Estimation of salivary and serum alkaline phosphatase level as a diagnostic marker in type-2 diabetes mellitus with periodontal health and disease: A clinico-biochemical study.

INTRODUCTION: Chronic periodontitis is a multifactorial disease resulting in the inflammation and destruction of the supporting structures around the teeth, leading to tooth mobility and subsequent loss of tooth. Metabolic disorders, such as diabetes mellitus, play a crucial role in the progression of periodontal inflammatory conditions. Alkaline phosphatase (ALP) enzyme plays a key role in gingival inflammation and bone resorption. Hence, the aim of the present study is to compare the serum and salivary alkaline phosphatase levels in chronic periodontitis patients with or without type-2 diabetes mellitus. MATERIALS AND METHODS: A total of 45 individuals were included in the study and divided into three groups: Group I (healthy individual), Group II (Chronic periodontitis without diabetes mellitus type-2) and Group III (Chronic periodontitis with type-2 diabetes mellitus) on the basis of clinical, radiographic and blood sugar examination. The serum and unstimulated saliva were collected from all patients in aseptic condition and samples were analyzed for alkaline phosphatase level using AVANTOR™ Benesphera ALP Kit by fully automated analyzer. RESULTS: The result showed that the concentration of serum and salivary alkaline phosphatase increases significantly in patients with chronic periodontitis with type-2 diabetes mellitus than chronic periodontitis without diabetes mellitus and healthy patients. CONCLUSION: We can conclude that alkaline phosphatase can be used as a key inflammatory diagnostic biomarker in periodontal diseases.

Responsiveness of prepubertal crossbred bull calves to exogenous GnRH and its impact on reproductive hormones under tropical conditions.

This study investigated the age related variations in luteinizing hormone (LH), androstenedione, testosterone, and total estrogens response to exogenous gonadotropin-releasing hormone (GnRH) in Holstein-Friesian (HF) × Tharparkar bull calves. Fifteen bull calves were selected and, based on their age, were divided into Group I (14-16 months, n = 5), Group II (9-12 months, n = 5), and Group III (6-8 months, n = 5). All bull calves were administered with 10 µg of GnRH intramuscularly. Blood samples were collected at an interval of 30 min commencing 1 h prior to GnRH treatment until 4 h post-GnRH treatment and thereafter, at an interval of 1 h for the next 3 h. Endocrine response in terms of pretreatment values, peak values, area under curve, and time taken to attain peak values for LH, androstenedione, testosterone, and total estrogens was evaluated in all the bull calves. Significant differences were observed in pretreatment values, peak concentrations, and area under curve for androstenedione and testosterone between the groups; with response being higher in Group I bull calves. The results indicated that the HF × Tharparkar bull calves of 14 months age and above respond to exogenous GnRH by secreting significant amounts of testosterone.

Development and Application of a Sensitive, Second Antibody Format Enzymeimmunoassay (EIA) for Estimation of Plasma FSH in Mithun (Bos frontalis).

Mithun (Bos frontalis) is a semi-wild rare ruminant species. A simple sensitive enzymeimmunoassay suitable for assaying FSH in the blood plasma of mithun is not available which thereby limits our ability to understand this species reproductive processes. Therefore, the aim of this article was to develop a simple and sensitive enzymeimmunoassay (EIA) for estimation of FSH in mithun plasma and apply the assay to understand the estrous cycle and superovulatory process in this species. To accomplish this goal, biotinylated FSH was bridged between streptavidin-peroxidase and immobilized antiserum in a competitive assay. Forty microlitre mithun plasma was used directly in the EIA. The FSH standards were prepared in hormone free plasma and ranged from 5-1280 pg/well/40 µL. The sensitivity of EIA was 5 pg/well FSH, which corresponds to 0.125 ng/mL plasma and the 50% relative binding sensitivity was 90 pg/well/40 µL. Although the shape of the standard curve was not influenced by different plasma volumes viz. 40 and 80 µL, a slight drop in the OD450 was observed with the increasing volume of plasma. Parallelism tests conducted between the endogenous mithun FSH and bovine FSH standards showed good homology between them. Plasma FSH estimated using the developed EIA and commercially available FSH EIA kit in the same samples were correlated (r = 0.98) and showed linearity. Both the Intra- and inter-assay CV were below 6%. Recovery of known concentrations of added FSH showed linearity (r = 0.99). The developed EIA was further validated biologically by estimating FSH in cyclic cows for the entire estrous cycle, in mithun heifers administered with GnRH analogues and in mithun cows during superovulatory treatment with FSH. In conclusion, the EIA developed for FSH determination in mithun blood plasma is simple and highly sensitive for estimation of mithun FSH in all physiological conditions.

Development and validation of a simple, sensitive enzyme immunoassay for quantification of androstenedione in bull plasma.

As an alternative to radioimmunoassay a simple and highly sensitive enzyme immunoassay (EIA) was developed and validated for androstenedione quantification in plasma of Karan Fries bulls using second antibody coating technique. The wells of the microtitreplate were coated with affinity-purified goat immunoglobulin (antirabbit IgG) that binds the hormone specific antibody. The EIA was performed to analyze androstenedione directly in 40 µl of bull plasma. The androstenedione standards ranged from 0.20 to 200 pg/40 µl /well and the sensitivity of the assay was 5 pg/ml plasma. Serially diluted bull plasma containing high endogenous androstenedione showed good parallelism with bovine androstenedione standard curve. Intra- and inter-assay coefficients of variation (CV) were found to be 8 and 9%, respectively. Peripheral plasma androstenedione concentrations determined in young and adult bull samples ranged between 104-990 pg/ml and 184-2040 pg/ml, respectively.

Comparison of endocrine changes, timing of ovulations, ovarian follicular growth, and efficacy associated with Estradoublesynch and Heatsynch protocols in Murrah buffalo cows (Bubalus bubalis).

Poor estrus expression and the difficulty encountered in predicting the time of ovulation compromise the reproductive efficiency of Murrah buffalo cows. Synchronization of ovulation and timed artificial insemination are able to precisely control the time of ovulation and thus avoid the need for estrus detection. Recently, the Estradoublesynch protocol (administration of a PGF2α injection 2 days before Heatsynch protocol; GnRH 0, PGF2α 7, estradiol benzoate [EB] 8) was developed that precisely synchronized ovulation twice, i.e., after GnRH and EB injections and resulted in satisfactory pregnancy rates in Murrah buffaloes. The present study was conducted on 104 cycling and 31 anestrus buffaloes to compare (1) the endocrine changes, timing of ovulations, ovarian follicular growth, and efficacy of Estradoublesynch and Heatsynch protocols in cycling and (2) the efficacy of Estradoublesynch and Heatsynch protocols for the improvement of fertility in cycling and anestrus Murrah buffalo cows. Ovulation was confirmed after all GnRH and EB treatments by ultrasonographic examination at 2-hour intervals. Plasma progesterone and total estrogen concentrations were determined in blood samples collected at daily intervals, beginning 2 days before the onset of protocols until the day of second ovulation detection. Ovulatory follicle size was measured by ultrasonography at six time points (first PGF2α administration of Estradoublesynch protocol every 2 days before the onset of Heatsynch protocol, GnRH administration of both protocols, 2 hours before ovulation detection after GnRH administration of both protocols, second PGF2α injection of Estradoublesynch protocol, PGF2α injection of Heatsynch protocol, EB injection of both protocols and, 2 hours before ovulation detection after EB administration of both protocols). Plasma LH, total estrogen, and progesterone concentrations were determined in blood samples collected at 30-minute intervals for 8 hours, beginning GnRH and EB injections, and thereafter at 2-hour intervals until 2 hours after the detection of ovulation. The first ovulatory rate was significantly higher (P<0.05) in the Estradoublesynch protocol (84.6%) than that in the Heatsynch protocol (36.4%). The first LH peak concentration (74.6±10.4 ng/mL) in the Estradoublesynch protocol was significantly higher (P<0.05) than that of the Heatsynch protocol (55.3±7.4 ng/mL). In Estradoublesynch protocol, the total estrogen concentration gradually increased from the day of GnRH administration coinciding with LH peak, and then gradually declined to the basal level until the time of ovulation detection. However, in Heatsynch protocol, the gradual increase in total estrogen concentration after GnRH administration was observed only in those buffalo cows, which responded to treatment with ovulation. In both Estradoublesynch and Heatsynch protocols, ovulatory follicle size increased by treatment with GnRH and EB until the detection of ovulation. The pregnancy rate after the Estradoublesynch protocol (60.0%) was significantly higher (P<0.05) than that achieved after the Heatsynch protocol (32.5%). Satisfactory success rate using the Estradoublesynch protocol was attributed to the higher release of LH after treatment with GnRH, leading to ovulation in most of the animals and hence creating the optimum follicular size at EB injection for ovulation and pregnancy to occur.

Comparative 2D-DIGE proteomic analysis of bovine mammary epithelial cells during lactation reveals protein signatures for lactation persistency and milk yield.

Mammary gland is made up of a branching network of ducts that end with alveoli which surrounds the lumen. These alveolar mammary epithelial cells (MEC) reflect the milk producing ability of farm animals. In this study, we have used 2D-DIGE and mass spectrometry to identify the protein changes in MEC during immediate early, peak and late stages of lactation and also compared differentially expressed proteins in MEC isolated from milk of high and low milk producing cows. We have identified 41 differentially expressed proteins during lactation stages and 22 proteins in high and low milk yielding cows. Bioinformatics analysis showed that a majority of the differentially expressed proteins are associated in metabolic process, catalytic and binding activity. The differentially expressed proteins were mapped to the available biological pathways and networks involved in lactation. The proteins up-regulated during late stage of lactation are associated with NF-κB stress induced signaling pathways and whereas Akt, PI3K and p38/MAPK signaling pathways are associated with high milk production mediated through insulin hormone signaling.

Endocrine changes, timing of ovulation, ovarian follicular growth and efficacy of a novel protocol (Estradoublesynch) for synchronization of ovulation and timed artificial insemination in Murrah buffaloes (Bubalus bubalis).

Experiments were conducted to investigate (1) ovarian follicular growth, timing of ovulation and associated endocrine changes (progesterone, estrogen, and LH) in cycling, and (2) efficacy in terms of pregnancy rate in cycling and anestrus Murrah buffaloes subjected to the Estradoublesynch protocol (prostaglandin F2α [PGF2α] 0, GnRH 2, PGF2α 9, estradiol benzoate, EB 10). Twelve cycling buffaloes were subjected to the Estradoublesynch protocol and observed for ovulation, follicle size, and endocrine changes after EB treatment. Ovulation occurred in 12 of 12 buffaloes (100%) at 48.5 ± 1.6 hours (range, 38.0-56.0) after EB treatment. Plasma LH, total estrogen, and progesterone concentrations were determined in intensive blood samples collected after EB treatment. Peak LH concentration of 34.2 ± 7.7 ng/mL (range, 17.8-178.5) occurred at 18.3 ± 0.8 hours (range, 14.0-22.0) after EB treatment. Peak total estrogen of 50.8 ± 6.9 pg/mL (range, 32.3-82.7) occurred 5.7 ± 1.0 hours (range, 2.0-14.0) after EB treatment. Follicle size was gradually increased from second PGF2α injection (9.7 ± 0.3 mm; range, 8.0-12.0) until ovulation was detected (12.9 ± 0.4 mm; range, 11.0-15.0). Fourteen cycling and 11 anestrus buffaloes were subjected to the Estradoublesynch protocol, with timed artificial insemination (TAI) 48 and 60 hours after EB treatment, and 58 cycling buffaloes were inseminated after spontaneous estrus (control group). Pregnancy rates were 62% for TAI of cycling buffaloes, 64% for anestrus buffaloes, and 34.5% for control group. Our observations demonstrated that the Estradoublesynch protocol followed by TAI satisfactory enhanced pregnancy rates in both cycling and anestrus buffaloes.

Development and validation of a sensitive enzyme immunoassay (EIA) for blood plasma cortisol in female cattle, buffaloes, and goats.

A highly sensitive enzyme immunoassay (EIA) that used the second antibody coating technique and the cortisol-horseradish peroxidase conjugate as a label for determination of free and total cortisol in blood plasma of dairy animals (cows, buffaloes, and goats) was developed. For biological validation of the EIA, blood samples were collected from the animals at 48 and 24 h before and 0, 12, 24, 36, 48, 60, 72, 84, 96, 108, 120, and 132 h after dexamethasone administration. The EIA was performed directly with 20 µL of fresh plasma (for free cortisol) and also with 20 µL of heat-treated plasma (for total cortisol) after 1:5 dilutions with PBS. Cortisol standards ranging from 0.39 to 200 pg/well/20 µL were used, and the sensitivity of the EIA procedure was found to be 0.39 pg/well/20 µL, which corresponded to 0.02 ng/mL. In comparison with RIA the EIA was at least 4 times more sensitive and required 5 times less cortisol antiserum. In female cattle, buffaloes, and goats, the total, free, and bound plasma cortisol before dexamethasone administration was significantly (P < 0.05) higher than the total, free, and bound cortisol after dexamethasone administration. It can be concluded from these studies that the direct, sensitive EIA validated for estimating the free and total cortisol concentrations was sufficiently reliable and quick for studying the dynamics of cortisol distribution in blood plasma of dairy animals.

Effect of supplementation of vitamin E, copper and zinc on the in vitro phagocytic activity and lymphocyte proliferation index of peripartum Sahiwal (Bos indicus) cows.

To study the effect of vitamin E (VE), copper (Cu) and zinc (Zn) supplementation on the in vitro phagocytic activity (PA) and lymphocyte proliferation response (LPR) of blood neutrophils and lymphocytes, thirty Sahiwal pregnant cows (six in each group) in their late gestation at 30 days before the expected date of calving were selected from the NDRI experimental herd and supplemented with various micronutrients from 30 days before calving to 45 days after calving. Cows were supplemented individually with VE (1000 IU/cow/day), Cu (20 ppm/cow/day) and Zn (80 ppm/cow/day) and also with a combination of VE, Cu and Zn to study cumulative effect of all micronutrients. One group without any supplementation acted as a control. Blood neutrophils and lymphocytes were isolated and studied for their PA and LPR. Supplementation of micronutrients like VE, Cu, Zn and a combination of all these nutrients significantly (p < 0.01) increased the PA of experimental cows as compared to control (unsupplemented) cows during the pre-partum period. During post-partum, all the micronutrients (VE, Cu, Zn and their combination) showed a significant (p < 0.01) increase in the PA of experimental cows as compared to control cows. Of all the groups, significant (p < 0.01) and maximum PA was observed in the combination group followed by Zn-supplemented group during both the pre- and post-partum period. A significant (p < 0.01) increase in LPR of B lymphocytes was observed in combination-supplemented group during the pre-partum period and during both the pre- and post-partum period in the Cu-supplemented group.

Selection of suitable reference genes for quantitative gene expression studies in milk somatic cells of lactating cows (Bos indicus).

We assessed the suitability of 9 internal control genes (ICG) in milk somatic cells of lactating cows to find suitable reference genes for use in quantitative PCR (qPCR). Eighteen multiparous lactating Sahiwal cows were used, 6 in each of 3 lactation stages: early (25 ± 5 d in milk), mid (160 ± 15 d in milk), and late (275 ± 25 d in milk) lactation. Nine candidate reference genes [glyceraldehyde 3-phosphate dehydrogenase (GAPDH), protein phosphatase 1 regulatory subunit 11 (PPP1R11), ß-actin (ACTB), ß-2 microglobulin (B2M), 40S ribosomal protein S15a (RPS15A), ubiquitously expressed transcript (UXT), mitochondrial GTPase 1 (MTG1), 18S rRNA (RN18S1), and ubiquitin (UBC)] were evaluated. Three genes, ß-casein (CSN2), lactoferrin (LTF), and cathelicidin (CAMP) were chosen as target genes. Very high amplification was observed in 7 ICG and very low level amplification was observed in 2 ICG (UXT and MTG1). Thus, UXT and MTG1 were excluded from further analysis. The qPCR data were analyzed by 2 software packages, geNorm and NormFinder, to determine suitable reference genes, based on their stability and expression. Overall, PPP1R11, ACTB, UBC, and GAPDH were stably expressed among all candidate reference genes. Therefore, these genes could be used as ICG for normalization of qPCR data in milk somatic cells through lactation.

The endocrine changes, the timing of ovulation and the efficacy of the Doublesynch protocol in the Murrah buffalo (Bubalus bubalis).

Experiments were conducted to investigate (a) the timing of ovulation and the associated endocrine changes (progesterone, estrogen and LH) during estrous cycle and (b) the efficacy, with respect to the pregnancy rate, in cycling and anestrus in Murrah buffaloes subjected to the Doublesynch protocol during the low breeding season. In experiment 1, 10 cycling buffaloes were administered PGF(2α) on day 0 (without regard to the estrous cycle stage), GnRH on day 2, a second PGF(2α) injection on day 9, and a second GnRH injection on day 11. Transrectal palpation was performed at 2-h intervals after the first and second GnRH treatments until ovulation was detected or for upto 96 h. The plasma progesterone and total estrogen concentrations were determined in blood samples collected at daily intervals starting 2 days before the onset of the protocol and continued until the day of the second detected ovulation. The plasma LH and total estrogen concentrations were measured in blood samples collected at 30-min intervals for 8h following the first and second GnRH injections and thereafter at 2-h intervals until 2h after the detection of ovulation. Ovulation occurred in 9/10 buffaloes (90%) at 22.2 ± 1.2 h (mean ± S.E.M.; range 18.0-26.0 h) and 10/10 buffaloes (100%) at 23.2 ± 1.0 h (mean ± S.E.M.; range 20.0-28.0 h) after the first and second GnRH treatments, respectively. The peak LH concentrations of 99.8 ± 28.5 ng/ml (range 37.8-320.0 ng/ml) and 62.3 ± 11.9 ng/ml (range 20.9-143.9 ng/ml) occurred 2.1 ± 0.3 h (range 1.0-3.5 h) and 2.3 ± 0.3 h (range 0.5-3.0 h) after the first and second GnRH treatments, respectively. The total estrogen concentration gradually increased from the day of both the first and second PGF(2α) administrations until the LH peak (with great variability) and then gradually declined to the basal level, which was reached at the time ovulation was detected. In experiment 2, 10 cycling and 11 non-lactating anestrus buffaloes were subjected to the Doublesynch protocol with timed artificial insemination (TAI) 16 and 24 h after the second GnRH treatment, and 55 cycling buffaloes were inseminated after spontaneous estrus was detected (control group). The pregnancy rates were 60% using TAI on cycling buffaloes (experiments 1 and 2), 55% for anestrus buffaloes (experiment 2), and 27.3% for cycling buffaloes inseminated following spontaneous estrus. The overall pregnancy success rates after the Doublesynch protocol in both cycling and anestrus buffaloes increased by 30.8% compared to spontaneous estrus (58.1% vs. 27.3%). In conclusion, the Doublesynch protocol effectively synchronized ovulation twice (after the first and second GnRH treatments) irrespective of the stage of estrous cycle in Murrah buffaloes. The study also demonstrated that the Doublesynch protocol followed by TAI significantly (P<0.005) enhanced the pregnancy rate in cycling and anestrus buffaloes in comparison to untreated controls during the low breeding season.

Influence of feeding Tinospora cordifolia peripartum on lactation parameters in crossbred cows.

For studying the effect of dietary supplementation of guduchi (Tinospora cordifolia) peripartum on lactation an investigation was conducted on 15 pregnant Karan Fries crossbred cows which were divided into two groups: treatment group of eight cows which were supplemented with guduchi at 60 g/day for 45 days prepartum and 120 g/day for 45 days postpartum; control group of seven pregnant cows which were not supplemented with guduchi. Jugular blood samples were collected from all cows during the periparturient period for analysis of various blood cell and plasma parameters. A significantly higher total leukocyte count, lymphocyte count, neutrophil count and neutrophil/lymphocyte ratio was recorded in the guduchi supplemented treatment group in comparison to untreated control cows throughout the experimental period. The increase of milk production over 305 days of lactation due to guduchi supplementation was significant (p < 0.05). A significant (p < 0.05) reduction in somatic cell count was also observed during the experimental period. Milk composition (fat, protein, lactose and SNF) was similar (p > 0.05) for both the groups. Plasma non esterified fatty acid (NEFA) concentrations were significantly higher (p < 0.01) in cows supplemented with guduchi throughout the course of study. Plasma concentration of growth hormone in the treated cows was also significantly higher beginning on the day of parturition up to 3 weeks postpartum (p < 0.05) in comparison to unsupplemented group.

Effects of supplementation of Tinospora cordifolia to crossbred cows peripartum.

Guduchi (Tinospora cordifolia), a medicinal plant used in ayurveda, is well documented for its immunomodulatory properties. Since the crossbred periparturient cow is highly susceptible to various diseases that effectively reduces its reproductive performance postpartum we explored the possibility of enhancing the reproductive performance of crossbred cows by guduchi supplementation peripartum. A total of 15 pregnant Karan Fries cows were selected and divided into two groups: treatment group of 8 cows which were supplemented with guduchi at 60 g/day for 45 days prepartum and 120 g/day for 45 days postpartum and unsupplemented control group of 7 cows. Jugular blood samples were collected from all cows during the periparturient period for analysis of endocrine (progesterone, total estrogens and PGFM), immunological and hematological parameters. Incidence of retention of fetal membranes, endometritis, pyometra and calf mortality were higher in control group of cows in comparison to those recorded in treated group. The guduchi supplemented cows exhibited faster uterine involution (28 days vs. 42 days) and early commencement of cyclicity (37 days vs. 58 days; based on plasma progesterone profiles) in comparison to untreated control group of cows. Mean birth weight of calves from treatment group of cows was significantly higher than those from control group however no significant difference was observed in average daily body weight gain of calves in both the groups. A higher total leukocyte, lymphocyte, neutrophil count along with increased neutrophil lymphocyte ratio was recorded in guduchi supplemented cows in comparison to untreated cows although plasma total antioxidant activity was similar between the two groups. Prepartum plasma progesterone concentration was significantly lowered in the treated group however there was no significant change in peripartum plasma total estrogens and PGFM levels due to guduchi supplementation.

Temporal changes in circulating levels of plasma interleukin-8 during peripartum period in Murrah buffaloes (Bubalus bubalis) under subtropical climate.

The objective of this study was to elucidate the changes in circulating levels of plasma interleukin-8 (IL-8) during peripartum period in Murrah buffaloes (Bubalus bubalis). IL-8 was estimated in blood plasma of healthy peripartum Murrah buffaloes (n=6) on days ± 30, ± 15, ± 5, ± 3, ± 1 and 0 pre- and postpartum with respect to the day of parturition (day 0) in each of the two different seasons (hot-humid and spring). The mean microclimate Temperature-Humidity Index (THI) during spring season was significantly lower (p<0.01) than the corresponding THI in hot-humid season. In both the seasons, plasma IL-8 remained lower in prepartum period (≤ 46.56 ± 14.08 pg/ml during spring and ≤ 73.18 ± 18.56 pg/ml during hot-humid season) than in the postpartum period (≥ 51.41 ± 13.82 pg/ml during spring and ≥ 84.13 ± 16.97 pg/ml in hot humid season). During spring, the IL-8 levels were significantly higher (P<0.05) on days+5 and +15 postpartum in comparison to the IL-8 levels on days -30, -5, and -3 prepartum. During hot-humid season, IL-8 level was significantly higher (P<0.05) on day +30 as compared to the IL-8 levels on days -30 and -5 prepartum. The correlation between IL-8 and mean microclimate THI was significant (r=0.25, P<0.01). From the results, it is concluded that peripartum period in buffaloes is associated with an inflammatory response leading to significantly higher plasma IL-8 during parturition and postpartum period than in the pre-partum period.

Divergent development of testosterone secretion in male zebu (Bos indicus) and crossbred cattle (Bos indicus x Bos taurus) and buffaloes (Bubalus bubalis) during growth.

Delayed pubertal development and low fertility of Bos indicus x Bos taurus crossbred male cattle and domestic buffaloes is hardly understood hence, a sensitive enzymeimmunoassay (EIA) was developed using the second antibody-coating technique and testosterone-3-O-carboxymethyloxime-horseradish peroxidase conjugate as a label for determination of testosterone in blood plasma. The EIA was validated by standard criteria. Blood samples were collected by venipuncture from growing male cattle (Karan Fries and Sahiwal) and buffalo (Murrah) and testosterone was estimated using the EIA procedure. Plasma testosterone concentrations increased significantly (P < 0.05) with advancing age. Testosterone concentrations were significantly (P < 0.01) higher in Sahiwal males in comparison to Karan Fries males. The low testosterone levels in crossbred than Sahiwal could imply that crossbred males have either not stabilized genetically or not adapted well in Indian climatic conditions resulting in poor libido and poor semen quality. The low testosterone levels in Murrah buffalo males may be the possible reason for delayed maturity in this species. The direct, sensitive EIA validated for estimating the plasma testosterone concentration was reliable for studying the testosterone profile in blood plasma of males. The results suggest that there could be a requirement for higher testosterone secretion by males during early stages of growth for attaining early sexual maturity.

Efficacy of Ovsynch protocol with antiprolactin treatment for timed artificial insemination during non-breeding seasons in yaks (Poephagus grunniens L.).

An attempt was undertaken to investigate the efficacy of Ovsynch protocol for timed artificial insemination (TAI) with or without Norprolac (antiprolactin) treatment during non-breeding season (winter months) in yaks (n = 25). During non-breeding season, plasma prolactin profile has been reported high due to cold and nutritional stress. The Norprolac dose of i.m. administration was standardized for prolactin suppression. Three different doses viz. 2.5, 5.0 and 7.5 mg were attempted and the dose of 7.5 mg Norprolac i.m. per animal was found to be suitable for suppression of prolactin levels up to 30 h. Ovsynch treatment with Norprolac induced more number of oestrous symptoms per animal (4.8 vs 2.1), higher LH peak concentration (24.01 vs 16.16 ng/ml), longer duration of LH surge (6.8 vs 5.2 h) and higher conception rate (70 vs 30%) in Ovsynch plus Norprolac treated animals compared with animals treated with Ovsynch alone. Therefore, this study clearly indicates the opportunity for practical application of the Ovsynch plus Norprolac protocol for TAI in yaks during non-breeding seasons.

Seasonal variations in plasma glucocorticoid levels in yaks (Poephagus grunniens L.) (Bos grunniens).

The changes of glucocorticoid concentrations were measured by highly sensitive enzymeimmunoassay procedures using second antibody coating technique and cortisol -HRP amplified system for one whole calendar year in yaks. The year was divided into seasons of cool, intermediate and hot seasons. The glucocorticoid values were lower (P < .01) in hotter months than in cool and intermediate seasons. Rectal temperature and respiration rates were indicative of heat stress in the hot temperature season and gave significant negative correlation with circulating corticoid concentration.

Temporal changes in endogenous estrogens and expression of behaviors associated with estrus during the periovulaory period in Murrah buffaloes (Bubalus bubalis).

The objective of this study were (1) to establish the duration of behavioral estrus signs and timing of ovulation in Murrah buffaloes (n = 10) and (2) to determine relationship between behavioral estrus signs with change in plasma estrogen concentrations. Estrus and its behavioral signs were detected at hourly intervals by visual observations, per recta examination of genitalia and bull parading four times in a day for 30 minutes each. Among the behavioral signs of estrus, swollen vulva (80%) was the best indicator of estrus followed by excitement (70%). Among the duration of behavioral estrus signs the first and longest duration of estrus signs was swollen vulva which was seen upto 19.8 +/- 0.8 h after onset of estrus. The mean total duration of estrus symptoms from appearance to disappearance of all the behavioral estrus symptoms was 23.5 +/- 1.7 h. All the behavioral estrus symptoms were observed during the period of estrogen surge. Endocrine profile during the periestrus period showed that the mean peak concentrations of total estrogen 23.9 +/- 3.9 pg/ml occurred at 8.8 +/- 1.7 h after onset of estrus. The average number of estrus symptoms observed per animal during onset of spontaneous estrus was 5.7. Ovulation occurred after 37.4 +/- 1.7 h after onset of estrus and 13.4 +/- 1.0 h after end of total estrogen surge respectively. In conclusion, our results suggest that all signs of behavioral estrus occurred during the preovulatory rise in estrogens. The first sign of estrus to be observed was a swollen vulva and this symptom persisted the longest.

Changes in endogenous estrogens and expression of behaviors associated with estrus during the periovulatory period in Heatsynch treated Murrah buffaloes (Bubalus bubalis).

The objective of this study was (1) to establish the duration of behavioral estrus signs and timing of ovulation in Murrah buffaloes (n = 10) and (2) to determine relationship between behavioral estrus signs with change in plasma estrogen concentrations in animals treated with Heatsynch protocol. Estrus and its behavioral signs were detected at hourly intervals by visual observations per rectal examination of genitalia and bull parading four times in a day for 30 min each. Among the behavioral signs of estrus, swollen vulva was the best indicator of estrus followed by excitement and chasing by bull (90%). Among the duration of behavioral estrus signs, the first and longest duration of estrus signs was swollen vulva, which was seen up to 21.6 +/- 1.1 h after onset of estrus. The mean total duration of estrus symptoms from appearance to disappearance of all the behavioral estrus symptoms was 40.2 +/- 1.8 h. Endocrine profile during the periestrus period showed that the mean peak concentrations of total estrogen 330.9 +/- 108.3 pg/ml occurred at 9.6 +/- 1.0 h after estradiol benzoate injection. The average number of estrus symptoms observed per animal during Heatsynch treatment was 7.1. Ovulation occurred after 50.0 +/- 2.0 h after estradiol benzoate treatment and 26.7 +/- 2.0 h after end of total estrogen surge, respectively. In conclusion, our results suggest that all signs of behavioral estrus occurred after the preovulatory rise in estrogens. The first sign of estrus was swollen vulva, and this symptom persisted the longest.