Short-term biological variation of serum thyroid hormones concentrations in clinically healthy cats.

Thyroid disease is common in cats, but little is known about the biologic variability of serum thyroid hormone concentrations and its impact on diagnostic utility in either healthy cats or cats with thyroid disease. The purpose of this study was to determine the biological variation, index of individuality, and reference change values for thyroid hormones and thyroid-stimulating hormone (TSH) in clinically healthy cats. Serum samples for analysis of total thyroxine (T4), triiodothyronine (T3), free T4 by dialysis, and TSH were obtained weekly for 6 wk from 10 healthy cats, then frozen until single-batch analyzed. Data were evaluated for outliers, and we determined the CV within individual cats (CVI) and between individual cats (CVG) for each hormone and the variation between duplicates or analytical variation (CVA). The index of individuality and reference change values for each hormone were then calculated. Serum concentrations of total T4, free T4, T3, and TSH all showed greater variation between cats (CVG) than within cats (CVI). Total and free T4 had an intermediate index of individuality (1.1 and 1.2, respectively), suggesting that these hormones would be best evaluated by a combination of their population-based reference intervals and reference change values. Serum TSH concentrations had high index of individuality (1.8), suggesting this hormone would be best evaluated with reference change values rather than the population-based reference interval. Total T3 also had a high calculated index of individuality (1.8); however, T3 had high ratio of analytical variation (CVA) to within cat variation (CVI), so RCV could not be accurately calculated. This study demonstrates that clinically normal cats show considerable interindividual biological variation in serum thyroid hormone and TSH concentrations, whereas the intraindividual variability in hormone concentrations is much narrower. This suggests that for all serum thyroid hormones, but especially serum TSH and T3 concentrations, comparing individual cat's hormone results to a population-based reference interval may be misleading, especially in those with early or subclinical thyroid disease. Clinicians might improve the diagnosis of feline thyroid disease by establishing baseline concentrations of T4, free T4, T3, and TSH for individual cats (ideally when healthy) and applying reference change values to subsequent measurements.

Effect of dry period dietary energy level in dairy cattle on volume, concentrations of immunoglobulin G, insulin, and fatty acid composition of colostrum.

The objective was to investigate the effect of different dry cow feeding strategies on the volume, concentration of IgG and insulin, as well as fatty acid composition of colostrum. Our hypothesis was that different dry period diets formulated to resemble current feeding practices on commercial dairy farms and differing in plane of energy would have an effect on IgG and insulin concentration, as well as composition of fatty acid of colostrum. Animals (n=84) entering parity 2 or greater were dried off 57 d before expected parturition and fed either a diet formulated to meet, but not greatly exceed energy requirements throughout the dry period (CON), or a higher energy density diet, supplying approximately 150% of energy requirements (HI). A third group received the same diet as group CON from dry-off until 29 d before expected parturition. After this time point, from 28 d before expected parturition until calving, they received a diet formulated to supply approximately 125% of energy requirements (I-med). Concentration of IgG and insulin in colostrum were measured by radial immunodiffusion and RIA, respectively. Composition of fatty acids was determined by gas-liquid chromatography. The IgG concentration was highest in colostrum of cows in group CON [96.1 (95% CI: 83.3-108.9) g/L] and lowest in group HI [72.4 (60.3-84.5) g/L], whereas insulin concentration was highest in group HI [1,105 (960-1,250) µU/mL] and lowest in group CON [853 (700-1,007) µU/mL]. Colostrum yield did not differ between treatments and was 5.9 (4.5-7.4), 7.0 (5.6-8.4), and 7.3 (5.9-8.7) kg in groups CON, I-med, and HI, respectively. A multivariable linear regression model showed the effect of dietary treatment group on IgG concentration was independent of the effect of dry matter. Cows in groups CON, I-med, and HI had an average colostral fat percentage of 5.0 (4.1-5.9), 5.6 (4.8-6.4), and 6.0 (5.2-6.8) and an average fat yield of 289 (196-380), 406 (318-495), and 384 (295-473) g, respectively. Colostrum of cows fed to exceed predicted energy requirements during the last 4 wk of the dry period (groups I-med and HI) exhibited a higher concentration and yield of de novo fatty acid and lower concentrations but similar yield of preformed fatty acid compared with cows in group CON. Feeding a controlled energy diet during the dry period increased colostral IgG concentration in this study, whereas overall colostrum yield was not affected. Exceeding energy requirements during the period of colostrogenesis led to higher concentrations of insulin and de novo fatty acid in colostrum.

Free thyroxine concentrations by equilibrium dialysis and chemiluminescent immunoassays in 13 hypothyroid dogs positive for thyroglobulin antibody.

OBJECTIVE: To determine if concentrations of free thyroxine (FT4) measured by semi-automated chemiluminescent immunoassay (CLIA) correspond to FT4 determined by equilibrium dialysis (ED) in hypothyroid dogs positive for thyroglobulin antibody (TGA). ANIMALS: Thirteen TGA-positive dogs classified as hypothyroid based on subnormal FT4 concentrations by ED. METHODS: Qualitative assessment of canine TGA was performed using an enzyme-linked immunosorbent assay. Serum total thyroxine and total triiodothyronine concentrations were measured by radioimmunoassay. Serum FT4 concentration was determined by ED, and also by semi-automated CLIA for human FT4 (FT4h) and veterinary FT4 (FT4v). Canine thyroid stimulating hormone concentration was measured by semi-automated CLIA. RESULTS: Each dog's comprehensive thyroid profile supported a diagnosis of hypothyroidism. For detection of hypothyroidism, sensitivities of CLIA for FT4h and FT4v were 62% (95% CI, 32-85%) and 75% (95% CI, 36-96%), respectively, compared to FT4 by ED. Five of 13 (38%) dogs had FT4h and 2 of 8 (25%) dogs had FT4v concentrations by CLIA that were increased or within the reference range. Percentage of false-negative test results for FT4 by CLIA compared to ED was significantly (P < .0001 for FT4h and P < .001for FT4v) higher than the hypothesized false-negative rate of 0%. CONCLUSIONS AND CLINICAL IMPORTANCE: Caution should be exercised in screening dogs for hypothyroidism using FT4 measured by CLIA alone. Some (25-38%) TGA-positive hypothyroid dogs had FT4 concentrations determined by CLIA that did not support a diagnosis of hypothyroidism.

Seasonal variation in serum concentrations of selected metabolic hormones in horses.

BACKGROUND: Determination of adrenocorticotropic hormone (ACTH) concentration is a commonly used test in the evaluation of endocrine causes of equine laminitis, but the concentration in healthy horses can be high at certain times of year, which alters the specificity of the ACTH test. OBJECTIVE: To determine if circulating concentrations of ACTH, cortisol, glucose, insulin, and thyroxine vary month to month in healthy horses and in horses with equine metabolic syndrome (EMS). ANIMALS: Nine healthy adult horses were studied on their farm/stable over the course of 1 year. After the diagnosis of EMS, 10 laminitic horses residing at the same farm/stable were also studied. METHODS: Prospective study of healthy and laminitic horses. Plasma/serum samples were analyzed for concentrations of hormones and glucose. RESULTS: ACTH was the only analyte to show a discrete seasonal pattern, with concentrations in healthy and EMS horses frequently outside of the reference range (9-35 pg/mL) in August through October. Insulin was elevated (>40 microIU/mL) in EMS horses during most months and median serum glucose was generally higher in EMS horses (100 mg/dL, range, 76-163 mg/ dL) than in controls (94 mg/dL, range, 56-110 mg/dL), but no seasonal patterns for insulin or glucose were found. CONCLUSIONS AND CLINICAL IMPORTANCE: An increased ACTH concentration in horses in late summer or autumn should be interpreted with caution. In contrast, insulin concentration is maintained within the reference range throughout the year in healthy horses, thus an increased insulin concentration at any time of year should raise suspicions of EMS, ECD, or both.

Comparison between vestibular and subcutaneous insertion of deslorelin implants for oestrus induction in bitches.

Investigations using sustained-release deslorelin implants at various insertion sites have shown that this method consistently induces oestrus in anoestus bitches. However, fertility comparisons between implant insertion sites have not been performed. Anestrous beagle bitches received one 2.1 mg deslorelin implant beneath the vestibular mucosa (VM group; n = 6) or in the subcutaneous tissue between the shoulder blades (SubQ group; n = 8). Vestibular implants were removed when serum progesterone concentrations first exceeded 1.5 ng/ml. Vaginal cytologies and blood samples were collected daily and bitches were inseminated during oestrus. Serum progesterone and deslorelin concentrations were measured and pregnancy status was determined using ultrasonography. There were no differences between groups in the intervals between implant administration and the onset of proestrus, the time of the luteinizing hormone surge and the onset of cytologic diestrus. There were also no differences in the number of corpora lutea or foetuses. However, conception rate was significantly lower in the SubQ group. The pregnancy rate did not differ significantly between the VM and SubQ groups [4 out of 6 (66.7%) and 3 out of 8 (37.5%), respectively]. One bitch (16.7%) in the VM group and three bitches (37.5%) in the SubQ group suffered distinct, premature declines in serum progesterone concentrations starting 1-4 weeks after cytologic diestrus. Serum progesterone concentrations did not recover (premature luteal failure), resulting in abortion. Bitches with premature luteal failure in the SubQ group still had serum deslorelin concentrations >100 pg/ml 20 days after implant insertion, suggesting a possible association between prolonged deslorelin release and luteal failure.

Plasma adrenocorticotropin, cortisol, and adrenocorticotropin/cortisol ratios in septic and normal-term foals.

BACKGROUND: Little information exists on the hypothalamic-pituitary-adrenal axis in septic foals. HYPOTHESIS: The plasma concentrations of adrenocorticotropin (ACTH) and cortisol are expected to be higher in septic foals as compared to normal foals. The concentrations of hormones in septic foals also are expected to differ further depending upon survival. ANIMALS: Twenty-eight control foals and 46 septic foals <14 days of age were included in this study. METHODS: Blood was collected in EDTA once from 28 normal foals born in the University of Georgia or Cornell University equine research herds and from 46 septic foals within 12 hours after admission to 1 of the 3 tertiary care referral centers involved in the study. Septic foal selection was based on a sepsis score of >11 or a positive blood culture. The control foals were age matched to the septic foals in the study. ACTH and cortisol concentrations were measured by a chemiluminescent immunoassay system. RESULTS: Cortisol concentrations in control foals did not vary with age. Septic foals had significantly higher mean ACTH, cortisol, and ACTH/cortisol ratios than did normal foals. Within the septic foal group, 28 foals survived to discharge, and 18 were euthanized or died. The mean age was not significantly different between the septic surviving and nonsurviving foals. The mean ACTH/cortisol ratio was significantly higher in the septic nonsurviving foals as compared to the septic surviving foals. CONCLUSIONS AND CLINICAL IMPORTANCE: Septic foals had higher hormone concentrations as compared to normal foals, which is an expected endocrine response to critical illness. The increased ACTH/cortisol ratio in nonsurviving septic foals in comparison to surviving septic foals could indicate hypothalamic-pituitary-adrenal axis dysfunction at the level of the adrenal gland in critically ill septic foals.

Failure of hCG to support luteal function in bitches after estrus induction using deslorelin implants.

Induction of estrus with deslorelin implants was followed by abortions in bitches that conceived during the induced estrus. Lowering the deslorelin dose and choosing a better implantation site prevented the abortions. This study investigated the hypothesis that induction of estrus with deslorelin is followed by reduced serum progesterone concentrations (SPC) during the ensuing diestrus. Assuming that reduced luteal function resulted from reduced LH secretion due to hypophyseal down-regulation of GnRH receptors, the effect of human chorionic gonadotropin (hCG) treatment on the SPC of diestrous bitches was also investigated. In Experiment 1, 10 spontaneously cycling bitches served as controls, whereas estrus was induced with deslorelin implants in 24 others. In Experiment 2, six diestrous bitches were treated with a single dose of hCG between Days 39 and 45 of diestrus. The SPC was lower in deslorelin-induced bitches from Days 35 to 56 of diestrus and hCG increased SPC during the first 24 h after treatment, followed by a dramatic decline thereafter. Although SPC recovered in pregnant bitches, it remained much lower (< or = 1 ng/mL) than in untreated, non-pregnant bitches. The suppression of progesterone secretion after hCG treatment suggested that decreased luteal activity in deslorelin-induced bitches may not be a simple consequence of down-regulation of hypophyseal GnRH receptors.

Subnormal concentrations of serum cobalamin (vitamin B12) in cats with gastrointestinal disease.

The present study sought to determine the spectrum of diseases associated with subnormal concentrations of serum cobalamin in cats undergoing investigation of suspected gastrointestinal problems. The solid-phase boil radioassay (RA) for cobalamin employed in the present study was immunologically specific, precise, and accurate, with a sensitivity of 15 pg/mL. The RA yielded results that strongly correlated with those obtained by bioassay (Spearmann rho = .805; P < .0001), although the absolute values were lower for the RA. Forty-nine of 80 serum samples submitted during the period of January 1996-January 1998 had cobalamin concentrations below the reference range for healthy cats (range 900-2,800 pg/mL; mean +/- SD, 1,775 +/- 535 pg/mL; n = 33). Cats with subnormal cobalamin concentrations (mean +/- SD; 384 +/- 272 pg/mL, range 3-883 pg/mL) were middle-aged or older and were presented for weight loss. diarrhea, vomiting, anorexia, and thickened intestines. Definitive diagnoses in 22 cats included inflammatory bowel disease (IBD), intestinal lymphoma, cholangiohepatitis or cholangits, and pancreatic inflammation. Serum concentrations of cobalamin were particularly low in cats with intestinal lymphoma, three-fifths of whom also had subnormal serum concentrations of folate (< 9 ng/mL). The simultaneous presence of disease in the intestines, pancreas, or hepatobiliary system in many cats made it difficult to determine the cause of subnormal cobalamin concentrations. The circulating half-life of parenteral cyanocobalamin was shorter in 2 cats with IBD (5 days) than in 4 healthy cats (12.75 days). The presence of subnormal serum concentrations of cobalamin in 49 of 80 cats evaluated suggests that the measurement of serum cobalamin may be a useful indirect indicator of enteric or pancreatic disease in cats. The rapid depletion of circulating cobalamin in cats suggests that cats may be highly susceptible to cobalamin deficiency. However, the relationship of subnormal serum cobalamin concentrations to cobalamin deficiency and the effect of cobalamin deficiency on cats remain to be determined.

Effect of fentanyl on the minimum alveolar concentration of isoflurane in swine.

BACKGROUND: Fentanyl is used in anesthetic protocols for swine, but there are no reports on its potency in this species. This study measured the extent to which fentanyl reduces the minimum alveolar concentration of isoflurane (MACISO) in swine. METHODS: Sixteen swine were randomly assigned to four groups. For each group, baseline MACISO was determined, and three groups received two of three fentanyl infusions as follows: 50 micrograms.kg-1.h-1 intravenously followed by 100 micrograms.kg-1.h-1, 50 micrograms.kg-1.h-1 followed by 200 micrograms.kg-1.h-1, or 100 micrograms.kg-1.h-1 followed by 200 micrograms.kg-1.h-1 (n = 8 for each dosage). A loading dose of fentanyl preceded each infusion. Each infusion was maintained for 60 min before initiating minimum alveolar concentration determination. The infusions were maintained throughout the period of minimum alveolar concentration determination. Plasma fentanyl samples were obtained after 30 min of each infusion, and plasma fentanyl and hemodynamic parameters were obtained immediately before stimulating swine for the final isoflurane concentration used in determining minimum alveolar concentration. A fourth group, control animals, received saline infusions. After each infusion, the MACISO was redetermined. Minimum alveolar concentration was determined using incremental changes in isoflurane concentrations until gross purposeful movement resulted when using a hemostat stimulus applied for 1 min to a rear dewclaw. RESULTS: MACISO for controls was 2.19 +/- 0.17% (mean +/- SEM) and changed minimally over time (-0.13 +/- 4.77%). MACISO decreased significantly (P < or = 0.01) 24.5 +/- 3.2%, 29.9 +/- 4.8%, and 45.9 +/- 5.5% with fentanyl dosages of 50, 100, and 200 micrograms.kg-1.h-1, respectively. Corresponding plasma fentanyl concentrations were 14 +/- 1 ng/ml, 26 +/- 3 ng/ml, and 59 +/- 5 ng/ml, respectively. A ceiling effect on reduction of MACISO was not observed. Changes over time or between groups were not observed for arterial blood gas tensions, blood pressure, heart and respiratory rate, or body temperature. CONCLUSIONS: These fentanyl dosages are larger than those commonly used in humans and other species. Anesthetic protocols using fentanyl for swine should be designed with the knowledge that a fentanyl infusion of 200 micrograms.kg-1.h-1 contributes approximately a 50% MACISO equivalent.

Estrogen production by bovine binucleate and mononucleate trophoblastic cells in vitro.

The bovine placenta has long been known as a source of steroid hormones. We performed three experiments to compare production of estrogens by bovine mononucleate and binucleate trophoblastic cells and examined effects of cortisol, progesterone, pregnenolone, testosterone, and androstenedione. In the first experiment, binucleate trophoblastic cells were purified by unit gravity sedimentation from six enzymatically dispersed placentas between 150 and 180 days of gestation. Cells (8 x 10(5)/ml) were incubated first at 37 degrees C for 6 h with Medium 199 alone (M199/6h) or with 10(-7) M cortisol (cortisol/6h). Medium then was replaced with 10(-7) M progesterone, 10(-7) M pregnenolone, 10(-7) M testosterone, or M199, and a second incubation was conducted for 4 h. Estradiol production did not differ between cells incubated for the first 6 h in M199 vs. cortisol and was not affected by progesterone or pregnenolone. Testosterone increased (p < 0.05) estradiol production. Estrone production did not differ between cells incubated for the first 6 h in M199 vs. cortisol; estrone production was not affected by either progesterone, pregnenolone, or testosterone. Mononucleate as well as binucleate cells were purified from placentas between 165 and 180 days of gestation and used in two other experiments. In the first of these, enriched populations of binucleate and mononucleate cells were incubated first for 6 h with Medium 199 (M199) or 10(-7) M cortisol. Medium then was replaced with 10(-7) M testosterone, 10(-7) M androstenedione, or M199 and incubation continued for 4 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of hyperlipemia on radioimmunoassays for progesterone, testosterone, thyroxine, and cortisol in serum and plasma samples from dogs.

Hyperlipemic serum and plasma samples often are received by clinical laboratories for endocrinologic analysis by radioimmunoassay. We designed a study to determine what effect, if any, hyperlipemia has on estimation of lipid-soluble hormone concentrations determined by solid-phase radioimmunoassays. Progesterone, testosterone, thyroxine, and cortisol concentrations were determined in canine plasma and serum with various degrees of lipemia. Samples of serum, heparinized plasma, and EDTA-treated plasma were obtained from blood collected from 4 female and 4 male Beagles by use of evacuated tubes. To induce hyperlipemia in vitro, IV fat emulsion was diluted in deionized water to produce 0 (water only), 33, 67, or 100% mixtures. Twenty microliters of each mixture then was added to the subsamples of serum and plasma from each dog. Hormone concentrations were determined, using validated radioimmunoassays. Triglyceride concentrations were determined by enzymatic assay. Addition of IV fat emulsion in vitro was an accurate and reproducible means of altering triglyceride concentrations in the samples. Triglyceride concentrations as high as 700 mg/dl had no effect on radioimmunoassays for progesterone, testosterone, and thyroxine in serum, heparinized plasma, or EDTA-treated plasma. Addition of 100% (but not 33 or 67%) fat emulsion reduced the mean cortisol concentration in heparinized plasma by 12% (P less than 0.05). This severe hyperlipemia did not affect quantification of cortisol in serum or EDTA-treated plasma.

Effects of hemolysis and storage on quantification of hormones in blood samples from dogs, cattle, and horses.

Veterinary diagnostic endocrinology laboratories frequently receive hemolyzed plasma, serum, or blood samples for hormone analyses. However, except for the previously reported harm done by hemolysis to canine insulin, effects of hemolysis on quantification of other clinically important hormones are unknown. Therefore, these studies were designed to evaluate effects of hemolysis on radioimmunoassay of thyroxine, 3,5,3'-triiodothyronine, progesterone, testosterone, estradiol, cortisol, and insulin in equine, bovine, and canine plasma. In the first experiment, hormones were measured in plasma obtained from hemolyzed blood that had been stored for 18 hours. Blood samples were drawn from pregnant cows, male and diestrous female dogs, and male and pregnant female horses. Each sample was divided into 2 equal portions. One portion was ejected 4 times with a syringe through a 20-gauge (dogs, horses) or 22-gauge (cows) hypodermic needle to induce variable degrees of hemolysis. Two subsamples of the blood were taken before the first and after the first, second, and fourth ejections. One subsample of each pair was stored at 2 to 4 C and the other was stored at 20 to 22 C for 18 to 22 hours before plasma was recovered and stored at -20 C. The second portion of blood from each animal was centrifuged after collection; plasma was recovered and treated similarly as was blood. Concentrations of thyroxine in equine plasma, of 3,5,3'-triiodothyronine, estradiol, and testosterone in equine and canine plasma, and of cortisol in equine plasma were not affected by hemolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Electronic probe measurements of cervico-vaginal mucus for detection of ovulation in dairy cows: sanitation, clinical observations and microflora.

An electronic vaginal probe with a sanitizing carrier unit has been designed to measure changes in the electrical resistance of cervico-vaginal mucus in the cow. Over 400 cows and heifers have been probed during the period prior to breeding. When 33 heifers and cows were probed 2-3X per day for 30 days, mild irritation of the vaginal mucosa developed during the luteal phase. In all other studies no grossly detectable lesions or other complications resulting from probing were observed either in experimental herds or in farmer herds where breeding tests were conducted. Fertility of probed cows was equal to cows inseminated when estrus was detected visually. Weekly culture of microorganisms from cervico-vaginal mucus collected from three cows probed 3X per week for 22 days resulted in no detectable change in the population of microorganisms. The number of isolates each successive week was 11, 8, 8 and 5, respectively. The sanitary procedures followed appeared to prevent introduction of organisms of any consequence, as judged by the decreasing number of isolates during the probing sequence.