D.L. Thompson

Prolonged interovulatory interval and hormonal changes in mares following the use of Ovuplant™ to hasten ovulation

In its first year of commercial availability in the United States, reports from the field indicated that Ovuplant™ (a deslorelin-containing slow-release implant for hastening ovulation in mares) was associated with a delayed return to estrus in mares not becoming pregnant. Supposedly this effect was particularly prevalent in mares subsequently administered PGF2α to cause luteal regression after embryo collection. The present experiment was conducted 1) to determine if the field observations were repeatable under controlled experimental conditions, and 2) to gather endocrine data that might yield information on the underlying cause(s) of this observation. Twenty-five light horse mares were used. Ovaries of each mare were examined by transrectal ultrasonography daily during estrus until ovulation. Once a follicle >30 mm was detected, the mare received either Ovuplant (treated group; N = 13) at the recommended dosage or a sham injection (controls; N = 12); treatments were administered in a manner to ensure that they were unknown to personnel involved with data collection. On day 7 after ovulation, each mare received a luteolytic injection of PGF2ð. Mares were examined every other day until return to estrus or development of a 30 mm follicle, at which time daily examination was performed until ovulation. Jugular blood samples were collected daily. Two mares receiving Ovuplant did not return to estrus within 30 days and their data were not included in the statistical analyses; in contrast, no control mare exhibited such an extended interovulatory interval. For all other mares receiving Ovuplant, the interval between the first and second ovulations was longer (P = .0001) than that of control mares by an average of 6.2 days. In addition, plasma LH concentrations were lower (P <.05) in the treated mares on days 0 through 4, 9, 11, 18, and 19 after the first ovulation. Plasma FSH concentrations were also lower (P = .017) in treated mares from days 4 to 11 and on days 6 and 5 prior to the second ovulation (P = .005). Differences in progesterone and estradiol were observed but were less consistent than for LH and FSH. Mares receiving Ovuplant had fewer small (P =.026), medium (P = .003) and large (P = .045) follicles prior to the second ovulation. In conclusion, Ovuplant treatment at the recommended dosage decreased follicular activity after ovulation and increased the interovulatory interval in mares short-cycled with PGF2ð. These effects appear to be mediated by a hyposecretion of LH and(or) FSH.

Effects of dexamethasone, glucose infusion, adrenocorticotropin, and propylthiouracil on plasma leptin concentrations in horses

In experiment 1, nine light horse geldings (three 3 x 3 Latin squares) received dexamethasone (DEX; 125 microg/kg BW, i.m.), glucose (0.2 g/kg BW, i.v.), or nothing (control) once per day for 4 days. DEX increased (P < 0.001) glucose, insulin, and leptin concentrations and resulted in a delayed increase (P < 0.001) in IGF-I concentrations. In experiment 2, mares were similarly treated with DEX (n = 6) or vehicle (n = 6). DEX again increased (P < 0.01) glucose, insulin, and leptin concentrations; the delayed elevation in IGF-I concentrations occurred on day 10, 12, and 19, relative to the first day of treatment. In experiment 3, six light horse geldings received either 200 IU of adrenocorticotropin (ACTH) i.m. or vehicle twice daily for 4 days. ACTH increased (P < 0.001) cortisol concentrations. Further, ACTH resulted in increases (P < 0.01) glucose, insulin, and leptin concentrations. In experiment 4, plasma samples from four light horse stallions that were fed 6-n-propyl-2-thiouracil (PTU) at 6 mg/kg BW for 60 days to induce hypothyroidism were compared to samples from control stallions. On day 52, stallions receiving PTU had lower concentrations of thyroxine (P < 0.05) and triiodothyronine (P < 0.01) and higher (P < 0.01) concentrations of TSH. Leptin concentrations were higher (P < 0.01) in PTU-fed stallions from day 10 through 52. In conclusion, circulating concentrations of leptin in horses was increased by administering DEX. Treatment with ACTH increased cortisol and resulted in lesser increases in leptin, glucose, and insulin. In addition, PTU feeding results in lesser increases in leptin concentrations.

Administration of sulpiride to anovulatory mares in winter: effects on prolactin and gonadotropin concentrations, ovarian activity, ovulation and hair shedding

Sixteen seasonally anovulatory mares were randomly allotted to two groups and injected daily with either sulpiride (1 mg/kg body weight) or vehicle from 14 January to 14 February. Sulpiride administration increased daily plasma prolactin concentrations (P < 0.05), although the prolactin response during the 6 h following sulpiride injections decreased markedly from the 1st to the 6th day of treatment (treatment by day, P < 0.0001). Plasma concentrations of LH and FSH were not affected by treatment (P > 0.1). Injection of GnRH and TRH on 15 February showed that the response of plasma prolactin to secretagogue was increased in sulpiride-treated mares (P < 0.005), while there was no effect (P > 0.1) of sulpiride treatment on the response of LH or FSH. Both treatment groups had similar changes in numbers of follicles 10-19 and > or = 20 mm during the experiment (P > 0.1). Similarly, the mean change in maximal follicular size was not affected by treatment (P > 0.9). No mare ovulated during the study, and plasma progesterone concentrations were similar in both groups (P > 0.1), always at levels < 1 ng/ml. Hairshedding increased with time in all mares (P < 0.001) and was increased by sulpiride injections (P = 0.09). It was concluded that sulpiride administration to seasonally anovulatory mares under the conditions of our experiment increased daily plasma prolactin levels but did not stimulate gonadotropin secretion or ovarian activity.

Effects of breed and wintering diet on growth, puberty and plasma concentrations of growth hormone and insulin-like growth factor 1 in heifers

Twenty-five Brangus (BR) and 15 Angus (AN) heifers were used to study the effects of breed and wintering diet on average daily gain (ADG), onset of puberty and plasma concentrations of growth hormone (GH) and insulin-like growth factor 1 (IGF-1). Wintering diets (fed for 107 days beginning November 15) consisted of the following: 1) native grass hay (NGH), 2) ammoniated NGH, 3) NGH plus cottonseed meal, 4) Diet 3 plus corn and 5) Diet 4 plus monensin. After wintering, heifers were transferred to ryegrass pasture for 70 days. Mean ADG during the wintering phase were -.20, -.10, .17, .29 and .39 kg for heifers fed Diets 1 through 5, respectively (P less than .01). ADG was greater (P less than .05) for BR than for AN heifers. Plasma concentrations of GH were higher (P less than .05) in heifers fed Diets 1 and 2 than in heifers fed Diets 3, 4 or 5. Plasma concentrations of IGF-1 were lowest in heifers fed Diet 1 and highest in heifers fed Diets 4 and 5. During ryegrass grazing, GH concentrations were similar for all groups. However, concentrations of IGF-1 were higher (P less than .05) in heifers fed Diets 3, 4 and 5 than in heifers fed Diets 1 and 2. Age at puberty (onset of cyclic progesterone concentrations) was greatest in heifers fed Diet 1 and lowest in heifers fed Diet 5. Weight at puberty was not affected (P greater than .10) by wintering diet but was greater (P less than .01) in BR than in AN heifers. Therefore, negative ADG appears to be associated with elevated plasma GH concentrations in heifers, and plasma IGF-1 concentration appears to be a more accurate indication of nutritional status than plasma concentrations of GH.

Hyperleptinemia in mares and geldings: assessment of insulin sensitivity from glucose responses to insulin injection.

Four experiments were conducted 1) to assess the use of glucose responses to insulin injections as a means of estimating insulin sensitivity in horses and 2) to compare the insulin sensitivities of normal horses vs. those displaying hyperleptinemia (HL). In Exp. 1, HL mares and geldings (n = 4 each) and 4 mares and geldings with normal leptin concentrations (NL) were injected intravenously with 20 and 100 mU/kg of BW of bovine insulin on 2 separate occasions in December 2008. In Exp. 2, the experimental protocol was repeated in late April 2009. In Exp. 1, the 20 mU/kg of BW dose of insulin caused a greater (P < 0.05) decline in glucose concentrations in NL mares and geldings compared with HL horses. The response of HL mares to the 100 mU/kg of BW dose was less (P < 0.05) than for the other groups. In Exp. 2, responses of all groups to the 20 mU/kg of BW dose were small and similar among groups (P > 0.1), whereas the greater dose revealed differences (P < 0.05) in sensitivity among groups consistent with those observed with the smaller dose in Exp. 1. Experiment 3 was conducted in June and July of 2009 to further examine the dose-response relationship in mares of potentially different insulin sensitivities in an attempt to standardize the approach for studying a wide range of sensitivities. Recombinant human insulin was used at doses of 8, 20, 50, and 125 mU/kg of BW, as needed, to estimate (by linear regression) the dose of insulin causing a 50% decline in glucose concentrations (ED50). Five mares each of reduced leptin concentrations (LL) and small BCS (3 to 5), LL and larger BCS (6 to 7.5), and increased leptin concentrations and increased BCS were studied. The ED50 was similar (P > 0.1) for LL mares, regardless of BCS, and was less (P < 0.01) than for mares with increased leptin concentrations. It was concluded that a dose of 50 mU/kg of BW of recombinant human insulin could be used safely to start the dose-response curve; smaller or larger doses could then be applied as appropriate to get sufficient data for estimation of ED50. Experiment 4, conducted in October of 2009, assessed the repeatability of the estimates for ED50 obtained in Exp. 3. Six mares with LL vs. increased leptin concentrations received the 50 mU/kg dose of insulin; appropriate larger or smaller doses were used to obtain estimates of ED50. Estimates obtained were highly correlated (r = 0.91) with those obtained in Exp. 3, with an average within-mare CV of 8.9%; this is equal to or better than the repetabilities of the currently used methods of assessing insulin sensitivity in horses. It was concluded that hyperleptinemic horses, which are also hyperinsulinemic and have exaggerated insulin responses to glucose injection, are indeed less sensitive to insulin than normal horses with reduced leptin concentrations of the same body condition.

Effects of short-term stress, xylazine tranquilization and anesthetization with xylazine plus ketamine on plasma concentrations of cortisol, luteinizing hormone, follicle stimulating hormone and prolactin in ovariectomized pony mares

Long-term ovariectomized pony mares were subjected to one of four treatments: 1) control group - no treatment, 2) stressed group - 5 min of restraint via a twitch, 3) tranquilized group - administered xylazine (1.1 mg i.v. per kg of body weight), and 4) anesthetized group - administered xylazine followed 2 min later by ketamine (2.2 mg i.v. per kg of body weight). Blood samples were taken at -40, -30, -20, -10, -0.5, 10, 20, 30, 40, 50, 60 and 90 min and at 2, 3, 4, 6, 8 and 24 h relative to onset of treatment. Stress increased (P<0.05) cortisol concentrations 20 to 50 min after treatment and again at 6 and 8 h. Tranquilization had no effect on cortisol concentrations, whereas anesthetization increased (P<0.05) cortisol concentrations from 90 min through 8 h after treatment. Concentrations of luteinizing hormone (LH) and follicle stimulating hormone (FSH) did not vary (P>0.1) relative to pretreatment in any group of mares. Concentrations of prolactin were 2.7-fold higher (P<0.05) 24 h after treatment in all four groups, indicating some procedural or environmental influence on prolactin secretion. There was a transient increase (P<0.06) in prolactin concentrations in anesthetized mares 30 min after treatment. Although two of these three commonly used methods of restraint did affect cortisol concentrations, there was no effect on plasma concentrations of LH or FSH. Thus, we conclude that such methods of restraint can be used in short-term situations without disturbing estimates of LH and FSH secretion. However, when prolactin concentrations are to be measured, anesthesia with ketamine should not be used.

Growth and metabolic characteristics of Suffolk and Gulf Coast Native yearling ewes supplemented with chromium tripicolinate

Abstract An experiment was conducted to investigate differences in production and physiological criteria in yearling ewe lambs from two distinct breeds fed a concentrate–base diet (13.5% CP) supplemented with 0 (BAS) or 370 ppb (CRP) chromium tripicolinate. The breeds used were Suffolk (SFK, n=8, BW 58.9±1 kg), representing a breed selected for meat production, and Gulf Coast Native sheep (GCN, n=8, BW 44.9±1 kg), representing a breed selected for adaptability and wool production. Lambs were fed the BAS diet for 10 days. On day 0 of the experimental period, the lambs were stratified by BW within breed and randomly assigned to the experimental diets. On days 0, 11 and 22, after 18 h of feed deprivation, lambs were bled via jugular venipuncture and weighed. Moreover, on day 22, lambs were catheterized and an i.v. glucose tolerance test (IVGTT; 500 mg glucose kg−1 BW) was conducted. Statistical analyses of plasma metabolite and hormone measurements were conducted on their concentrations, as well as the relative changes (percent change) of these measurements for days 11 and 22 compared with day 0. Growth rate, feed intake and glucose and insulin kinetics in response to an IVGTT were not affected (P>0.10) by CRP. Relative to BAS, CRP decreased plasma NEFA concentration (P 0.10) plasma glucose, albumin, triacylglycerol or cortisol concentrations. CRP reduced plasma NEFA, but did not affect glucose kinetics. There were major differences in physiological measurements between the two distinct breeds of sheep that could explain the differences observed in performance.

Effects of age, season and active immunization against estrogen on serum prolactin concentrations in stallions

Prolactin concentrations in stallion serum were measured by radioimmunoassay based on antiserum generated against equine prolactin and radioiodinated canine prolactin. Prolactin concentrations in serum collected from 152 stallions at a slaughterhouse were higher (P less than .01) in summer than in winter (5.7 +/- .15 vs 2.0 +/- .17 ng/ml). Moreover, there was an effect of age (P less than .02) in the analysis of variance; there was no interaction between age and season. In general, prolactin concentrations increased with age up to 3 to 5 years. Samples of serum collected from five control and five estrogen-immunized stallions from 6 to 26 months of age also were assessed for prolactin concentration. Active immunization against estrogen had no effect (P greater than .10) on prolactin concentrations. In contrast, whether analyzed by age or by month of the year, prolactin concentrations varied with time (P less than .01) in these stallions. Averaged over both groups, concentrations were high during summer and low during winter and were also gradually increasing with time as indicated by a linear trend (P less than .01) in the data. It is concluded that serum prolactin concentrations in stallions 1) are seasonal, as in many other seasonally breeding species, 2) in general are highest in older stallions and lowest in young stallions and 3) are not affected by active immunization against estrogens.

Hyperleptinemia in horses: Responses to administration of a small dose of lipopolysaccharide endotoxin in mares and geldings

Mares and geldings in good body condition selected for hyperleptinemia vs. normal leptin concentrations were studied to determine whether the hyperleptinemic condition affected various characteristics of the hematologic and hormonal systems after a challenge with lipopolysaccharide endotoxin. Four mares and 4 geldings that were determined to be hyperleptinemic (mean plasma leptin concentrations of 10.0 to 15.5 ng/mL) and 4 mares and 4 geldings with mean plasma leptin concentrations between 2.4 and 5.5 ng/mL were administered Escherichia coli O55:B5 endotoxin (35 ng/kg of BW in 500 mL of saline over a 30-min infusion), or saline only, in pairs in a single-switchback design, with horses and treatments randomly assigned for the first infusion. Physiological variables and blood components were monitored for 24 h after the onset of infusions. Treatments were switched and the second infusions were administered 8 d later. Relative to vehicle infusion, endotoxin infusion increased (P < 0.01) the rectal temperature, heart rate, respiration rate, plasma total protein concentration, and blood packed cell volume; there was an interaction of leptin status, endotoxin treatment, and time for heart rate (P = 0.039), respiration rate (P = 0.018), and plasma total protein concentration (P = 0.054). Blood concentrations of leukocytes, lymphocytes, and neutrophils all decreased (P < 0.001) after endotoxin infusion; there was an interaction (P = 0.0057) between sex and leptin status for blood platelet concentration. Plasma leptin concentrations increased (P = 0.013) after endotoxin infusion in both hyperleptinemic horses and those with reduced leptin concentrations. There were interactions (P < 0.037) of sex with endotoxin treatment and time for plasma concentrations of cortisol and prolactin, whereas plasma GH concentrations were affected (increased; P < 0.001) only by time after infusion. Given that the effects of hyperleptinemia were generally minor, it was concluded that the hyperleptinemic condition, and its associated type-2 diabetic symptoms, has a minimal impact on the components of the hematologic and hormonal systems studied.

Effect of administration of adrenocorticotropic hormone on plasma concentrations of testosterone, luteinizing hormone, follicle stimulating hormone and cortisol in stallions

Summary In boars and rabbits, administration of adrenocorticotropic hormone (ACTH) results in a testis-dependent, short-term increase in concentrations of testosterone in peripheral plasma. This experiment was designed to assess the short-term effects of a single ACTH injection on plasma concentrations of testosterone, luteinizing hormone (LH), follicle stimulating hormone (FSH) and cortisol in stallions. Eight light horse and two pony stallions were paired by age and weight and then one of each pair was randomly assigned to the treatment (ACTH, .2 IU/kg of body weight) or control (vehicle) group. Injection of ACTH increased (P .10) in concentrations of cortisol during the blood sampling period. Control stallions exhibited a midday increase (P>.05) in concentrations of testosterone similar to that reported previously; ACTH treatment prevented or delayed this increase such that concentrations of testosterone in treated stallions were lower (P