Highly sensitive second-antibody enzyme immunoassay for determination of estradiol-17beta concentration in blood plasma of the mithun (Bos frontalis)

The objective of the present study was to develop and validate a simple, sensitive, quick and economic enzyme immunoassay (EIA) for estradiol-17beta (E2) in mithun (Bos frontalis) plasma on microtiter plates using a second-antibody coating technique and hormone-horseradish peroxidase as a label. For the assay, the wells of microtiter plates were coated with affinity-purified goat anti-rabbit IgG that binds the hormone-specific antibody. One milliliter of mithun plasma was extracted using benzene and 50 microl of 300 microl volume reconstituted with assay buffer was run in the assay along with standards ranging from 0.10-100 pg/well prepared in assay buffer. The sensitivity of the assay was 0.72 pg/ml. The intra- and inter-assay coefficients of variation were below 10%, and the extraction efficiency was >93%. Linearity of recovery of the added hormone concentrations was recorded. The assay developed was further validated biologically by estimating the hormone concentrations in six female and five male mithun calves, 12 cyclic mithuns for the entire reproductive cycle, and four pregnant mithun cows. The EIA developed can estimate low concentrations of E2 (2.2-5.2 pg/ml) in growing calves as well as very high concentrations of the hormone during pregnancy (E2=85.6-143.5 pg/ml). Apart from being non-radioactive, the assay developed has several advantages over conventional radioimmunoassays: it is more sensitive, less labor intensive, simpler to perform, and less time consuming. In conclusion, the EIA procedure described herein is sufficiently reliable, economic, safe, quick and sensitive to estimate the hormone at all physiological levels in bovine plasma.     

Chronic stress, hormone profiles and estrus intensity in dairy cattle

The objectives of the present study were to determine if lameness, a model for a natural chronic stressor, affects hormone concentrations in milk prior to estrus and/or the subsequent expression of estrus in the postpartum period. Dairy cows>20 days postpartum were scored for lameness and observed for estrus intensity using a weighted scoring system (>100 points=estrus=Day 0). Increasing lameness score was not associated with daily profiles of milk progesterone (throughout Days -18 to 0), estradiol (Days -6 to 2) or cortisol (Days -18 to 2) around estrus, maximum estradiol values or estradiol concentrations on Day 0. However, post hoc pair wise comparisons revealed that prior to estrus, severely lame cows had lower maximum progesterone concentrations compared to nonlame cows (1.3+/-0.1, 1.2+/-0.2, 0.7+/-0.1 ng/ml milk; P=0.042). Furthermore, severely lame cows expressed behavioral estrus with lower intensity (284+/-128 points, n=9) compared to moderately lame (662+/-310 points, n=9) or nonlame animals (583+/-275 points, n=18; P=0.05 and P=0.02, respectively). Resting concentrations of cortisol (Days 20-80 postpartum) did not vary between days postpartum or lameness score. The incidence of behavioral estrus was not affected by increasing lameness score, as 54.2%, 56.2% and 50.0% periods with low progesterone were associated with spontaneous estrus expression, respectively. Concluding, in this biological model of chronic stress, lameness did not affect the incidence of behavioral estrus but did reduce estrus intensity once ovarian cyclicity had resumed after calving. This reduced intensity of estrus was associated with lower maximum progesterone values prior to estrus but not abnormal daily cortisol or estradiol values in milk.     

Relationship between plasma growth hormone concentrations and temperament in mithuns (Bos frontalis)

The aim of the present study was to verify whether or not plasma growth hormone (GH) concentrations are correlated with temperament in mithuns (Bos frontalis), a semiwild ruminant. Therefore, a total of 69 female mithuns from four different strains, viz., Arunachal, Nagaland, Mizoram, and Manipur, were divided into six age groups (Group I, 0-6 months; Group II, >6-12 months; Group III, >1-2 years; Group IV, >2-2.5 years; Group V, >2.5-3.0 years; Group VI, >3.0 years). Blood samples were collected weekly for 6 consecutive weeks and assayed for plasma GH. Temperament was scored on a 6-point scale, 6 were being very aggressive and 1 docile. Body weights of all animals were recorded once a week for 6 consecutive weeks GH concentrations and temperament scores were found to differ significantly between groups. Strain had significant effects on blood GH levels and temperament. Blood GH concentrations and temperament of Manipur mithuns were significantly higher than those of the other three strains within each group, for all groups. Across groups III to VI, blood GH levels and temperament among Nagaland, Mizoram, and Arunachal mithuns did not differ. Overall, the strain with the highest blood GH concentrations also had highest temperament scores. The Manipur strain had the highest blood GH levels and exhibited the most aggression (r = 0.95), and Arunachal mithuns, the lowest (r = 0.93). Temperament scores tended to decrease with increasing age for all four strains. Coefficients of correlation between blood GH and temperament among strains within each group, for all groups, were found to be significant. The highly positive correlation (r = 0.94) between blood GH concentrations and temperament for all animals, regardless of age and strain differences, clearly indicates the relationship between blood GH and temperament in mithuns. In conclusion, our results suggest that peripheral blood GH levels can influence temperament in mithuns.     

Infradian rhythmicity in lactogenic hormone (prolactin,growth hormone,cortisol and thyroid hormone) secretion throughout the lactation cycle in mithun cows (Bos frontalis): variation among strains

The role of lactogenic hormones (prolactin, growth hormone, cortisol and thyroid hormone) on lactation yield in Mithun cows as well as their rhythmicity throughout the lactation cycle were studied in Mizoram (n = 4) and Nagaland (n = 7) strain of mithun (Bos frontalis). Blood samples were collected from all the animals from the day of calving to the complete dry off at an interval of 15 days. All the hormones were estimated in the serum by commercially available ELISA kits. Plasma level of cortisol (μg/dl), growth hormone (GH, in ng/ml), prolactin (PRL, in μIU/ml), triiodothyronine (T₃, in nmol/μl) and thyroxin (T₄, in ng/ml) were 20.84 ± 0.29, 28.08 ± 0.56, 9.87 ± 0.20, 27.82 ± 0.56 and 51.33 ± 0.48, respectively, in mithun irrespective of strains during the lactation period. Levels of all the hormones varied significantly (p ≤ 0.01) during different days of lactation cycle but, there was no significant difference among strain. Levels of PRL, GH, cortisol and T₃ were significantly (p < 0.01) higher around calving and declined sharply. The hormones remained in almost steady state during mid-lactation and declined during late lactation. All the hormones stated above were positively correlated with lactational yield thus their role on lactogenesis and galactopoiesis was established.     

Importance of estrus on pregnancy per insemination in suckled Bos indicus cows submitted to estradiol/progesterone-based timed insemination protocols

The objective was to evaluate the effect of estrus occurrence (based on removal of tail-head marks) on ovarian responses and pregnancy per AI (P/AI; 30 d after AI) in suckled Bos indicus beef cows submitted to timed AI (TAI) protocols. Cows received an intravaginal device containing 1.0 g progesterone, and 2.0 mg estradiol benzoate im; 8 d later, the intravaginal device was removed, and they were given PGF_2α (0.25 mg of cloprostenol sodium) and 300 IU of eCG, with TAI 48 to 52 h later. In Experiment 1, cows were assigned to receive one of three treatments: 1 mg of estradiol cypionate (ECP) im at progesterone (P4) device removal (N = 178); 10 μg of GnRH im at TAI (N = 190); or both treatments (N = 172). In cows given estradiol (ECP or ECP + GnRH), more displayed estrus (P = 0.002) and became pregnant (P < 0.0001) compared with those receiving only GnRH. In Experiment 2, the effect of the occurrence of estrus on ovarian responses was evaluated in cows (N = 53) synchronized using ECP at device removal. Cows that displayed estrus had a greater diameter of the largest follicle (LF) at device removal (P < 0.0001), a greater diameter at TAI (P < 0.0001), a greater ovulation rate (P = 0.02), a larger CL (P = 0.02), and a greater P4 concentration (P < 0.0001) than cows that did not display estrus. In Experiment 3, the effect of GnRH treatment on P/AI at TAI was evaluated in cows that received ECP at device removal, and either displayed, or did not display, estrus (N = 726). There was no estrus by GnRH interaction (P = 0.22); the P/AI was greater (P < 0.0001) in cows that displayed estrus (61.9%) than cows that did not display estrus (41.4%). However, GnRH did not improve (P = 0.81) P/AI (GnRH = 53.7% vs. no GnRH = 52.6%). In conclusion, exogenous estradiol at device removal increased both the proportion of suckled Bos indicus cows that displayed estrus and P/AI. Cows that displayed estrus had better ovarian responses (i.e., larger follicles at TAI, a greater ovulation rate, larger CL, and greater P4 concentrations) following an estradiol/P4-based synchronization protocol. Although occurrence of estrus improved pregnancy outcomes, GnRH at TAI did not improve P/AI in suckled Bos indicus cows treated with ECP, regardless of estrus occurrence.     

Serum estradiol-17beta concentrations during spontaneous silent estrus and after prostaglandin treatment in the mare

Serum estradiol-17beta concentrations were determined during silent estrus in the mare. Relationships between serum estradiol-17beta concentration, corpus luteum regression, follicular development, ovulation, prostaglandin treatment and behavioral estrus were investigated. The expression of behavioral estrus was found to be related to the patterns of progesterone and estradiol-17beta secretion during the periovulatory period. When compared to normal estrous cycles, silent estrus was accompanied by a significantly lower maximum serum estradiol-17beta concentration (47.8 vs 34.6 pg/ml), a significantly longer interval from maximum estradiol-17beta concentration to ovulation (1.7 vs 4.0 days), and a significantly shorter interval from corpus luteum regression to ovulation (5.3 vs 2.8 days). Silent estrus following prostaglandin treatment was related to a significantly shorter interval from prostaglandin treatment to ovulation (3.6 +/- 0.4 days) than from normal corpus luteum regression to ovulation (5.3 +/- 0.3 days). Silent estrus appeared to be related to changes in follicular estradiol-17beta secretion and to the pattern of its secretion as related to regression of the corpus luteum. There appeared to be not only less estradiol-17beta present, but also less time available after luteal regression for it to interact with the central nervous system to elicit the changes necessary to cause behavioral estrus. There fore, unusual relationships between luteal function and folliculogenesis can result in one type of silent estrus. Significant correlations (P<0.05) were found between follicle size and serum estradiol-17beta concentration whenever behavioral estrus occurred [follicle diameter in mm = 0.96 (serum estradiol-17beta in pg/ml) + 6.08 and 0.73 (serum estradiol-17beta + 13.32 for control and normal estrus following prostaglandin treatment groups, respectively]. During silent estrus, however, no significant correlations between follicle size and serum estradiol-17beta concentration were observed.     

Effect of induced suprabasal progesterone levels around estrus on plasma concentrations of progesterone, estradiol-17beta and LH in heifers

A controlled study was carried out to investigate the effects of suprabasal plasma progesterone concentrations on blood plasma patterns of progesterone, LH and estradiol-17beta around estrus. Heifers were assigned to receive subcutaneous silicone implants containing 2.5 g (n=4), 5 g (n=4), 6 g (n=3), 7.5 g (n=3) or 10 g (n=4) of progesterone, or implants without hormone (controls, n=5). The implants were inserted on Day 8 of the cycle (Day 0=ovulation) and left in place for 17 d. The time of ovulation was determined by ultrasound scanning. Blood was collected daily from Days 0 to 14 and at 2 to 4-h intervals from Days 15 to 27. Control heifers had the lowest progesterone concentrations on Days 20.5 to 21 (0.5 +/- 0.1 nmol L(-1)); a similar pattern was observed in heifers treated with 2.5 and 5 g of progesterone. In the same period, mean progesterone concentrations in the heifers treated with 6, 7.5 and 10 g were larger (P < 0.05) than in the controls, remaining between 1 and 2.4 nmol L(-1) until implant removal. A preovulatory estradiol increase started on Days 16.4 to 18.4 in all the animals. In the controls and in heifers treated with 2.5 and 5 g of progesterone, estradiol peaked and was followed by the onset of an LH surge. In the remaining treatments, estradiol release was prolonged and increased (P < 0.05), while the LH peak was delayed (P < 0.05) until the end of the increase in estradiol concentration. The estrous cycle was consequently extended (P < 0.05). In all heifers, onset of the LH surge occurred when progesterone reached 0.4 to 1.2 nmol L(-1). The induction of suprabasal levels of progesterone after spontaneous luteolysis caused endocrine asynchronies similar to those observed in cases of repeat breeding. It is suggested that suprabasal concentrations of progesterone around estrus may be a cause of disturbances oestrus/ovulation.     

社群压力对圈养雌性亚洲象动情的影响

亚洲象是中国一级濒危保护动物,改善圈养亚洲象的繁殖状况是亚洲象保护工作的重要内容之一。本研究旨在通过分析社群压力对圈养雌性亚洲象应激和动情状况的影响,从而探究圈养雌性亚洲象长期不动情的原因。通过放射性免疫法检测雌性亚洲象尿液中皮质醇、孕酮和雌二醇水平以及粪便中皮质醇和雌二醇水平,以雌激素水平反映亚洲象的动情状况,以皮质醇水平反映亚洲象的应激状态,以争斗行为和友好行为的发生情况反映圈养群体的社群压力。研究发现：(1)处于优势地位的个体表现出更多的斗争行为,处于从属地位的个体受到更多的攻击行为;(2)优势个体的皮质醇水平显著高于处于从属地位个体的皮质醇水平;(3)优势个体的雌二醇和孕酮具有小幅周期性波动,而从属个体的雌二醇和孕酮水平没有表现出明显的波动。从本研究结果可以看出圈养亚洲象群体中优势个体对从属个体的行为抑制可能是导致从属个体雌激素水平低,长期不能参与繁殖的原因。     

大额牛产犊行为观察

采用直接观察法对大额牛(Bos frontalis)的产前离群与产后带犊归群、分娩等产犊行为进行了观察。结果表明,(1)野外观察17头临产母牛中,76.47%(13头)的个体在产前(11.23±7.14)d离开社群独行觅食,寻找分娩场所;分娩地点多选择在靠近溪河、地势较高、食物丰富、光照好、地面相对平坦的草地、林边或林中。母牛野外产犊后带犊归群的时间为产后(5.57±2.64)d。(2)在设围栏的人工草地上放牧条件下对6头母牛分娩行为观察表明,产前24～12 h母牛的警觉巡视行为明显增多;产前12～5h表现为烦燥不安和频繁爬跨;产前3～1 h领地行为明显,对分娩地点4～6 m2内长势较高的牧草等进行清理,为新生犊牛行动清除障碍;分娩时间为6:00～18:00时,其中在6:00～7:00时分娩的占66.67%(4/6);母牛从卧地进入分娩至犊牛产出期间起卧(7.00±6.00)次,从卧地到犊牛完全产出需(25.00±5.00)min,胎衣排出的时间为产后(5.64±1.80)h;犊牛(n=6,♂1+♀5)初生体重为(16.46±2.56)kg,母牛无难产现象;母牛对犊牛能及时舔饰,定时授乳和用心守护。犊牛从产出至第一次站稳需(52.70±29.69)min,第一次站稳到吮吸初乳需(38.7±29.14)min,第一次吃初乳到向周边活动需(61.33±1.53)min。     

独龙牛遗传多样性及其种群遗传结构的等位酶分析

采用水平片淀粉凝胶电泳技术,进行３０头独牛牛４１种蛋白质共计４４个遗传座位的等位酶分析,只在Ｔｒ,Ｈｐ,Ａｍｙ,Ｅｓｔ等４个座位发现多态性。每个座位等基因的平均数、多态座位百分比和平均杂合度值分别为Ａ＝１．０９０９、Ｐ＝０．０６８２和Ｈ＝０．０２６２。贡山县和福贡县独龙牛群体从酶基因的角度上看遗传多样性贫乏,可能是分别由小种群引种而来,受到瓶颈效应的作用,并伴随着创立者事件的发生。我们结合独龙牛在     

Effect of norgestomet on peripheral levels of progesterone and estradiol-17beta in beef cows

Peripheral levels of progesterone and estradiol 17beta were quantified in 27 cycling cows following administration of a single Hydron ear implant (G. D. Searle and Co.) containing 2, 4 or 6 mg norgestomet or controls which received no implant. Implants were inserted subcutaneously in the ear on day 15 of the estrous cycle (day of estrus = day 0) and removed 9 days later. The 4 mg (seven of seven cows) and 6 mg (six of six cows) implants suppressed estrus; however, three of eight cows in the 2 mg group exhibited estrus prior to implant removal. The 6 mg implant group had a significantly longer interval from implant removal to estrus than either the 2 or 4 mg group. Failure to detect differences in the rate at which progesterone declined indicated norgestomet treatment did not affect normal corpus luteum regression. Estradiol levels rose at a similar rate approaching estrus in all treatments. There was no indication of increased endogenous estradiol levels due to norgestomet treatment.     

Diurnal changes in blood metabolites and their relation to plasma growth hormone and time of feeding in mithun heifers (Bos frontalis)

The aim of the present study was to investigate what, if any, diurnal changes occur in blood metabolites in relation to plasma growth hormone (GH) and feeding time among mithun (Bos frontalis), a semi-wild ruminant. Blood samples were collected at hourly intervals during a 24 h span from 6 mithun heifers (averaging 2.5 yr of age and averaging 230 kg in weight) that were fed twice a day at 11:00 and 16:00 h. Samples were assayed for plasma GH and blood metabolites, non-esterified fatty acids (NEFA), glucose, and alpha-amino nitrogen. The total sampling period was divided into a 1) postprandial (after meal) period (period I: 11:00 to 21:00 h) and 2) interprandial period (period II: 22:00 to 10:00 h) and also into night (20:00 to 05:00 h) and day (06:00 to 10:00 h) periods for statistical analysis. Plasma glucose and alpha-amino nitrogen levels increased (p<0.01), and plasma NEFA and GH decreased (p<0.01) after each meal. No diurnal rhythmicity was detected in plasma glucose or alpha-amino nitrogen levels. Interestingly, plasma NEFA and GH levels were higher (p<0.01) during the interprandial (period II) and night periods, indicating an energy deficit that occurred progressively during the interprandial period of nocturnal feed deprivation. In twice-daily-fed mithuns we conclude that: 1) plasma metabolites and GH exhibited a definite pattern of change with time of feeding; 2) concentrations of plasma NEFA were higher nocturnally due to an energy deficit and that GH levels were higher during the interprandial period after the second meal; 3) the interprandial period after the second feeding may be considered to constitute a short-term food deprivation; 4) the longer interprandial period of 19 h in this study between the second and subsequent morning meal may be changed into equally divided feedings to minimize the short-term energy deficit; and 5) blood sampling for blood metabolites in mithuns should be conducted at a fixed time of day with special emphasis on time of feeding.     

Simultaneous quantification of progesterone, estrone, estradiol-17 beta and corticosterone in female American kestrel plasma

Progesterone, estrone, estradiol-17 beta and corticosterone were quantified simultaneously for the first time in female American kestrel (Falco sparverius) plasma. A mean level for each hormone was determined for the laying and non-laying periods of the summer (April-September), and for February. Means were comparable to those of other wild avian species and were significantly higher for the laying period than for the other 2 periods. The mean corticosterone level for February was higher than that for the non-laying summer period. Plasma from laying kestrels, unlike that from other avian species, required lipid removal before column chromatography. Of 2 lipid removal techniques compared, i.e. the cold methanol and hexane:methanol techniques, the latter proved superior.     

The effect of active immunization against progesterone on plasma concentrations of total and free progesterone, estradiol-17beta and LH in the cyclic ewe

Nine mature cyclic ewes were actively immunized against progesterone which was rendered immunogenic by conjugation to bovine serum albumin (BSA). Seven control ewes were immunized with BSA. In ewes immunized against progesterone, the concentration of total plasma progesterone increased to 24.3 ng/ml vs 2.8 ng/ml in control animals (P<0.001). However, immunization did not affect the plasma levels of free, unbound progesterone. The correlation coefficient between total plasma progesterone concentrations on Days 4 to 11 of the estrous cycle and antibody titer was r=0.983. Estradiol-17beta concentrations in immunized ewes were higher than in controls on Days 6 to 15 of the estrous cycle (P approximately 0.05). Frequent sampling for LH over a 6-h period on Days 2, 5, 8, 11 and 14 of the cycle revealed no significant differences in the frequency and amplitude of LH pulses between immunized and control ewes. The immunized ewes had estrous cycles of normal length and maintained normal pregnancies. It is suggested that the immunized cyclic ewe is capable of maintaining adequate levels of free progesterone by greatly increasing progesterone synthesis, thus neutralizing the effect of the antibodies.     

笼养雌性黑叶猴尿中性腺激素水平变化与性行为、等级序位的关系

2003年10月30日至2004年11月21日,采集了3只笼养雌性黑叶猴妊娠期和1 只笼养雌性黑叶猴发情期的晨尿,并用放射免疫分析法检测尿液中孕酮和雌二醇浓度,同时用全事件观察法观察黑叶猴的行为,分析雌性黑叶猴的行为与其尿中的性腺激素的关系。研究结果发现:1)进入妊娠期,雌性黑叶猴尿中的雌二醇浓度逐渐上升,中后期达高峰;而尿中孕酮浓度在妊娠初期仍维持在放免法的灵敏度以下,妊娠约1 个月后才升高至灵敏度以上(13.39 ng/ mgCr);2)在妊娠中期,雌性黑叶猴仍维持着较高频率的邀配行为,但与雌激素的变化无明显的相关关;3)发情期,雌猴尿中的雌二醇浓度与其邀配行为呈正相关;4)雌性黑叶猴的性激素高低与其等级序位无关,但序位高的雌猴繁殖较早。     

Diurnal Changes in Blood Metabolites and Their Relation to Plasma Growth Hormone and Time of Feeding in Mithun Heifers (Bos frontalis)

The aim of the present study was to investigate what, if any, diurnal changes occur in blood metabolites in relation to plasma growth hormone (GH) and feeding time among mithun (Bos frontalis), a semi‐wild ruminant. Blood samples were collected at hourly intervals during a 24 h span from 6 mithun heifers (averaging 2.5 yr of age and averaging 230 kg in weight) that were fed twice a day at 11:00 and 16:00 h. Samples were assayed for plasma GH and blood metabolites, non‐esterified fatty acids (NEFA), glucose, and alpha‐amino nitrogen. The total sampling period was divided into a 1) postprandial (after meal) period (period I: 11:00 to 21:00 h) and 2) interprandial period (period II: 22:00 to 10:00 h) and also into night (20:00 to 05:00 h) and day (06:00 to 10:00 h) periods for statistical analysis. Plasma glucose and alpha‐amino nitrogen levels increased (p<0.01), and plasma NEFA and GH decreased (p<0.01) after each meal. No diurnal rhythmicity was detected in plasma glucose or alpha‐amino nitrogen levels. Interestingly, plasma NEFA and GH levels were higher (p<0.01) during the interprandial (period II) and night periods, indicating an energy deficit that occurred progressively during the interprandial period of nocturnal feed deprivation. In twice‐daily‐fed mithuns we conclude that: 1) plasma metabolites and GH exhibited a definite pattern of change with time of feeding; 2) concentrations of plasma NEFA were higher nocturnally due to an energy deficit and that GH levels were higher during the interprandial period after the second meal; 3) the interprandial period after the second feeding may be considered to constitute a short‐term food deprivation; 4) the longer interprandial period of 19 h in this study between the second and subsequent morning meal may be changed into equally divided feedings to minimize the short‐term energy deficit; and 5) blood sampling for blood metabolites in mithuns should be conducted at a fixed time of day with special emphasis on time of feeding.     

Influence of lameness on follicular growth, ovulation, reproductive hormone concentrations and estrus behavior in dairy cows

The objective of this study was to examine the effect of a chronic stressor, lameness, on reproductive parameters. Seventy cows 30-80 days post-partum were scored for lameness and follicular phases synchronized with GnRH followed seven days later by prostaglandin (PG). Fifteen Lame animals did not respond to GnRH ovarian stimulation. Milk progesterone for 5 days prior to PG was lower in the remaining Lame cows than Healthy herdmates. Fewer Lame cows ovulated (26/37 versus 17/18; P = 0.04) and the interval from PG to ovulation was shorter in Lame cows. In Subset 1 (20 animals), the LH pulse frequency was similar in ovulating animals (Lame and Healthy) but lower in Lame non-ovulators. An LH surge always preceded ovulation but lameness did not affect the interval from PG to LH surge onset or LH surge concentrations. Before the LH surge, estradiol was lower in non-ovulating cows compared to those that ovulated and estradiol concentrations were positively correlated with LH pulse frequency. In Subset 2 (45 cows), Lame ovulating cows had a less intense estrus than Healthy cows, although Lame cows began estrus and stood-to-be-mounted earlier than Healthy cows. In conclusion, we have identified several parameters to explain poor fertility in some chronically stressed animals. From 30 to 80 days post-partum, there was a graded effect that ranged from 29% Lame cows with absence of ovarian activity, whereas another 21% Lame cows failed to express estrus or ovulate a low estrogenic follicle; in 50% cows, many reproductive parameters were unaffected by lameness.     

Prediction of estrus cyclicity in Asian elephants (Elephas maximus) through estimation of fecal progesterone metabolite: development of an enzyme-linked immuno-sorbent assay

Asian elephants (Elephas maximus), prominent “flagship species”, are listed under the category of endangered species (EN - A2c, ver. 3.1; IUCN Red List 2009) and there is a need for their conservation. This requires understanding demographic and reproductive dynamics of the species. Monitoring reproductive status of any species is traditionally being carried out through invasive blood sampling and this is restrictive for large animals such as wild or semi-captive elephants due to legal, ethical, and practical reasons. Hence, there is a need for a non-invasive technique to assess reproductive cyclicity profiles of elephants, which will help in the species’ conservation strategies. In this study, we developed an indirect competitive enzyme linked immuno-sorbent assay (ELISA) to estimate the concentration of one of the progesterone-metabolites i.e., allopregnanolone (5α-P-3OH) in fecal samples of Asian elephants. We validated the assay which had a sensitivity of 0.25 μM at 90% binding with an EC₅₀ value of 1.37 μM. Using female elephants, kept under semi-captive conditions in the forest camps of Mudumalai Wildlife Sanctuary, Tamil Nadu and Bandipur National Park, Karnataka, India, we measured fecal progesterone-metabolite (5α-P-3OH) concentrations in six animals and showed their clear correlation with those of serum progesterone, measured by a standard radio-immuno assay. Statistical analyses using a Linear Mixed Effect model showed a positive correlation (P < 0.1) between the profiles of fecal 5α-P-3OH (range: 0.5-10 μg/g) and serum progesterone (range: 0.1-1.8 ng/mL). Therefore, our studies show, for the first time, that the fecal progesterone-metabolite assay could be exploited to predict estrus cyclicity and to potentially assess the reproductive status of captive and free-ranging female Asian elephants, thereby helping to plan their breeding strategy.     

Genotype/age interactions on aggressive behavior in gonadally intact estrogen receptor beta knockout (betaERKO) male mice

Estrogen, as an aromatized metabolite of testosterone, has a facilitatory effect on male aggressive behavior in mice. Two subtypes of estrogen receptors, alpha (ER-alpha) and beta (ER-beta), in the brain are known to bind estrogen. Previous studies revealed that the lack of ER-alpha gene severely reduced the induction of male aggressive behavior. In contrast, mice that lacked the ER-beta gene tended to be more aggressive than wild type (WT) control mice, although the behavioral effects of ER-beta gene disruption were dependent on their social experience. These findings lead us to hypothesize that estrogen may facilitate aggression via ER-alpha whereas it may inhibit aggression via ER-beta. In the present study, we further investigated the role of ER-beta in the regulation of aggressive behavior by examining developmental changes starting at the time of first onset, around the age of puberty. Aggressive behaviors of ER-beta gene knockout (betaERKO) mice were examined in three different age groups, puberty, young-adult, and adult. Each mouse was tested every other day for three times in a resident-intruder paradigm against olfactory bulbectomized intruder mice and their trunk blood was collected for measurements of serum testosterone after the completion of the study. Overall, betaERKO mice were significantly more aggressive than WT. These genotype differences were more pronounced in puberty and young adult age groups, but not apparent in the adult age group, in which betaERKO mice were less aggressive than those in two younger age groups. Serum testosterone levels of betaERKO mice were significantly higher than those of WT mice only in the pubertal age group, but not in young adult (when betaERKO mice were still significantly more aggressive than WT mice) and adult (when no genotype differences in aggression were found) age groups. These results suggest that ER-beta mediated actions of gonadal steroids may more profoundly be involved in the inhibitory regulation of aggressive behavior in pubertal and young adult mice.