Use of Nonconventional Sample Matrices for Endocrine Studies of Pit Vipers: Assay Validation and Applications

Investigation of keratinase digestion to improve steroid hormone extraction from diverse keratinous tissues

In order to clarify the functional relationship between thyroid, adrenal and gonadal hormones, hypothyroidism was induced by administration of thiuoracil in adult male and female rats, and the effects of hypothyroidism on the adrenal and the gonadal axes were investigated in the present study. 1. The functional relationship between thyroid and adrenal hormones: Adrenal weights and corticosterone were lowered, whereas the secretion of ACTH, corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP) increased in hypothyroid rats compared to euthyroid rats. These results indicate that hypothyroidism causes adrenal dysfunction directly and results in hypersecretion of CRH and AVP from the hypothalamus. 2. The functional relationship between thyroid and gonadal hormones: The pituitary response to LHRH was lowered, whereas the testicular response to hCG was not changed in hypothyroid rats. Hypothyroidism suppressed copulatory behavior in male rats. These results suggest that hypothyroidism probably causes dysfunction in gonadal axis at the hypothalamic-pituitary level in male rats. In adult female rats, hypothyroidism inhibited the follicular development accompanied estradiol secretion, whereas plasma concentrations of progesterone and prolactin (PRL) increased in hypothyroid female rats. Hypothyroidism significantly increased the pituitary content of vasoactive intestinal peptide (VIP) though it did not affect dopamine synthesis. These results suggest that hypothyroidism increases pituitary content of VIP and this increased level of VIP likely affects PRL secretion in a paracrine or autocrine manner. In female rats, inhibition of gonadal function in hypothyroid rats mediated by hyperprolactinemia in addition to hypersecretion of endogenous CRH.

The hippocampal formation: Morphological changes induced by thyroid, gonadal and adrenal hormones

Recent studies have demonstrated that some hormones are present in baleen powder from bowhead (Balaena mysticetus) and North Atlantic right (Eubalaena glacialis) whales. To test the potential generalizability of this technique for studies of stress and reproduction in large whales, we sought to determine whether all major classes of steroid and thyroid hormones are detectable in baleen, and whether these hormones are detectable in other mysticetes. Powdered baleen samples were recovered from single specimens of North Atlantic right, bowhead, blue (Balaenoptera [B.]musculus), sei (B. borealis), minke (B. acutorostrata), fin (B. physalus), humpback (Megaptera novaeangliae) and gray (Eschrichtius robustus) whales. Hormones were extracted with a methanol vortex method, after which we tested all species with commercial enzyme immunoassays (EIAs, Arbor Assays) for progesterone, testosterone, 17β-estradiol, cortisol, corticosterone, aldosterone, thyroxine and tri-iodothyronine, representing a wide array of steroid and thyroid hormones of interest for whale physiology research. In total, 64 parallelism tests (8 species × 8 hormones) were evaluated to verify good binding affinity of the assay antibodies to hormones in baleen. We also tested assay accuracy, although available sample volume limited this test to progesterone, testosterone and cortisol. All tested hormones were detectable in baleen powder of all species, and all assays passed parallelism and accuracy tests. Although only single individuals were tested, the consistent detectability of all hormones in all species indicates that baleen hormone analysis is likely applicable to a broad range of mysticetes, and that the EIA kits tested here perform well with baleen extract. Quantification of hormones in baleen may be a suitable technique with which to explore questions that have historically been difficult to address in large whales, including pregnancy and inter-calving interval, age of sexual maturation, timing and duration of seasonal reproductive cycles, adrenal physiology and metabolic rate.

Determination of plasma kisspeptin concentrations during reproductive cycle and different phases of pregnancy in crossbred cows using bovine specific enzyme immunoassay

Validation of a shed skin corticosterone enzyme immunoassay in the African House Snake (Lamprophis fuliginosus) and its evaluation in the Eastern Massasauga Rattlesnake (Sistrurus catenatus catenatus)

Glucocorticoid receptor mediates corticosterone-thyroid hormone synergy essential for metamorphosis in Xenopus tropicalis tadpoles

Aphasia in a farmer after viper bite

Dorper is a South African sheep breed characterized by its fast growth and excellent body confirmation that is used for mutton production. Hematological profiling is useful in assessing the health, nutritional, and metabolic status of an animal and can be affected by non-genetic factors such as sex, age, season, body condition score, and physiological status of an animal. Hence, this study focused on the effect of the factors mentioned above on the hematological parameters of Dorper sheep. Sex, age, physiological status, and body condition score for 30 (11 males and 19 females) Dorper sheep were recorded, and blood samples were collected from each sheep. Red blood cell count (RBC), white blood cell count (WBC), haemoglobin (Hb), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), absolute neutrophil count (ANC), absolute monocyte count (AMC), absolute lymphocyte count (ALC), lymphocyte count (LYM), normoblast count (NB), platelet count (Plt), haematocrit (Hct), eosinophils (Eos), monocytes (Mn), red blood cell distribution width (RDW), basophil count (BASO) and absolute eosinophil count (AEC) were determined. Pearson correlation analysis showed a correlation (p<0.05) between some of the haematological parameters in Dorper sheep. Pearson correlation analysis results indicated that RBC was positively highly (p<0.01) correlated with Hct, WBC, ALC, Eos, and AEC and negatively highly correlated (p<0.01) with MCV, Mn, and NB. However, RBC had no significant (p>0.05) correlation with MCHC, RWD, NEU, ANC, AMC, BASO, ABC, and Plt in Dorper sheep. Body condition score and age had no effects (p>0.05) on the haematological parameters (p>0.05), while physiological status affected only RDW. The results further showed that sex influenced (p<0.05) Hb, RDW, LYM, and NB. The findings of this study suggest that sex and physiological status can influence some of the haematological parameters of Dorper sheep. Therefore, it is important to consider factors such as sex and physiological status of an animal when evaluating the nutritional, physiological and health status to improve the management and productivity of Dorper sheep.

Thyroid hormones in small ruminants: effects of endogenous, environmental and nutritional factors

Glucocorticoids play a key role in mediating stress responses in vertebrates. Corticosterone (CORT) is the main glucocorticoid produced in amphibians, birds, and reptiles, and regulates several metabolic functions. The most common methods for quantifying CORT are competitive binding immunoassays: radioimmunoassay (RIA) and enzyme immunoassay (EIA). RIA has been broadly used since the 1980’s but it requires radioactivity. Commercial EIA kits permit quantifying hormone levels without radioactivity although the requirement for a larger sample volume may be a strong limitation for measurements involving larval amphibians. Here we used Xenopus laevis tadpoles to compare the performance of three commonly used procedures for determination of CORT: RIA on a chloroform extract of whole-body homogenate, EIA on plasma, and EIA on supernatant of whole-body homogenate. We treated tadpoles with exogenous CORT at 0, 25, 50, and 100 nM. RIA could distinguish between 0 and 25 nM, and EIA on plasma between 0 and 50 nM, whereas whole-body homogenate EIA only detected significant differences between 0 and 100 nM. Each procedure presents advantages and disadvantages regarding sensitivity, the use of radioactivity, sample size, handling time, and economic cost. RIA is preferred when studying small-bodied animals from which blood samples cannot be obtained. When CORT level differences are intermediate and blood sampling is possible, EIA on plasma is a good non-radioactive alternative. EIA on whole-body homogenates may be useful to assess qualitative changes in CORT levels when considerable differences are expected. Finally, we discuss our findings in the context of previous studies on CORT in amphibians.

Mammals can use a variety of physiological mechanisms to adapt to changes in their environment. Thyroid hormones (THs) are key modulators of growth and mediators of environmental conditions by regulating developmental processes and metabolism in animals. In recent years, advancements in non-invasive sampling have allowed monitoring of the fluctuations of THs and their metabolites in wild mammals. Triiodothyronine (T3) represents the major metabolite of THs excreted in feces so that it can be monitored in fecal samples. In this study, fecal samples collected during the hunting season from legally culled European roe deer (Capreolus capreolus; n = 160) were assayed to investigate the potential fluctuations of fecal TH metabolites (FTMs) in response to environmental (e.g., the temperature, local densities) and individual (e.g., sex, age, body, and nutritional conditions) variables. For this aim, we validated a TH enzyme immunoassay in the feces of roe deer. Our results show that FTMs can be successfully measured with satisfactory accuracy and precision. Extraction recovery (70%-120%), intra- and inter-day repeatability (<15%), linearity dilutions (80%-120%), and parallelism (<20%) were consistent with international guidelines. Environmental temperature (p < 0.001) showed a strong inverse correlation with FTM levels. THs can thus represent a reliable indicator in studying animals' adaptative responses to environmental temperature changes, providing perspectives for the study of the impact of climate change on ungulates and mammals. Further analyses, comparing samples collected all year round, are needed to investigate the correlations of TH values versus the other investigated variables.

Antivenom cross neutralisation in a suspected Asian pit viper envenoming causing severe coagulopathy

Administration of exogenous thyroid hormones precociously increase intestinal sucrase activity (SA) in suckling rats. To differentiate the effects of thyroid and adrenal hormones on SA, intact suckling Sprague-Dawley rats were given TSH (0.5U) bid or water subcutaneously from day 14 until sacrifice on day 18. Results are shown below (x+SEM). Serum T4 and corticosterone (CS) as well as SA in the jejunum (J), mid-jejunum (MJ) and ileum (I) were all significantly increased in intact TSH treated rats.Sucklings were then adrenalectomized (ADX) on day 14 and given TSH (0.5 or 1.0U) bid or water subcutaneously until sacrifice on day 18. No significant differences in SA were found between ADX sucklings given TSH(1 or 2U/day) and ADX controls given water only.In the absence of the adrenal glands, physiologic elevation of endogenous thyroid hormones by TSH does not alter the maturational time course of intestinal sucrase activity.

Molecular mechanisms of corticosteroid synergy with thyroid hormone during tadpole metamorphosis