Effects of nutrient restriction and subsequent realimentation in pregnant beef cows: Maternal endocrine profile, umbilical hemodynamics, and mammary gland development and hemodynamics.

Our hypothesis was that maternal nutrient restriction would negatively impact the endocrine and metabolic status of the pregnant cow, therefore influencing the mammary gland in preparation for lactation. We further hypothesized that earlier timing of realimentation could prevent negative impacts of nutrient restriction. The objectives were to investigate the influence of nutrient restriction and realimentation during early to late gestation on endocrine profile, umbilical hemodynamics, and mammary gland development and hemodynamics in pregnant beef cows. In Experiment 1, on d 30 of pregnancy cows (initial BW = 667.5 ± 13.4 kg, BCS = 6.2 ± 0.1) were randomly assigned to one of 3 treatments: 1) 100% NRC requirements from d 30 to 254 of gestation (CCC; n = 6); 2) 60% NRC from d 30 to 85, thereafter being re-alimented to 100% NRC to d 254 (RCC; n = 5); 3) or receive 60% NRC from d 30 to 140, thereafter being re-alimented to 100% NRC to d 254 (RRC; n = 6). Cows were returned to a common outdoor facility for calving thereafter and were fed ad libitum. In Experiment 2, on d 30 of pregnancy, cows (initial BW = 620.5 ± 11.3 kg, BCS = 5.1 ± 0.1) were randomly assigned to dietary treatments including: control (CON; 100% NRC; n = 18) and nutrient restriction (RES; 60% NRC; n = 30). On d 85 of pregnancy, cows were either slaughtered (CON, n = 6 and RES, n = 6), remained on control (CC; n = 12) and restricted (RR; n = 12) treatments, or were realimented to control (RC; n = 11). On d 140 of pregnancy, cows were either slaughtered (CC, n = 6; RR, n = 6; RC, n = 5), remained on control (CCC, n = 6; RCC, n = 5), or were realimented to control (RRC, n = 6). On d 254 of pregnancy, all remaining cows were slaughtered (CCC, n = 6; RCC, n = 5; RRC, n = 6). Mammary hemodynamics and endocrine profile were measured. Serum urea nitrogen, NEFA, as well as fetal parameters were measured in Experiment 1; whereas in Experiment 2, mammary gland development was recorded. In Experiment 1, RRC cows had lower dry matter intake (P = 0.001) and consequently lower BW change (P = 0.06). However, maternal nutrition did not alter mammary hemodynamics, hormonal patterns, and fetal characteristics (P > 0.11). In Experiment 2, CCC cows had increased (P = 0.02) mammary gland blood flow ipsilateral to the gravid horn as well as greater (P = 0.02) mammary gland fat on d 254. Nevertheless, plane of nutrition did not alter hormonal concentrations nor mammary gland characteristics (P > 0.15). These data indicate that nutrient restriction did not alter mammary hemodynamics nor endocrine profile throughout gestation.

Uteroplacental secretion of progesterone and estradiol-17ß in an ovine model of intrauterine growth restriction.

Using a mid to late gestation model of intrauterine growth restriction, uteroplacental secretion of progesterone and estradiol-17ß were examined. From day 50 to 130 of gestation, 31 ewe lambs were allocated to receive 100% (ADQ) or 60% (RES) of nutrient requirements. At day 130, umbilical and uterine artery blood flows were determined and blood samples were collected from maternal saphenous artery, gravid uterine vein, umbilical vein, and umbilical artery. Uteroplacental secretion of progesterone was increased in RES compared to ADQ fed dams. There was a net secretion and net metabolism of estradiol-17ß in RES, and ADQ fed dams, respectively. In relation to steroid synthesis, cotyledonary abundance of steroidogenic acute regulatory protein was greater in RES compared with ADQ fed dams, while abundance of aromatase was not different between dietary treatments. Caruncular aldo-keto reductase 1C abundance was less in RES compared to ADQ fed dams. The increase in progesterone secretion, therefore, is due in part to an increase in synthesis and a decrease in placental catabolism. Caruncular cytochrome P450 3A, which catalyzes the conversion of estrogens to catechol-estrogens, was in lesser abundance in RES compared to ADQ fed dams. Opposite responses in estradiol-17ß uteroplacental secretion compared with metabolism may be mediated through placental estrogen metabolism via cytochrome P450 enzymes.

Short communication: Effects of fluoxetine on lactation at weaning in sheep.

Selective serotonin reuptake inhibitors have been considered for use in the dairy industry to aid in dry-off procedures because of their ability to delay the onset of lactation. Fluoxetine (a selective serotonin reuptake inhibitor; FLX) is an agent that has been shown to delay the onset of lactogenesis stage II when taken during pregnancy and lactation in women. Two experiments were conducted to determine whether ewes would be an appropriate model to evaluate the effects of FLX on milk production at weaning. In the first experiment, 12 Suffolk cross ewes (body weight = 83.4 ± 12.2 kg; body condition score = 2.1 ± 0.4) in late lactation were assigned to treatments of 0 (control), 40, or 80 mg of FLX. They were given a single subcutaneous injection with the appropriate level of FLX mixed with propylene glycol at 0700 h on approximately d 78 of lactation (the day lambs were removed). In the second experiment, 18 Suffolk cross ewes (body condition score = 1.8 ± 0.3) from a previous lactation study were selected in late lactation. On approximately d 66 following parturition, weaning was initiated and ewes received a single oral bolus treatment (0, 80, or 160 mg of FLX). Treatment was administered using gelatin capsules containing the appropriate dose of FLX. For both experiments, milk production was estimated: in experiment 1 on d 0 (before treatment), 1, 2, and 3 (after treatment) at 0800 and 1100 h, and in experiment 2 on d 0, 1, and 2 following treatment at 0800 or 1100 h. Milk production was measured over a 3-h period. We observed no treatment differences or day effects on milk production in either experiment. In experiments 1 and 2, as the dose of FLX increased, milk production decreased linearly. Serum lactose concentrations were depressed in ewes treated with FLX in experiment 1 but similar across treatments in experiment 2. Overall, FLX depressed milk production in ewes; therefore, there is potential to use FLX as a dry-off agent in the dairy industry.

Effects of feeding ground redberry juniper (Juniperus pinchotii) to gestating ewes on pre- and postpartum performance, serum metabolites and hormones, milk fatty acid composition, and progeny preweaning performance.

The objective of this research was to evaluate effects of replacing sorghum × Sudangrass hay with ground juniper in gestating ewe supplements on pre- and postpartum growth performance, serum metabolites and hormonal concentrations, milk fatty acid composition, and progeny preweaning performance. In a completely randomized design, commercial Rambouillet ewes (age = 3 to 5 yr; initial BW = 65.2 ± 1.6 kg) on a base diet of long-stem sorghum × Sudangrass hay were assigned to 1 of 4 dietary supplements in which ground juniper replaced 0% (CNTL), 33% (18JUN), 66% (36JUN), or 100% (54JUN) of the ground sorghum × Sudangrass hay in a pelleted supplement with ground juniper from d 38 ± 4 of gestation to 2 d postpartum. Treatment DM diet intake overall (g/kg BW) in ewes receiving no juniper was similar ( ≥ 0.38) to that of those receiving increasing concentrations of juniper. Changes in ewe BW and BCS were similar ( ≥ 0.24) in ewes throughout gestation. All serum metabolites and hormones were within normal clinical ranges; however, serum IGF-1 decreased linearly ( = 0.003), alanine increased (linear; = 0.003), and serum Na decreased (linear; = 0.049) as the percentage of juniper increased in the diet. Ewe milk fatty acid composition was similar ( > 0.05) for the majority of fatty acids across treatment groups, with the exception of arachidonic acid (C20:4n6) being greater ( < 0.02) in 54JUN vs. CNTL ewe milk. Lamb birth weights were similar ( = 0.13), whereas lamb ADG tended to differ (quadratic; = 0.06) from d 0 to 14, with 18JUN being the least. At weaning, BW tended ( = 0.09) to linearly decrease in lambs born to ewes consuming greater concentrations of juniper but were not different ( = 0.26) between CNTL and 18JUN, 36JUN, and 54JUN. Results indicated that feeding increasing levels of ground juniper in supplements did not negatively alter ewe performance or serum metabolites and hormones measured pre- and postpartum. Lamb birth weight and preweaning performance appeared unaffected by maternal consumption of ground juniper containing supplements. Results also provide novel information regarding the effects of plant secondary compound consumption throughout pregnancy on ewe and progeny performance and health.

Protein kinase B is required for follicle-stimulating hormone mediated beta-catenin accumulation and estradiol production in granulosa cells of cattle.

Follicle-stimulating hormone regulation of ovarian estradiol (E2) production requires involvement of beta-catenin (CTNNB1), a transcriptional co-factor. In cultured granulosa cells (GC) of cattle, FSH treatment increased protein abundance of CTNNB1 as well as protein kinase B (AKT), a molecule known to regulate components of the CTNNB1 degradation complex. However, whether FSH induction of CTNNB1 is through direct modulation of AKT remains to be determined. To investigate specific contributions of AKT to CTNNB1 accumulation, GC were treated with insulin-like growth factor-I (IGF-I), a well-established AKT activator, in the presence or absence of FSH. Granulosa cells treated with FSH, IGF-I, and IGF-I plus FSH had increased CTNNB1 accumulation compared with controls (P ≤ 0.02; n=6). E2 medium concentrations were greater (P=0.09; n=4) in FSH treated cells compared to controls (166 and 100 ± 28 pg/mL, respectively). Treatment with IGF-I and IGF-I plus FSH increased (P<0.01) E2 to comparable concentrations. Subsequently, GC treated with lithium chloride (LiCl), a pharmacological activator of AKT, provided a response consistent with IGF-I treated cells, as LiCl, FSH, and FSH plus LiCl increased CTNNB1 accumulation compared with non-treated controls (P ≤ 0.03; n=3). In contrast, inhibition of AKT signaling with LY294002 suppressed the ability of FSH and IGF-I to regulate CTNNB1. Additionally, LY294002 treatment reduced FSH and IGF-I mediated E2 medium concentrations (P ≤ 0.004). These results demonstrate that activation of AKT is required for gonadotropin regulation of CTNNB1 accumulation and subsequent ovarian E2 production.

Arginine supplementation between 41 and 146 days of pregnancy reduces uterine blood flow in dairy heifers.

We hypothesized that arginine (Arg) supplementation during early pregnancy could foster placental vascularization. The objective of this study was to evaluate the effects of Arg supplementation on uterine artery hemodynamics between 40 and 140 days of gestation. At 41 days of gestation, 17 Holstein heifers (448 ± 73.9 kg) were surgically fitted with a catheter in the peritoneal cavity. After surgery, nine heifers received a daily intraperitoneal infusion of 40 mg of Arg/kg of body weight (BW; ARG), whereas the remaining eight heifers received a daily intraperitoneal infusion of saline solution at equivalent volumes as ARG heifers relative to BW (CTRL). Daily infusions took place every 12 hours until heifers reached 146 days of pregnancy. At 41, 62, 83, 104, 125, and 146 days of pregnancy, all heifers were body weighed, bled, and uterine blood flow volume (FV) and other hemodynamics were determined using Doppler ultrasonography. The measurements included heart rate, FV, pulsatility index, and resistance index. Plasma concentrations of amino acids (AAs), nitric oxide, glucose, insulin growth factor, progesterone (P4), growth hormone, and prolactin were analyzed. In a second experiment, we evaluated Arg metabolism when infused either intravenously or intraperitoneally. Overall, FV did not differ between the treatments, but it increased (P < 0.001) with pregnancy day. Uterine blood FV evolved differently between the treatments, with CTRL heifers having a greater (P < 0.05) FV (1104 ± 65.42 mL/min) at 146 days of pregnancy than in ARG heifers (806 ± 65.32 mL/min). Pulsatility index and resistance index decreased throughout pregnancy but did not differ between the treatments. Heart rate was decreased (P < 0.05) in ARG (74 ± 1.4 beats/min) compared with CTRL heifers (81 ± 1.5 beats/min). Plasma concentrations of Arg tended (P < 0.09) to be greater in ARG than those in CTRL heifers, but carnitine, valine, isoleucine, leucine, phenylalanine, and tryptophan were lower (P < 0.05) in ARG than those in CTRL heifers. The proportion of essential AAs (with respect to total AAs) in plasma was greater (P < 0.05) in CTRL (73 ± 2.4%) than that in ARG heifers (65 ± 2.3%). No differences between the treatments were found in plasma prolactin (P = 0.43), insulin growth factor (P = 0.97), glucose (P = 0.45), growth hormone (P = 0.39), or nitric oxide (P = 0.89) concentrations. However, plasma P4 concentration was greater (P < 0.05) in ARG than in CTRL heifers. Contrary to our hypothesis, Arg supplementation did not increase blood flow to the uterus but did change other parameters that could influence placental and fetal growth such as heart rate, maternal concentration of plasma AAs, or P4 synthesis. Furthermore, infusion sites (blood vs. peritoneum) of Arg render different metabolic responses.

Temporal concentrations of cortisol and LH in virgin ewes acutely exposed to rams during the transition into the breeding season.

The objectives of this study were to determine if exposing seasonally anovular ewes to rams would alter patterns of cortisol concentrations, and if these changes are associated with changes in characteristics of LH concentrations. Seasonally anestrous ewes were assigned to be exposed to rams (RE; n=11) or wethers (NE; n=12). Blood samples were collected at 15-min intervals beginning 120 min before introduction of males (time=0 min), and continued for 360 min after male exposure. Characteristics of cortisol and LH concentrations included: mean and baseline concentrations, pulse amplitude, duration, frequency, and time to first pulse. Mean and baseline cortisol concentrations, and cortisol pulse amplitude, frequency, and time to first pulse after male exposure did not differ between RE and NE ewes. Cortisol pulse duration was longer (P<0.05) in RE ewes than in NE ewes. Mean LH and LH pulse amplitude, duration, and time to first pulse after male exposure did not differ between RE and NE ewes. Baseline LH concentrations and LH pulse frequency were greater (P<0.05) in RE than in NE ewes. In RE ewes, but not NE ewes, LH pulse frequency tended to increase (P=0.06) as pulse frequency of cortisol decreased. In conclusion, exposing ewes to mature rams during the transition into the breeding season increased LH pulse frequency which hastened ovulatory activity. However, the results do not support the hypothesis that changes in cortisol concentrations plays a significant role in the 'ram effect'.

Effect of anabolic implants on adrenal cortisol synthesis in feedlot beef cattle implanted early or late in the finishing phase.

Implantation of anabolic steroids to increase growth rate in beef cattle impacts adrenal glucocorticoid production. The mechanism by which combination androgen and estrogen implants reduce cortisol biosynthesis in heifers is not clear. The objective of this study was to identify whether pituitary or adrenal gene expression and liver enzyme activity may contribute to altered serum cortisol concentrations in heifers receiving a combination implant. On d 0 of a 122-d finishing phase, 187 predominantly Angus heifers (361 kg) approximately 14 months old were randomly assigned to one of three implant groups: (1) non-implanted control, (2) implanted at the beginning of the finishing phase (d 0; early implant) with a combination implant (200mg TBA+20mg E2; Revalor 200®), and (3) implanted during the late stage of the finishing phase (d 56; late implant) with Revalor 200®. At d 56, body weight (BW) was greater (P<0.0001) for the early implanted heifers (456 ± 1.9 kg) compared to 437 and 435 (± 1.8) kg for control and late implanted heifers, respectively. Final BW (d 122) was similar between both implanted groups and heavier than non-implanted controls (P<0.0001). Serum cortisol was similar among groups at d 0 (P=0.86) however, by d 28 heifers receiving the combination implant had reduced (P<0.05) serum cortisol concentrations (31.2 ng/mL) compared to controls (49.4 ng/mL) and late (48.2 ng/mL) groups. On d 84 cortisol was similar (P=0.75) among implanted heifers and was less (P<0.01) than non-implanted heifers. Expression of pituitary and adrenal genes involved in glucocorticoid synthesis was evaluated at d 28/29 or 84/85; however, despite decreased serum cortisol in implanted heifers, no change in mRNA expression was demonstrated. Liver CYP3A enzyme activity at d 28/29 was decreased 59% in early implanted heifers compared to control heifers (P=0.01). Additionally, at d 84/85 AKR1C activity was greatest (P=0.01) in control heifers compared to both implanted groups. Data suggest that components of hypothalamic-pituitary-adrenal axis are influenced by exposure to exogenous hormones and this should be recognized when considering cortisol levels as a marker for stress response.

Effects of a dietary sweetener on growth performance and health of stressed beef calves and on diet digestibility and plasma and urinary metabolite concentrations of healthy calves.

Two experiments were conducted to explore the effects of a sodium saccharin-based dietary sweetener (Sucram) on growth performance, health, and physiological responses of feedlot steers. In Exp. 1, 173 newly-received male calves purchased from auction barns were fed 0, 100, 200, or 300 g of Sucram/t of DM over 56 d. Overall, ADG and G:F (P > 0.10) were not different among treatments, but steers receiving 200 g Sucram/t displayed numerically greater ADG (23%). In addition, DMI was 17% greater for steers receiving 200 g of Sucram/t compared to steers fed the control diet (cubic effect, P = 0.09). The morbidity rate for respiratory disease did not differ (P > 0.50) among treatments. In Exp. 2, 15 steers (initial BW = 261 ± 28 kg) were used to evaluate the effects of Sucram on apparent total tract digestibility, plasma metabolite concentrations, and urine monoamine metabolite concentrations. Treatments consisted of ad libitum access to a 60% concentrate diet (Control), ad libitum access to Control + 200 g of Sucram/t of DM (Adlib), and Control + 200 g of Sucram/t of DM with feed intake paired to the Control (Paired). By design, steer DMI during the metabolism period did not differ (P = 0.34) between Paired and Control, but DMI tended (P = 0.14) to be 8.2% greater for Adlib than for Control. Treatments did not alter (P > 0.17) apparent total tract nutrient digestibility. Postprandial plasma citrulline concentration was lower (P = 0.03) for Adlib than for Control and tended to be lower (P = 0.13) for Paired than for Control. Plasma homocysteine concentration was reduced (P < 0.03) by feeding Sucram. Urinary concentrations of ethylmalonic acid, vanillymandelic acid, and 5-hydroxyindolacetic acid were greater (P < 0.06) for Adlib than for Control; Paired steers had a greater (P = 0.02) urine vanillymandelic acid concentration than Control steers and tended (P < 0.12) to have a greater urinary concentration of ethylmalonic and 5-hydroxyindolacetic acid than Control steers. Serum insulin was greater for Adlib than for Control steers (P = 0.04) and tended to be greater for Paired than for Control steers (P = 0.14), but serum prolactin area did not differ (P > 0.22) among treatments. Supplementation with Sucram may increase feed intake by newly-received, stressed feedlot calves. Saccharin supplementation reduced plasma homocysteine and increased urinary excretion of vanillymandelic acid, suggesting an improved activity of the dopamine reward system.

Dietary selenium and nutritional plane alter specific aspects of maternal endocrine status during pregnancy and lactation.

Objectives were to examine effects of selenium (Se) supply and maternal nutritional plane during gestation on placental size at term and maternal endocrine profiles throughout gestation and early lactation. Ewe lambs (n = 84) were allocated to treatments that included Se supply of adequate Se (ASe; 11.5 µg/kg BW) or high Se (HSe; 77 µg/kg BW) initiated at breeding and nutritional plane of 60% (RES), 100% (CON), or 140% (EXC) of requirements beginning on day 40 of gestation. At parturition, lambs were removed from their dams, and ewes were transitioned to a common diet that met requirements of lactation. Blood samples were taken from a subset of ewes (n = 42) throughout gestation, during parturition, and throughout lactation to determine hormone concentrations. Cotyledon number was reduced (P = 0.03) in RES and EXC ewes compared with CON ewes. Placental delivery time tended (P = 0.08) to be shorter in HSe ewes than in ASe ewes, whereas placental delivery time was longer (P = 0.02) in RES ewes than in CON and EXC ewes. During gestation, maternal progesterone, estradiol-17ß, and GH were increased (P < 0.05) in RES ewes and decreased (P < 0.05) in EXC ewes compared with CON ewes. In contrast, maternal cortisol, IGF-I, prolactin, triiodothyronine, and thyroxine were decreased in RES ewes and increased in EXC ewes compared with CON ewes during gestation. Selenium supply did not alter maternal hormone profiles during gestation. During parturition and lactation, maternal hormone concentrations were influenced by both Se and maternal nutritional plane. During the parturient process, HSe ewes tended to have greater (P = 0.06) concentrations of estradiol-17ß than ASe ewes. Three hours after parturition a surge of GH was observed in ASe-RES ewes that was muted in HSe-RES ewes and not apparent in other ewes. Growth hormone area under the curve during the parturient process was increased (P < 0.05) in ASe-RES vs HSe-RES ewes. Ewes that were overfed during gestation had reduced (P < 0.05) estradiol-17ß but greater IGF-I, triiodothyronine, and thyroxine (P < 0.05) compared with RES ewes. Even though ewes were transitioned to a common diet after parturition, endocrine status continued to be affected into lactation. Moreover, it appears that gestational diet may partially affect lactational performance through altered endocrine status.

Performance by spring and fall-calving cows grazing with full, limited, or no access to toxic Neotyphodium coenophialum-infected tall fescue.

Replacing toxic, wild-type Neotyphodium coenophialum-infected tall fescue (E+) with nontoxic, N. coenophialum-infected tall fescue (NE+) has improved cow performance, but producer acceptance of NE+ has been slow. The objective was to compare performance by spring- and fall-calving cows grazing either E+ or NE+ at different percentages of the total pasture area. Gelbvieh×Angus crossbred cows (n=178) were stratified by BW and age within calving season and allocated randomly to 1 of 14 groups representing 5 treatments for a 3-yr study: i) Fall-calving on 100% E+ (F100); ii) Spring-calving on 100% E+ (S100); iii) Fall-calving on 75% E+ and 25% NE+ (F75); iv) Spring-calving on 75% E+ and 25% NE+ (S75); and v) Spring-calving on 100% NE+ (SNE100). Groups allocated to F75 and S75 grazed E+ until approximately 28 d before breeding and weaning, then were then moved to their respective NE+ pasture area for 4 to 6 wk; those allocated to F100, S100, and SNE100 grazed their pastures throughout the entire year. Samples of tall fescue were gathered from specific cells within each pasture at the time cows were moved into that particular cell (∼1 sample/mo). Blood samples were collected from the cows at the start and end of the breeding season. Stocking rate for each treatment was 1 cow/ha. Forage IVDMD, CP, and total ergot alkaloid concentrations were affected (P<0.05) by the treatment×sampling date interaction. Hay offered, cow BW, and BCS at breeding, end of breeding, and at weaning were greater (P<0.05) from fall-calving vs. spring-calving. Cow BW at weaning was greater (P<0.05) from F75 and S75 vs. F100 and S100. The calving season×NE+ % interaction affected (P<0.05) calving rates. Preweaning calf BW gain, actual and adjusted weaning BW, ADG, sale price, and calf value at weaning were greater (P<0.05) from fall-calving vs. spring-calving and from SNE100 vs. S75 except for sale price which was greater (P<0.05) from S75 vs. SNE100. Cow concentrations of serum prolactin at breeding and serum NEFA at the end of breeding were affected (P<0.05) by the calving season×NE+ % interaction. Serum Zn and Cu concentrations from cows were affected (P<0.05) by calving season. A fall-calving season may be more desirable for cows grazing E+, resulting in greater calving rates, cow performance, and calf BW at weaning, whereas limited access to NE+ may increase calving rates, serum prolactin, and NEFA concentrations during certain times in the production cycle, particularly in spring-calving cows.

Serum hormone profiles, pregnancy rates, and offspring performance of Rambouillet ewes treated with recombinant bovine somatotropin before breeding.

An experiment was conducted to examine effects of bovine ST (bST) on serum hormone concentrations, pregnancy rates, and offspring performance. Before initiation of a fall breeding period, 75 Rambouillet ewes (68.8 ± 1.5 kg) received an intravaginal insert containing 0.3 g of progesterone (P4) to synchronize onset of estrus. After 12 d, inserts were removed (d 0), and ewes (stratified by BW and age) received either 0 (control, n = 37) or 250 (n = 38) mg of recombinant bST (Posilac, Monsanto, St. Louis, MO, subcutaneously). Ewes were joined with fertile rams 24 h after insert removal. Blood samples were collected from 12 ewes in each treatment group daily from d 0 to 20 after insert removal. Serum IGF-I concentrations were 315 and 437 (± 58) ng/mL in control and bST-treated ewes 2 d after receiving bST (P = 0.02) and remained increased (P < 0.03) in bST-treated ewes throughout the 13-d period (P < 0.05). Serum prolactin (P > 0.10) and estradiol (P = 0.65) were similar between treatments. Serum triiodothyronine (T3) and thyroxine (T4) concentrations were similar (P > 0.20) between treatments from d 0 through 8. Controls had greater (P < 0.04) serum T3 and T4 concentrations than treated ewes did until d 18. Serum P4 was similar (P > 0.10) in control and bST-treated ewes from d 0 through 3 but was increased (P < 0.05) from d 4 to 8 in control ewes. Serum P4 was again similar (P > 0.10) between treatments from d 9 to 20. Serum insulin concentrations were 0.44 and 1.74 (± 0.19) ng/mL in control and bST-treated ewes, respectively, 1 d after receiving bST (P < 0.001) and remained increased (P < 0.03) in bST-treated ewes through d 9 (P < 0.03). Serum glucose was increased (P = 0.003) from d 0 to 10 in bST-treated ewes compared with controls. Thirty-three of 37 (89%) control ewes were pregnant, whereas 27 of 38 (71%) bST-treated ewes were pregnant (P = 0.05). As a percentage of ewes lambing, 61% and 39% of control ewes produced single and twin lambs, respectively, compared with 41% and 59% of bST-treated ewes (P = 0.12). Lamb 60-d adjusted weaning weights were 23.0 and 21.2 (± 0.65) kg for offspring produced by control and bST-treated dams, respectively (P = 0.04). In conclusion, serum IGF-I, insulin, and glucose were greater whereas serum T3, T4, and P4 were less in bST-treated ewes than in controls. Pregnancy rates and offspring adjusted weaning weights were decreased by bST treatment immediately before breeding.

Follicle-stimulating hormone regulation of estradiol production: possible involvement of WNT2 and ß-catenin in bovine granulosa cells.

Follicle-stimulating hormone regulation of estrogen biosynthesis in the adult rodent ovary requires ß-catenin (CTNNB1), but whether CTNNB1 is involved in FSH-induced estrogen production in cattle is unknown. To elucidate the effect of FSH in regulating specific wingless-type mouse mammary tumor virus integration site (WNT)/CTNNB1 pathway components in bovine folliculogenesis and steroidogenesis, granulosa cells and follicular fluid were collected from large antral follicles (8 to 22 mm) from ovaries containing stage-III corpora lutea (d 11 to 17 of an estrous cycle). Follicles were categorized as high estradiol (n = 3; ≥ 25 ng/mL) or low estradiol (n = 3; ≤ 14 ng/mL) based on intra-follicular estradiol concentrations. Protein fractions were collected from granulosa cells and CTNNB1 abundance was analyzed by Western blot. Follicles with increased estradiol concentrations had 6-fold greater (P < 0.001) abundances of CTNNB1 compared with those classified as low-estradiol follicles, indicating that the hormonal milieu responsible for increased estradiol content could result in CTNNB1 accumulation. To ascertain specific contributions of FSH to increases in CTNNB1 protein abundances, granulosa cells were isolated from small ovarian follicles (1 to 5 mm) and cultured in the presence or absence of 100 ng/mL FSH for 24 or 48 h. Real-time PCR quantification of aromatase (CYP19A1) and select WNT family members were evaluated in response to FSH treatment. Successful stimulation of granulosa cells with FSH was confirmed by induction of CYP19A1 mRNA and parallel temporal increases of medium estradiol concentrations. Additionally, protein kinase b (AKT), a known FSH target, increased 1.7-fold (P = 0.07). Of the WNT family members analyzed, only WNT2 mRNA was induced after 24 h of FSH treatment compared with controls (0.12-fold and 3.7-fold for control and FSH-treated, respectively; P < 0.05), and WNT2 expression tended (P = 0.11) to remain increased at 48 h in FSH-treated cells compared with controls (1.0- and 3.14-fold, respectively). Furthermore, FSH-treated granulosa cells had greater abundances of total CTNNB1 (P = 0.04) protein. These data demonstrate for the first time that FSH regulates CTNNB1 protein and WNT2 mRNA expressions in bovine granulosa cells, suggesting a potential role of canonical WNT signaling in ovarian steroidogenesis and follicular growth of cattle. Future studies are necessary to determine if FSH directly regulates CTNNB1 through modulation of AKT or indirectly by up regulating WNT2, which subsequently activates the canonical WNT pathway.

Effects of bacterial lipopolysaccharide injection on white blood cell counts, hematological variables, and serum glucose, insulin, and cortisol concentrations in ewes fed low- or high-protein diets.

Bacterial lipopolysaccharide endotoxins (LPS) elicit inflammatory responses reflective of acute bacterial infection. We determined if feeding ewes high-CP (15.5%) or low-CP (8.5%) diets for 10 d altered inflammatory responses to an intravenous bolus of 0 (control), 0.75 (L75), or 1.50 (L150) µg of LPS/kg of BW in a 2 × 3 factorial arrangement of treatments (n = 5/treatment). Rectal temperatures, heart and respiratory rates, blood leukocyte concentrations, and serum cortisol, insulin, and glucose concentrations were measured for 24 h after an LPS bolus (bolus = 0 h). In general, rectal temperatures were greater (P ≤ 0.05) in control ewes fed high CP, but LPS increased (P ≤ 0.05) rectal temperatures in a dose-dependent manner at most times between 2 and 24 h after the bolus. Peak rectal temperatures in L75 and L150 occurred 4 h after the bolus. A monophasic, dose-independent increase (P ≤ 0.023) in serum cortisol occurred from 0.5 to 24 h after the bolus, with peak cortisol at 4 h. Serum insulin was increased (P ≤ 0.016) by LPS in a dose-dependent manner from 4 to 24 h after the bolus. Insulin did not differ between control ewes fed high- and low-CP diets but was greater (P < 0.001) in L75 ewes fed low CP compared with high CP and in L150 ewes fed high CP compared with low CP. Increased insulin was not preceded by increased serum glucose. Total white blood cell concentrations were not affected (P ≥ 0.135) by LPS, but the neutrophil and monocyte fractions of white blood cells were increased (P ≤ 0.047) by LPS at 12 and 24 h and at 24 h after the bolus, respectively, and the lymphocyte fraction was increased (P = 0.037) at 2 h and decreased (P ≤ 0.006) at 12 and 24 h after the bolus. Red blood cell and hemoglobin concentrations and hematocrit (%) were increased (P ≤ 0.022) by LPS at 2 and 4 h after the bolus. Rectal temperatures and serum glucose were greater (P ≤ 0.033) in ewes fed a high-CP diet before LPS injection, but these effects were lost at and within 2.5 h of the bolus, respectively. Feeding high-CP diets for 10 d did not reduce inflammation in ewes during the first 24 h after LPS exposure but may benefit livestock by preventing acute insulin resistance when endotoxin exposure is mild.

Effects of body condition on measures of intramuscular and rump fat, endocrine factors, and calving rate of beef cows grazing common bermudagrass or endophyte-infected tall fescue.

Multiparous beef cows were managed to achieve marginal (BCS = 4.7 ± 0.07; n = 106) or good (BCS = 6.6 ± 0.06; n = 121) body condition (BC) to determine the influence of forage environment on BW and BC changes, intramuscular fat percentage (IMF), rump fat (RF), and serum hormones during 2 yr. Cows within each BC were randomly assigned to graze either common bermudagrass (CB; n = 3 pastures/yr) or toxic endophyte-infected tall fescue (EI; n = 3 pastures/yr) during a 60-d breeding season. Blood samples were collected at d 0, 30, and 60 of the breeding season, and serum concentrations of prolactin (PRL), IGF-I, and cortisol (CORT) were quantified; PRL and progesterone (P(4)) also were quantified 10 d before the breeding season (d -10). Body weight and BCS were recorded during the breeding season (d 0, 30, and 60). Cow IMF and RF were measured via ultrasonography at the start and end of the breeding season. Cows with increased (>1 ng/mL) P(4) at the beginning of the breeding season (cyclic) had greater (P < 0.02) concentrations of PRL on d 30 and 60 compared with anestrous cows. A forage environment × BC interaction tended (P = 0.07) to influence PRL. Cows grazing CB independent of BC had increased PRL compared with cows grazing EI. Prolactin was decreased in good-BC cows grazing EI compared with cows grazing CB, and cows in marginal BC grazing EI had the least concentrations of PRL. Concentrations of IGF-I were similar (P > 0.10) among good- and marginal-BC cows grazing CB, as well as good-BC cows grazing EI; however, marginal-BC cows grazing EI had reduced (P < 0.04) concentrations of IGF-I compared with all other groups. Cows in marginal BC grazing CB gained (P = 0.02) the most BW during the breeding season, whereas good-BC cows grazing EI gained the least amount of BW. Marginal-BC cows grazing CB tended (P = 0.06) to increase BC during the breeding season, whereas good-BC cows grazing either CB or EI lost BC. Rump fat tended (P = 0.07) to increase during the breeding season in marginal-BC cows compared with cows in good BC. Calving rates were similar (P > 0.10) among good- (82%) and marginal- (84%) BC cows grazing CB, and good-BC cows grazing EI (79%); however, marginal-BC cows grazing EI had a reduced (P = 0.04) calving rate (61%). Cattle grazing EI during the breeding season lost BC. That reduction in BC may be communicated to the pituitary via hormones that include IGF-I or PRL or both, resulting in decreased calving rates.

Technical note: comparison of salivary and serum cortisol concentrations after adrenocorticotropic hormone challenge in ewes.

An ACTH challenge was conducted to determine if salivary cortisol concentration reflects serum cortisol concentration in ewes. Twelve yearling ewes (64.0 +/- 1.2 kg) were administered ACTH (100 IU, intravenously) or saline. Serum and salivary samples were collected at 30-min intervals for 2 h before ACTH administration, at 15-min intervals for 2 h after treatment, and at 30-min intervals for an additional 3 h, and cortisol concentration was determined by RIA. Although ewes responded to ACTH and saline, cortisol concentration was greater (P < 0.001) in ACTH-treated ewes from 15 to 120 min and tended to be greater (P = 0.054) at 150 min after challenge in serum. In saliva, cortisol concentration was greater (P < 0.001) in ACTH-treated ewes from 30 to 120 min and tended to be greater (P = 0.092) at 15 min after challenge. No difference was observed between ACTH-treated ewes and controls for time to peak serum cortisol concentration (P = 0.126) and time to peak salivary cortisol concentration (P = 0.109), or between saliva and serum for time to peak cortisol concentration (P = 0.220) and return to baseline cortisol concentration (P = 0.341). The serum (P = 0.009) and salivary (P = 0.050) cortisol areas under the curve between 0 and 150 min were greater for ACTH-treated ewes than controls, and serum (P = 0.002) and salivary (P < 0.001) cortisol return to baseline concentration was longer for ACTH-treated ewes. The correlation coefficient between serum and salivary cortisol concentrations was 0.88 (P < 0.001). These data indicate that salivary cortisol concentration is closely related to serum cortisol concentration and that the former may represent a suitable noninvasive alternative to blood collection for measurement of cortisol in sheep.

High periconceptional protein intake modifies uterine and embryonic relationships increasing early pregnancy losses and embryo growth retardation in sheep.

The effects of supplemented protein level (PL) during the periconceptional period and their interaction with body condition were evaluated in sheep. Multiparous Rambouillet ewes (n = 12) received two PL of rumen undegradable protein (UIP) during a 30-day pre-mating and 15-day post-mating period: low [LPL, 24% crude protein (CP), 14 g UIP and 36 g/CP animal/day] and high [HPL, 44% CP, 30 g UIP and 50 g/CP animal/day]. While ovulation rate (OR) did not differ between treatments (1.6 +/- 0.5, mean +/- SEM), a lower fertility rate, a decreased embryo number and a reduced uterine pH (UpH) was observed in the HPL group (p < 0.05), irrespective of BC. Luteal tissue weight, volume and progesterone secretion did not differ among treatments. Sheep with lower UpH also had lower conceptus weight (Cwt; p < 0.05, r = 0.65) and conceptuses with lower mass tended to secrete less INF-tau and IGF-1, and the correspondent endometrial explants had a higher basal PGF(2alpha) release. Current study indicates that high protein diets during the periconceptional period in sheep modify uterine and embryonic relationships, increasing early pregnancy losses and inducing embryo growth retardation. Surviving embryos were affected by weight reductions, which could compromise later foetal growth and birth weight. Results evidence the key role of a balanced diet in reproductive success and indicate that the quality and nutrient composition of the maternal diet are essential for an adequate establishment of pregnancy, having paramount effects on the interplay of the embryo and the uterus.

Effects of dietary protein and bacterial lipopolysaccharide infusion on nitrogen metabolism and hormonal responses of growing beef steers.

Purified lipopolysaccharide (LPS) infusion in cattle induces clinical and metabolic responses similar to gram-negative bacterial infection. Effects of LPS and dietary protein on rectal temperature, serum hormones, haptoglobin, plasma urea N and AA, and N balance were evaluated in 24 steers (250 +/- 2.8 kg of BW). Treatments were a 2 x 3 factorial of LPS (0 vs. 1.5 microg/kg of BW; -LPS vs. +LPS) and diets containing (DM basis) 1) 14.5% CP, 11.6% ruminally degradable protein (RDP), and 2.9% ruminally undegradable protein (RUP; CP14.5CON); 2) 16.3% CP, 13.4% RDP, and 2.9% RUP (CP16RDP); and 3) 16.1% CP, 11.2% RDP, and 4.9% RUP (CP16RUP). Diet RDP and RUP were altered using casein, fish meal, and corn gluten meal. Steers were adapted to diets (1.1 Mcal/kg of NE(g); DM fed at 1.8% BW) for 14 d and were infused (intravenously 1 mL/min) with LPS (in 100 mL of saline) on d 15. Rectal temperature and serum cortisol, prolactin, haptoglobin, and insulin increased, glucose initially increased and then declined, and serum thyroxine and triiodothyronine decreased for +LPS vs. -LPS steers (LPS x hour; P < 0.01). Serum IGF-I was less (P < 0.01) for +LPS vs. -LPS steers. Plasma urea N increased in response to LPS (LPS x hour; P = 0.02) and was greater for +LPS steers fed CP16RDP and CP16RUP vs. CP14.5CON, but greater in -LPS steers fed CP16RUP vs. CP16RDP and CP14.5CON (LPS x diet; P = 0.04). Plasma Met, Thr, Leu, Ile, Phe, Trp, Gly, Ser, Asn, and Tyr decreased, and plasma Ala increased in response to LPS (LPS x hour; P < 0.01). Plasma Orn initially increased and then decreased in +LPS vs. -LPS steers (LPS x hour; P < 0.01). No LPS x diet interactions (P > or = 0.15) occurred for DM, OM, NDF and N intake, fecal excretion, or apparent digestibility. Dietary DM, OM, NDF, and N intake, and retained N were less (P < 0.01) for +LPS than -LPS steers. Total N intake, apparent N digestibility, and retained N were greater (P < or = 0.05) for steers fed CP16RDP and CP16RUP vs. CP14.5CON. An LPS x diet interaction (P = 0.05) occurred for N retention (% N intake) because N retention was less for +LPS than -LPS steers when fed CP14.5CON, but not different between +LPS and -LPS steers when fed CP16RDP and CP16RUP. These results demonstrate that LPS infusion alters serum hormones, plasma AA, and N balance in cattle and imply that growing steers exposed to LPS may require greater dietary protein concentrations to account for altered intake and metabolic AA demand.

Dynamics of GHRH in third-ventricle cerebrospinal fluid of cattle: relationship with serum concentrations of GH and responses to appetite-regulating peptides.

Objectives were to (1) characterize the relationship of third-ventricle (IIIV) cerebrospinal fluid (CSF) concentrations of growth hormone-releasing hormone (GHRH) with concentrations of GH in the peripheral circulation; and (2) assess the influence of acute administration of appetite-regulating peptides leptin (anti-orexigenic) and neuropeptide Y (NPY; orexigenic) on the release of GHRH. Six mature beef cows fitted with IIIV and jugular vein cannulae were treated intracerebroventricularly with saline, and leptin (600 microg) and NPY (500 microg) in saline, in a replicated 3x3 Latin square design. Third-ventricle CSF and blood were collected 10 min before and continued 220 min after treatments. Mean concentrations of GHRH and frequency of pulses after treatments were 2.2+/-0.13 ng/mL and 1.2+/-0.15 pulses/220 min, respectively. These measures were not influenced by treatments. Concentrations of GHRH in CSF were weakly correlated (r=0.15; P<0.03) with serum concentrations of GH; however, 58% of the GH pulses were preceded by a pulse of GHRH and 90% of the GHRH pulses occurred within 20 min preceding a pulse of GH. Leptin tended (P<0.10) to suppress GH area under the curve (AUC) compared to saline. Concomitantly, NPY tended (P<0.10) to increase GH AUC, which appeared to be a consequence of increased (P<0.05) pulse amplitude. Infusion of NPY also increased (P<0.05) AUC of GHRH relative to saline. No differences were detected among treatments in serum concentrations of insulin-like growth factor-I or its AUC. Sampling CSF from the IIIV appears to be a viable procedure for assessing hypothalamic release of GHRH coincident with anterior pituitary gland secretion of GH in cattle. These data also demonstrate the differential responsiveness of the GH axis to appetite-regulating peptides.

Short-term intake of beta-carotene-supplemented diets enhances ovarian function and progesterone synthesis in goats.

The effect of beta-carotene supplementation upon luteal activity, measured as number (CLT) and volume (VLT) of corpus luteum, and P4 synthesis in goats, was evaluated. Goats (n = 22, 34 months) were randomly assigned to one of two experimental groups: (i) beta-carotene [Beta, n = 10; body weight (BW = 44.8 +/- 1.45 kg), body condition score (BCS = 3.25 +/- 0.07)], and (ii) Control (Control, n = 12; BW = 45.30 +/- 1.32 kg, BCS = 3.33 +/- 0.06). Upon oestrus synchronization, the Beta group received 50 mg of beta-carotene per day during 35 days pre- and 17 days post-ovulation. The day 4, 8, 12 and 16 post-ovulation, blood samples were collected for quantification of serum P4 concentrations by radioimmmunoassay, and transrectal ultrasonographic scanning was performed at day 18 for evaluating CLT and VLT. Overall, CLT and VLT mean were 3.10 and 2211.1 mm(3) respectively. The Beta-goats depicted both the largest values for CLT (p = 0.07) and serum P4 levels (p = 0.05), with no differences (p = 0.53) for VLT between treatments. Results suggest a higher efficiency within the cellular-enzymatic groups defining the steroidogenic pathways in the beta-carotene-supplemented goats, generating a larger P4 synthesis. The last is essential for ovulation of healthy oocytes, maintenance of uterine quiescence, nourishment and survival of the embryo around implantation; all of them of paramount significance during the maternal recognition of pregnancy process.