 2 (p<0.05). In the second experiment, concentrations of both the glucose and insulin started increasing significantly from Day 2 and remained with elevated levels until day 7 as compared with mean pre-infusion period or compared to control groups (p<0.05). The mean Follicle stimulating hormone concentration in treatment group was not affected by glucose infusion as compared with pre-infusion periods or control group. The mean LH (concentration and pulse frequency) was not affected by glucose or saline infusion in both experiments. Conclusion: The present studies suggest that an increase in the glucose availability does not control the gonadotropin secretion as a stimulatory mediator in ovariectomized goats.

PMON257 Oral Ingestion of Macronutrients Selectively and Variably Decrease Circulating Testosterone Concentration in Healthy Men

Abstract We have previously reported acute suppression of testosterone in men after oral ingestion of dextrose. In the present study, we aimed to explore a potentially similar effect in response to other macronutrients. Eleven healthy men (age: 33-70 yrs, BMI 20.4-31.5 kg/m2) were studied in random order after an overnight fast on 4 separate days, involving oral ingestion of 300 ml of either water (control), dextrose, protein, or lipid solutions. Sessions were 6.5 h long, starting at 0800-0900 hrs. Test meals were isocaloric (400 kcal) and were ingested after the 4th blood draw (30 min point). Blood was collected at 10-min intervals for testosterone (ng/dL), and LH (IU/L) measurements. Means of the 1st 4 pre-intervention measures were used as the first value in all the time series. Mean delta-serum (macronutrients-water) testosterone and LH concentrations sampled at 10-minute intervals over 6 hour period were adjusted to the corresponding 1st value to correct for among the sessions inconsistent baseline testosterone and LH measures. Compared to the water day (control), mean (± SE) serum testosterone concentrations (ng/dL) in 11 subjects decreased from 415 ± 36 (control) to 370 ± 31 (dextrose: P=0.009), 340 ± 28 (protein: p=0.002), and 355 ± 30 (lipid: p=0.01). Corresponding LH (IU/L) values for dextrose (3.16 ± 0.29), protein (3.17 ± 0.39), and lipid (3.07 ± 0.29) were not significantly different from the control day (3.3 ± 0.29). Mean delta-testosterone time series revealed decreased testosterone of 63 ng/dL at 130 min after dextrose, 80 ng/dL at 200 min after protein, and 10 ng/dL at 70 min after lipid ingestion. Duration of suppression was variable among the 3 macronutrients, but most prolonged for protein lasting for 320 min. As oppose to testosterone, delta-LH concentrations were more positive rather than negative, suggestive of a potential testicular insult. In conclusion, oral ingestion of all macronutrients decrease circulating testosterone concentration, which appear to be independent of LH secretion. The effect is more intense and prolonged after protein intake, and minimal with lipid ingestion. Future studies are required to define the etiology, and the macronutrient effect on more prolonged basis. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

252 Impact of Pre- and Postnatal Nutritional Extremes on Tonic Secretion of Gonadotropins and Feedback Responsiveness to Estradiol in Sexually Mature Heifers

Abstract Objectives were to test the hypotheses that nutritional extremes during prenatal development interact with postnatal nutrition during the juvenile period of heifers to impact 1) tonic secretion of gonadotropins, and 2) estradiol-17β (E2) negative and positive feedback responsiveness in adulthood. Heifers were selected from a larger population programmed nutritionally using a 3 x 2 factorial arrangement of pre- and postnatal diets. Beginning at 90 days of pregnancy, Bos indicus-influenced cows (n = 95) were fed to achieve body condition scores (BCS; 1–9 scale) of 3–3.5 (L; thin), 5.5–6 (M; moderate), or 7.5–8 (H; obese) by onset of the third trimester and maintained thereafter. Heifer offspring were weaned at 3–3.5 months of age and assigned to either a low- (L; 0.5 kg/day) or high-gain (H; 1.0 kg/day) diet until 8 mo. of age, then fed a common diet until puberty. Heifers (n = 18; 6/grp) representing HH, MH and LL combinations were ovariectomized postpubertally (17.1 mo. of age) and received E2 replacement. In Exp. 1, blood samples were collected at 10-min intervals to evaluate pulsatile secretion of LH and FSH for 5.5 hours. In Exp. 2, heifers received E2 (2.4 ug/kg I.M.), with blood sampling at 30-min to 1-h intervals for 30 h. Heifers in the MH (1.25 ± 0.11 ng/mL) group tended (P < 0.09) to have greater LH pulse amplitude compared to HH (0.91 ± 0.14 ng/mL) and LL (0.96 ± 0.09 ng/mL); otherwise, frequency, amplitude, and mean concentrations of LH and FSH did not differ among groups. Exogenous E2 (Exp. 2) suppressed (P < 0.0001) mean plasma concentrations of LH and FSH equally among groups, then stimulated equivalent surges of LH beginning at 14 ± 0.02 h, with only two FSH surges detected. Neither pre- nor postnatal nutrition affected E2 negative or positive feedback in this study.

Chronic Undernutrition Inhibits KNDy Neurons in Ovariectomized Ewe Lambs

Undernutrition negatively impacts reproductive success, at least in part, through a central suppression of GnRH secretion. Given that GnRH neurons are devoid of receptors for peripheral metabolic hormones such as leptin and insulin, nutritional regulation of GnRH secretion must be through afferent input. Neurons which co-express kisspeptin, neurokinin B (NKB), and dynorphin, termed KNDy neurons, are a unique population of cells in the arcuate nucleus (ARC) of the hypothalamus and are believed to play a critical role in GnRH/LH pulse generation. With our recent evidence that chronic feed restriction reduced kisspeptin and NKB in young, castrated male sheep, we hypothesized that nutrient restriction would inhibit expression of the KNDy neuron peptides kisspeptin and NKB, while increasing dynorphin expression in young, ovariectomized female sheep. Fifteen ewe lambs were ovariectomized and were fed to maintain body weight (n=7; Fed) or feed-restricted to lose 20% of pre-study body weight (FR; n=8). Blood samples were taken weekly every 12 minutes for 4.5 hours via jugular venipuncture and plasma was stored at -20°C until assessment of LH using radioimmunoassay. Body weights were recorded weekly and feed amounts were adjusted to achieve desired body weights. Following blood collection at Week 13, animals were euthanized, brain tissue was perfused with 4% paraformaldehyde, and tissue containing the hypothalamus was collected for assessment of KNDy neuropeptide mRNA abundance by in situ hybridization (RNAscope). At Week 13, the average percent change in body weight was clearly evident (Fed, 7.35 +/- 2.2% vs FR, -21.36 +/- 0.9%), and mean LH concentrations were lower in FR ewes (10.70 +/- 3.1 ng/ml) compared to Fed controls (20.98 +/- 3.8 ng/ml). Data analyzed to date for kisspeptin, NKB, and dynorphin in the ARC show that feed restriction reduced the number of kisspeptin mRNA-expressing cells (Fed, 165 +/- 25 vs FR, 9.25 +/- 6), the number of NKB mRNA-expressing cells (Fed, 141 +/- 28 vs FR, 24 +/- 5), and the number of dynorphin mRNA-expressing cells (Fed, 109 +/- 41 vs FR, 29.5 +/- 24). Together, these findings demonstrate that chronic feed restriction suppresses KNDy neurons and supports a role for these key reproductive neurons in the central mechanism governing GnRH/LH secretion during undernutrition in female sheep.

OR32-04 Dynamic Interactions Between Luteinizing Hormone and Testosterone in Healthy Community-Dwelling Men: Impact by Age and Body Composition

Context. Aging is associated with diminished testosterone (Te) secretion, which could be attributed to Leydig cell dysfunction, decreased pituitary stimulation and altered Te feedback. Objective. The goal was to quantify all three regulatory nodes of the GnRH-LH-Leydig cell- axis in the same cohort of healthy men, by measuring (1) indirectly the strength of the endogenous GnRH signal on the gonadotrope, (2) the strength of Te feedback on LH by ketoconazole (KTCZ), and (3) the effect of LH infusions on Te secretion, in relation to age and body composition. Design. This was a placebo-controlled, blinded, prospectively randomized cross-over study in 40 men, age 19–73 yr, BMI 20–34.3 kg/m2. A submaximal dose of ganirelix (GnRH antagonist) was used to assess outflow of GnRH, by calculating the difference between LH output during the control and ganirelix arm. Ketoconazole (steroidogenic inhibitor) was used to estimate feedback, by the difference in LH output during ketoconazole and control arm. High-dose ganirelix and repeated 6-min LH (18.75 IU) infusions were used to measure testicular responsivity. Blood sampling was at 10-min intervals. The 4 sessions were concluded with, a single submaximally GnRH stimulus to assess the responsiveness of the gonadotrope during ganirelix inhibition. Setting. The study was performed in a Clinical Translational Research Unit. Interventions. In 3 of the 4 experiments subjects underwent 5 h of blood sampling at 10-min intervals, starting at 0800 h. At 1100 h GnRH was injected and sampling was continued for another 2 h. Admission was at 1700 h the day before. At 2000 h they received KTCZ, dexamethasone or ganirelix and/or placebo. KTCZ and dexamethasone (or placebo) were administered again at 0700 when the IV catheter was placed. High-dose ganirelix was used to test the testicular responsiveness, and 7 LH pulses (90 min intervals) were given., with blood sampling from 1500 till 1300 h next day. Outcome measures. Mean concentrations of LH and (bio)Te, deconvolution analysis, endogenous dose-response LH-bioTe relation, and approximate entropy. Abdominal visceral fat (AVF) was calculated from single slice CT. Results. There were age-, but not body composition-related decreases in estimated endogenous GnRH secretion, Te’s feedback strength on LH, and Leydig cell responsivity to LH, accompanied by changes in approximate entropy. Bioavailable Te levels were negatively related to both age and AVF, without interaction between these variables. The LH response to a submaximal dose of GnRH was independent of age and AVF. Conclusion. Advancing age is associated with 1) attenuated bioavailable Te secretion caused by diminished GnRH outflow and not by decreased GnRH responsivity of the gonadotrope, 2) diminished testicular responsivity to infused LH pulses, and 3) partial compensation by diminished Te feedback on central gonadotropic regulation.

Central obestatin administration affect the LH and FSH secretory activity in peripubertal sheep

Obestatin – a 23 amino acid peptide is synthesized as another product of the ghrl gene and its synthesis occurs mainly in gastric mucosa cells. This hormone is involved in complex gut-brain neurohormonal networks, thereby can participates in the modulation of gonadotrophic axis activity.The aim of this study was to investigate the consequence of intracerebroventricular infusions of obestatin on LH and FSH pituitary cells secretory activity in peripubertal female sheep. Animals were randomly divided into two groups: the control group (n = 14) received intracerebroventricular infusions of Ringer-Lock solution (120 μL h−1), and the obestatin group (n = 14) was infused with obestatin (25 μg/120 μL h−1) diluted in Ringer-Lock solution. A series of four infusions was performed on three consecutive days. Blood samples were collected on day 0 and day 3. The sheep were slaughtered immediately after the end of the experiment. For molecular biological analysis, pituitaries from 7 sheep from each group (n = 7 + 7) were prepared and frozen in liquid nitrogen immediately after collection and then stored at −80 °C until Real Time RT-qPCR and RIA analyzes. For immunohistochemical analysis, pituitary tissues from the remaining animals (n = 7 + 7) was fixed in situ for further examination.Real-Time qPCR and immunohistochemistry analyses revealed substantial changes in the LH and FSH pituitary cells secretory activity in obestatin-infused sheep. Exogenous obestatin administration reduced LHβ mRNA expression and increased the accumulation of immunoreactive LH in gonadotrophic cells of the adenohypophysis. These changes were accompanied by a decrease in the mean LH concentration in the peripheral blood resulting from the lower LH pulse amplitude. Moreover, an increase in both FSHβ mRNA expression and FSH immunoreactivity and amount in pituitary cells were noted, while mean blood FSH concentration remained unchanged after obestatin treatment.The obtained results showed that exogenous obestatin affected LH secretory activity at the level of protein synthesis, accumulation and release as well as obestatin increase FSHβ mRNA expression and accumulation of this hormone but at the same time have no effect on FSH release to blood. Thus, obestatin can participate in the neuroendocrine network, which modulates gonadotrophic axis activity in sheep.

373 Response to kisspetin as a possible reference for puberty age in heifer

Abstract The objective of this study was to compare the variation of LH concentrations in response to kisspeptin as a way to evaluate the neuroendocrine maturation of Nellore heifers. The hypothesis was that the LH response to kisspeptin increases with age as the number of kisspeptin receptors increase in the CNS. Initially 50 weaned heifers were segregated according to the weaning weight as light (174±1 kg) or heavy (232±2 kg). Both groups received a kisspeptin injection (10μg/kg; IM) monthly from weaning to 16 months, and blood samples were collected 20 minutes after each application. After 16 months of age, heifers were exposed to bulls (25:1) for 60 days and pregnancy was determined 30 days after the end of the breeding season. The LH concentration in response to kisspeptin increased over time (P = 0.001), but there was no difference between light and heavy heifers over the 8-month period (3.62±0.14, 3.34±0.16 ng/ml; P = 0.3853). Similarly, no differences were observed between heifers that became pregnant or not (3.42±0.12, 3.67±0.20 ng/ml; P = 0.3169). Due to the high LH concentrations at 8 months of age (3.06±0.19 ng/ml), another experiment was performed in younger heifers. Fifty newborn Nellore heifers (29±0.3 kg) from the same herd received monthly injections of kisspeptin until weaning. There was also an increase in LH concentrations in response to kisspeptin over time (P = 0.001), but the mean LH concentrations in this experiment (4.05±0.22ng/mL) were considerably greater than those in postweaning heifers. Our interpretation is that genetic differences likely contributed to the different responses to kisspeptin between the two experiments. In order to test this hypothesis, the LH response to kisspeptin will be evaluated in a large number of animals from different herds at weaning to determine the effect of genetic variability on kisspeptin response and possibly to use this phenotype as a marker for age at puberty.

Effect of short-term nutritional supplementation on hormone concentrations in ovarian follicular fluid and steroid regulating gene mRNA abundances in granulosa cells of ewes

This study was conducted to investigate effects of short-term nutritional supplementation on concentrations of reproductive hormones in follicular fluid and on mRNA abundance in granulosa cells (GCs) during the luteal phase of ewes. Eighteen ewes were randomly assigned to treatment or control groups (n = 9, each group). All the ewes were subjected to estrous synchronization using a Controlled Intravaginal Releasing Device (CIDR). From the second to the eleventh day of estrous synchronization, ewes were fed a diet with a relatively greater (treatment group) or a maintenance (control group) energy content. Samples of follicular fluid and GCs were collected from antral follicles of estrous ewes after CIDR removal. The results indicate mean FSH concentration of fluid in small follicles and mean LH concentrations of fluid in large follicles of the ewes in the treatment group were greater (P < 0.05) than that of ewes in the control group. Follicular fluid E2 concentrations were directly related (P < 0.05) to follicular diameter. The ewes of the treatment group had greater (P < 0.05), compared with the control group, abundances of Follicle Stimulating Hormone Receptor (FSHR) in small and medium follicles, and (P<0.05) Luteinizing Hormone Receptor (LHR), Steroid Acute Regulatory Protein (STAR), cytochrome P450 (CYP17A1, CYP19A1) enzyme and Estrogen Receptor (ESR1) in large follicles. The results of this study provide evidence for a potential mechanism by which short-term nutritional supplementation improves follicular development possibly because of increased expression of steroid synthesis-regulating genes in large follicles.

Effect of Inhibin-B hormone and its relationship with a number of sex hormones in men with Azoospermia

Inhibin-B is produced by sertoli cells and acts as a negative feedback mechanism of follicular stimulation hormone (FSH). In order to evaluate the effect of Inhibin-B in men with Azoospermia, the study measured the concentrations of the following hormones: Inhibin-B, FSH, Luteinizing Hormone Prolactin (PRL), (LH) and Testosterone (Testo). Serum for 30 normal men semen and 50 men with Azoespermia (zero sperm count in sperm). The mean concentration of the hormone Inhibin-B in patients with Azoospermia was (135.7 ±87.6) pg/ml, while in normal men (175.6 ±47.9) Pg / ml with a significant value of the group of healthy patients at a significant level (p≤0.01). The mean serum FSH concentration in patients was (13.7 ± 12.27) lU / L whereas in normal men (4.1 ± 2.84) lU / L, the relationship was significant between patients and healthy at a significant level (P≤0.01) the correlation with the Inhibin-B hormone is reversible relationship. The mean concentration of LH in patients was (12.8±11.87) IU/L and in the healthy (7.9±5.57) IU/L and the relationship between them was significant (P≤0.05) and the correlation with the Inhibin-B hormone is positive relationship. The mean concentration of the Testosterone In the patients was (2.37 ± 1.22) IU / L and in the healthy (5.1 ± 1.96) IU/ L, the relationship was of significant value at P≤0.01 and the correlation with the Inhibin-B hormone is positive relationship, and there is a difference in the mean concentration of PRL between the patients and the healthy where it reaches (8.88 ±5.16) IU / L in patients and (8.03 ± 4.2) IU / L in the healthy. where the relationship was non-significant value between both patients and healthy and the correlation with the Inhibin-B hormone is positive relationship

Patterns of preoptic-hypothalamic neuronal activation and LH secretion in female sheep following the introduction and withdrawal of novel males.

The neuroendocrine response of female sheep to a novel male involves neural activation in the hypothalamus. However, if males are removed, the gonadotrophic signal declines, so the neural activity is likely to change. We examined Fos-immunoreactive (IR) cells in hypothalamic tissues from seasonally anovulatory female sheep exposed to males for 2 or 6h, or for 2h followed by 4h isolation from males. Control females were killed in the absence of male exposure. Male introduction increased LH secretion in all females; male removal was associated with a reduction only in mean and basal LH concentrations. Females exposed to males for 2h had more Fos-IR cells in the arcuate nucleus (ARC), ventromedial nucleus of the hypothalamus (VMH) and organum vasculosum of the lamina terminalis (OVLT) than control females. Fos-IR cells in the preoptic area (POA) were only greater than in control females after 6h exposure to a male. Removal of males decreased the number of Fos-IR cells in the ARC, VMH and OVLT, but not in the POA. Thus, hypothalamic neural activation and LH secretion in female sheep are stimulated by males and decline after male removal. However, activation in the POA persists after removal and may explain the incomplete decline in the LH response.

Stężenia gonadotropin i α podjednostki w surowicy krwi u pacjentów z gruczolakami gonadotropowymi przysadki w zależności od immunoreaktywności guza

Although active gonadotropin-secreting pituitary adenomas are considered very rare, the vast majority of pituitary tumours diagnosed as "non-functioning" express gonadotropins or their free β or α subunits. However, systemic investigations comparing the serum concentrations of follitropin (FSH), lutropin (LH), and α-subunit (αSU) before surgery with the immunoreactivity of the respective substances in the excised tumours are still lacking. Immunostaining of FSH, LH, and αSU was compared in 43 surgically removed gonadotropin - expressing pitu-itary adenomas with serum concentrations of the above-mentioned substances before surgery in the same patients. The serum concentrations of FSH were elevated (> 11.6 mU/mL) in 8/12 (66.7%) cases of FSH-positive adenomas. By contrast, in FSH-negative tumours the elevation of FSH is absent. Moreover, only 1/25 (4%) patients with LH-positive adenoma had the elevated serum concentration of LH (51.5 mU/mL). The overproduction of LH was not observed in adenomas expressing free β LH or in LH-negative tumours. In patients with αSU-positive adenomas elevated serum levels of αSU were observed in 3/15 (20%) cases. No αSU elevations were observed in patients with αSU-negative adenomas. The mean serum FSH, LH, and αSU concentrations were higher in patients with FSH, LH, and/or αSU immunopositive tumours in comparison with immunonegative. However, the differences are not statistically significant. Although "silent" gonadotropinomas constitute a frequent subtype of pituitary adenomas, the "active" subtype (i.e. manifesting by gonadotropin excess) are rare (approx. 4% of all pituitary adenomas). Gonadotropinomas are difficult to diagnose before surgery. The measurement of gonadotropins including αSU is needed but often not sufficient for presurgical diagnosis.

Resistin regulates reproductive hormone secretion from the ovine adenohypophysis depending on season

Work in cattle and rodents has shown that resistin, in addition to its roles in insulin resistance and inflammation, is involved in the regulation of gonadal steroidogenesis. However, the role of resistin in the regulation of reproductive processes in other species, such as seasonally breeding sheep, is completely unknown. Herein, we tested the hypothesis that resistin can influence the secretion of anterior pituitary hormones and that its effect in ewes is dependent on the day length. Thirty Polish Longwool ewes, a breed that exhibits a strong seasonal reproductive pattern, were ovariectomized with estrogen replacement using subcutaneously inserted estradiol implants. Ewes were fed ad libitum and housed under a natural photoperiod (longitude: 19°57′ E, latitude: 50° 04′ N). Intravenous treatments consisted of control or recombinant bovine resistin (rbresistin) in saline: (1) control (saline; n = 10), (2) low resistin dose (1.0 μg/kg BW; n = 10), and (3) high resistin dose (10.0 μg/kg BW; n = 10). Experiments were conducted during both short-day (SD) and long-day (LD) seasons using 5 sheep per group within each season. Blood samples were collected every 10 min over 4 h. Blood plasma concentrations of FSH, LH, and prolactin (PRL) were assayed using RIA. A season × dose interaction was observed for all hormonal variables measured. Greater concentrations (P < 0.001) of LH and FSH were observed during SDs than during LDs in all groups. During SDs, the high dose (10 μg/kg BW) decreased (P < 0.001) basal LH levels and amplitude (P < 0.05) of LH pulses and increased (P < 0.001) circulating concentrations of FSH. However, the low dose of resistin decreased (P < 0.001) FSH concentrations compared to those of controls. During LDs, both the low and high resistin doses increased mean concentrations of LH (P < 0.001 and P < 0.05, respectively) and FSH (P < 0.001). A high dose of rbresistin increased (P < 0.001) the mean circulating concentrations of PRL during both seasons. However, in all groups, concentrations of PRL were greater during LDs than SDs. These results demonstrate for the first time that resistin is involved in the regulation of pituitary hormone secretion and that this effect is differentially mediated during LDs and SDs.

Intravaginal instillation of gonadotropin-releasing hormone analogues with an absorption enhancer induced a surge of luteinizing hormone in lactating dairy cows

Our objectives were to evaluate circulating LH concentrations after intravaginal (IVG) instillation of GnRH analogs in lactating dairy cows. In 2 experiments, lactating Holstein cows (experiment 1: n = 32; experiment 2: n = 47) received the experimental treatments 48 h after the first of 2 PGF2α treatments given 12 h apart and 7 d after a modified Ovsynch protocol (GnRH at -7 d, PGF2α at -24 h, PGF2α at -56 h, GnRH at 0 h). In experiment 1, cows were stratified by parity and randomly allocated to receive the following treatments: 2 mL of saline IVG (SAL, n = 6), 100 µg of gonadorelin (Gon) i.m. (G100-IM, n = 5), and 100 (G100, n = 7), 500 (G500, n = 8), or 1,000 µg of Gon IVG (G1000, n = 7). In experiment 2, treatments were SAL (n = 8), G100-IM (n = 8), G1000 (n = 7), 1,000 µg of Gon plus 10% citric acid (CA) IVG (G1000CA, n = 8), 80 µg of buserelin IVG (B80, n = 8), and 80 µg of buserelin plus 10% CA IVG (B80CA, n = 8). In both experiments, blood was collected every 15 min from -15 min to 4 h, and every 30 min from 4 to 6 h after treatment. Data for area under the curve (AUC), mean LH concentrations, and time to maximum LH concentration were analyzed by ANOVA with (mean LH only) or without repeated measures using PROC MIXED of SAS (version 9.4, SAS Institute Inc., Cary, NC). The proportion of cows with a surge of LH was evaluated with Fisher's exact test using PROC FREQ of SAS. In both experiments, LH concentrations were affected by treatment, time, and the treatment by time interaction. In experiment 1, the AUC for LH and maximum LH concentration were greatest for the G100-IM treatment and were greater for the G1000 than for the SAL and G500 treatments. The proportion of cows with an observed surge of LH was 100 and 0% for cows that received Gon i.m. and IVG, respectively. In experiment 2, the AUC and maximum LH concentrations were greater for the G100-IM, G1000CA, and B80CA treatments than for the other IVG treatments. The proportion of cows with a surge of LH differed by treatment (SAL = 0%, G100-IM = 100%, G1000 = 14%, G1000CA = 88%, B80 = 13%, and B80CA = 100%). For the treatments with a surge of LH, time to maximum concentration of LH was the shortest for the G100-IM treatment, intermediate for the G1000CA treatment, and the longest for cows in the B80CA treatment. In conclusion, Gon (up to 1,000 µg) absorption through intact vaginal epithelium after a single IVG instillation was insufficient to elicit a surge of LH of normal magnitude. Conversely, IVG instillation of 1,000 µg of Gon and 80 µg of buserelin with the addition of citric acid as absorption enhancer resulted in a surge of LH of similar characteristics than that induced after i.m. injection of 100 µg of Gon.

Characterization of the variability and repeatability of gonadotropin-releasing hormone-induced luteinizing hormone responses in dairy cows within a synchronized ovulation protocol.

The primary objective was to determine the variability and repeatability of GnRH-induced LH responses. The secondary objective was to evaluate the associations among plasma LH, FSH, estradiol (E2), and progesterone (P4) concentrations. One hundred lactating Holstein cows (35 primiparous, 65 multiparous) were initially subjected to a presynchronization protocol (d 0, PGF2α; d 3, GnRH) followed 7 d later by Ovsynch (d 10, GnRH; d 17, PGF2α; 56 h later, GnRH) and timed artificial insemination 16 h after the last GnRH. Blood samples were collected immediately before the GnRH injection of presynchronization and the second GnRH of Ovsynch to determine plasma concentrations of LH, FSH, and P4. A second blood sample was collected 2 h after each of the above GnRH injections to determine GnRH-induced LH and FSH concentrations. Plasma concentrations of E2 were also determined in samples collected immediately before the second GnRH of Ovsynch. Cows that (1) had higher LH concentrations at 0 h than at 2 h after GnRH, (2) showed an ongoing spontaneous LH surge, (3) did not respond to GnRH, and (4) had P4 ≥ 0.5 ng/mL at GnRH of presynchronization and the second GnRH of Ovsynch were excluded from the analysis. The variability (coefficient of variation) and repeatability [between animal variance/(within animal variance + between animal variance)] of GnRH-induced LH response were determined from samples collected 2 h after the GnRH of presynchronization and the second GnRH of Ovsynch. The associations among plasma LH, FSH, E2, and P4 were determined at the second GnRH of Ovsynch. Mean (±SEM) LH concentrations before GnRH were 0.5 ± 0.04 and 0.6 ± 0.03 ng/mL, whereas mean LH concentrations 2 h after GnRH were 9.8 ± 1.0 and 12.1 ± 0.8 ng/mL at GnRH of presynchronization and the second GnRH of Ovsynch, respectively. The variability of GnRH-induced LH was 76.1 and 52.1% at GnRH of presynchronization and the second GnRH of Ovsynch, respectively. The repeatability estimate for GnRH-induced LH concentration between GnRH of presynchronization and Ovsynch assessments was 0.10. Plasma concentrations of LH were positively associated with FSH and E2 (r = 0.61 and 0.30, respectively) and negatively associated with P4 (r = -0.46) at the second GnRH of Ovsynch. In summary, GnRH-induced LH responses were highly variable and unrepeatable, and LH concentrations were positively associated with FSH and E2 and negatively associated with P4.

Rumen-protected arginine alters blood flow parameters and luteinizing hormone concentration in cyclic beef cows consuming toxic endophyte-infected tall fescue seed.

The objective of this study was to determine the effect of rumen-protected arginine on median caudal artery blood flow and LH dynamics in cows fed toxic endophyte-infected tall fescue seed. Four ruminally cannulated nonlactating beef cows (539 ± 30 kg) were used in a 2 × 2 factorial arrangement of treatments utilizing a 4 × 4 Latin square design with 4 periods of 31 d each. Each cow was assigned to individual pens and fed orchardgrass hay (10.3% CP and 85% NDF; OM basis) during a 10-d adaptation period, followed by a 21-d collection period in which each cow was assigned 1 of 4 treatments: 1) rumen-protected Arg (180 mg/kg of BW) and 1.0 kg/d of toxic endophyte-infected fescue seed (AE+), 2) rumen-protected Arg (180 mg/kg of BW) and 1.0 kg/d of endophyte-free fescue seed (AE-), 3) 1.0 kg/d of toxic endophyte-infected fescue seed (E+) alone, or 4) 1.0 kg/d of endophyte-free fescue seed (E-) alone. In each period, Doppler ultrasound measurements for blood flow parameters were quantified on d 1, 5, 10, 15, and 20. On d 20 of each period, blood samples were collected every 10 min for 6 h and then once every hour for 12 h for LH response following exogenous GnRH. There was an Arg × fescue seed type interaction ( = 0.05) for median caudal artery blood flow due to an increase in blood flow in cows fed rumen-protected Arg with endophyte-free fescue seed. In addition, mean blood flow velocity in the artery was greater ( = 0.01) with the inclusion of rumen-protected Arg in the diet. Median caudal artery area ( = 0.03) and diameter ( = 0.01) were decreased in cows consuming E+ compared to those consuming E- with no effect ( ≥ 0.38) by Arg inclusion. Circulating nitric oxide (NO) concentrations tended to be influenced ( = 0.09) by the interaction of Arg × fescue seed type with E+ alone decreasing NO concentrations. Circulating NO concentrations were unaffected by rumen-protected Arg ( = 0.48). Mean serum LH concentration exhibited ( = 0.02) an Arg × fescue seed type interaction. Cows consuming E+ had decreased ( < 0.05) LH concentrations compared to all other treatments. However, cows consuming AE+ had ( ≥ 0.67) LH concentrations similar to those of cows consuming AE- and E-. Thus, supplementing rumen-protected Arg to cows consuming toxic endophyte-infected fescue seed has the potential to increase reproductive performance and peripheral blood flow.

Comparative studies on testicular and epididymal morphology, and serum hormone concentrations in foxes and the hybrids during the breeding season

The silver fox and the blue fox belong to different genera, and the hybrid males are fully or partially sterile. In the present study, the objective was to evaluate the causes of hybrid male sterility, and therefore analyze the differences in testicular, and epididymal morphology and serum hormone concentrations among silver foxes, blue foxes, and the hybrids during the breeding season. Samples were collected from 20 male silver foxes, 20 male blue foxes, 15 male HSBs (silver fox female×blue fox male hybrids) and 14 male HBSs (blue fox male×silver fox female hybrids), respectively. Seminal evaluation showed large numbers of sperm present in the semen of blue foxes and silver foxes, but no sperm present in the hybrids. Mean testicular volume and the diameter of seminiferous tubules in silver foxes and blue foxes were greater than in the hybrids; and there were many Sertoli cells, spermatogenic cells, and sperm in silver foxes and blue foxes, while spermatogenic cells decreased with no sperm in the hybrids. Mean serum LH and prolactin concentrations in silver foxes and blue foxes were less and testosterone was greater than in the hybrids (P<0.05). The results indicate that germ cell meioses in the hybrids were arrested at the prophase stage of meiosis, and that lesser concentrations of testosterone and greater concentrations of LH and prolactin can inhibit the completion of spermatogenesis.

Feedback effects of estradiol and progesterone on ovulation and fertility of dairy cows after gonadotropin-releasing hormone-induced release of luteinizing hormone

An experiment was conducted with the objective to determine the effects of estradiol, progesterone, presence of a corpus luteum (CL), and size of a dominant follicle on the characteristics and patterns of GnRH-induced LH release and subsequent ovulation during a timed artificial insemination (TAI) program, or a combination of these. In 70 lactating dairy cows, a total of 163 blood collection periods resulting in a GnRH-induced LH release were analyzed. Concentrations of LH were measured in hourly samples (0 through 6 h after GnRH) during each of the blood collection periods, whereas concentrations of progesterone and estradiol were measured in the sample before GnRH treatment (0 h). Measures of LH included time to LH peak concentration during the 6-h blood collection period, the 2 largest concentrations of LH, mean, and variance of the 6 LH concentrations under each LH curve. Individual and combination effects of CL presence and a dominant follicle≤or >13.5mm, in addition to individual and combination effects of progesterone: low (<0.45 ng/mL; n=83), medium (0.53 to 2.41 ng/mL; n=25), and high (2.66 to 10.7 ng/mL; n=55), and estradiol: low (<4.0 pg/mL; n=89) and high (≥4.0 pg/mL; n=74) were independent variables in models to determine their influence on characteristics of LH and ovulation. Injections of GnRH induced LH release during 6 h after each of 163 injections. Measures of GnRH-induced LH concentration were inhibited at greater concentrations of progesterone and in the presence of a CL. In contrast, GnRH-induced LH concentrations were increased when estradiol was ≥4.0 pg/mL, but relatively unaffected by the size of the dominant follicle. Furthermore, resulting incidences of ovulation were decreased at greater progesterone concentrations and presence of a CL, and increased at greater estradiol concentrations and presence of follicles >13.5mm. In cows with or without a CL, the presence of a follicle >13.5mm did not increase mean LH concentration or incidence of ovulation. We conclude that when progesterone concentration exceeded 0.5 ng/mL at the time of GnRH treatment, subsequent LH concentrations and ovulation were suppressed. At that same concentration of progesterone or when concentrations of estradiol were ≥4 pg/mL, TAI pregnancy outcomes were improved in the face of similar incidences of ovulation suggesting greater progesterone or lesser estradiol at the time of AI may inhibit pregnancy establishment by other mechanisms.

A comparative study about toxicity of CdSe quantum dots on reproductive system development of mice and controlling this toxicity by ZnS coverage

Objective(s): Medicinal benefits of quantum dots have been proved in recent years but there is little known about their toxicity especially in vivo toxicity. In order to use quantum dots in medical applications, studies ontheir in vivo toxicity is important. Materials and Methods: CdSe:ZnS quantum dots were injected in 10, 20, and 40 mg/kg doses to male mice10 days later, mice were sacrificed and five micron slides were prepared structural and optical properties of quantum dots were evaluated using XRD. Results: Histological studies of testis tissue showed high toxic effect of CdSe:ZnS in 40 mg/kg group. Histological studies of epididymis did not show any effect of quantum dots in terms of morphology and tube structure. Mean concentration of LH and testosterone and testis weight showed considerable changes in mice injected with 40 mg/kg dose of CdSe:ZnS compared to control group. However, FSH and body weight did not show any difference with control group. Conclusion: Although it has been reported that CdSe is highly protected from the environment by its shell, but this study showed high toxicity for CdSe:ZnS when it is used in vivo which could be suggested that shell could contribute to increased toxicity of quantum dots. Considering lack of any previous study on this subject, our study could potentially be used as an basis for further extensive studies investigating the effects of quantum dots toxicity on development of male sexual system.

Ovarian and Hormonal Responses to Follicular Phase Administration of Investigational Metastin/Kisspeptin Analog, TAK‐683, in Goats

This study evaluated the effects of follicular phase administration of TAK-683, an investigational metastin/kisspeptin analog, on follicular growth, ovulation, luteal function and reproductive hormones in goats. After confirmation of ovulation by transrectal ultrasonography (Day 0), PGF2α (2 mg/head of dinoprost) was administered intramuscularly on Day 10 to induce luteal regression. At 12 h after PGF2α administration, intravenous administration of vehicle or 35 nmol (50 μg)/head of TAK-683 was performed in control (n = 4) and treatment (n = 4) groups, respectively. Blood samples were collected at 6-h intervals for 96 h and then daily until the detection of subsequent ovulation (second ovulation). After the second ovulation, ultrasound examinations and blood sampling were performed every other day or daily until the subsequent ovulation (third ovulation). Mean concentrations of LH and FSH in the treatment group were significantly higher 6 h after TAK-683 treatment than those in the control group (12.0 ± 10.7 vs 1.0 ± 0.7 ng/ml for LH, 47.5 ± 28.2 vs 15.1 ± 3.4 ng/ml for FSH, p < 0.05), whereas mean concentrations of oestradiol in the treatment group decreased immediately after treatment (p < 0.05) as compared with the control group. Ovulation tended to be delayed (n = 2) or occurred early (n = 1) in the treatment group as compared with the control group. For the second ovulation, ovulatory follicles in the treatment group were significantly smaller in maximal diameter than in the control group (3.8 ± 0.5 vs 5.4 ± 0.2 mm, p < 0.05, n = 3). Administration of TAK-683 in the follicular phase stimulates gonadotropin secretion and may have resulted in ovulation of premature follicles in goats.

Pharmacologic application of native GnRH in the winter anovulatory mare, I: Frequency of reversion to the anovulatory state following ovulation induction and cessation of treatment

The continuous, subcutaneous infusion of native GnRH into seasonally anovulatory mares stimulates the synthesis and secretion of LH without pituitary refractoriness, offering opportunities to markedly accelerate the timing of ovulation within the operational breeding season. Herein, we tested the hypothesis that ovarian cycles induced in winter anovulatory mares using continuous administration of native GnRH for 28 days, beginning in either early February or early March (North America) would not revert to an anovulatory state after treatment withdrawal. Anovulatory mares received sham pumps (control) or native GnRH (100 μg/h) for 28 days beginning from February 2 or 3 (GnRH-Feb) or March 2 or 3 (GnRH-Mar). Mean concentrations of LH were five- to seven-fold greater during February in the GnRH-Feb group compared with control and GnRH-Mar groups through February and ending on March 2 or 3. However, concentrations of LH returned to the winter baseline within 3 to 11 days after pump removal and all GnRH-Feb mares failed to remain cyclic after treatment withdrawal. Correspondingly, during March, concentrations of LH in the GnRH-Mar group were greater (P < 0.001) than in the control and GnRH-Feb groups during the 28-day treatment period. Follicular growth and frequency of ovulation (6/10 GnRH-Feb; 9/10 GnRH-Mar, 1/11 controls, respectively) were greater (P < 0.01) in GnRH-treated mares. Ovulatory cycles continued in five of nine GnRH-Mar mares that ovulated, with interovulatory intervals of 15 to 24 days; whereas, three of nine mares had extended (33–42 days) interovulatory intervals and one of nine mares had a persistent CL after cessation of treatment. In summary, continuous administration of native GnRH for 28 days, beginning in early February or March, elevated circulating LH adequately to stimulate follicular growth and ovulation up to 60 days earlier than in untreated controls. However, if continuous, subcutaneous infusion of GnRH is selected as the only pharmacologic or managerial intervention, and mares are not pregnant, treatment must be continued at least until the end of March. This will improve the likelihood of a normal interovulatory interval after treatment withdrawal.