Background and Purpose-The neuroprotective role of mild therapeutic hypothermia was established in animal models of cerebral ischemia. Still, several issues, including optimal target temperature, remain unclear. The optimal depth of hypothermia in a rat model of focal cerebral ischemia was investigated. Methods-Eighty-four male Wistar rats (nϭ84) were subjected to filament occlusion of the middle cerebral artery for 90 minutes. Sixty animals were equally split into 6 groups kept at core temperatures of 37°C, 36°C, 35°C, 34°C, 33°C, and 32°C over a period of 4 hours starting 90 minutes after middle cerebral artery occlusion. Twenty-four hours later, after performing a neuroscore, animals were killed and brains examined for infarct size, edema, and invasion of leukocytes.In the second part, 24 animals (8 per group) were kept at 33°C, 34°C, and 37°C for 4 hours, allowed to survive for 5 days, and underwent additional investigation of transferase dUTP nick-end labeling. Results-In the first part, one animal in each treatment group and 2 animals in group 37°C died. The infarct size and edema were smaller for 34°C and 33°C compared with all other groups (PϽ0.05) over 24 hours. These animals also had better functional outcome (PϽ0.05) with an advantage for 34°C versus 33°C (PϽ0.05). Leukocyte count was lower for 34°C and 33°C as compared with the 37°C group. Similar results were obtained in the second part of the study with an advantage for 34°C versus 33°C. Conclusion-Our results suggest that the optimal depth of therapeutic hypothermia in temporary middle cerebral artery occlusion is 34°C.

Amphibian metamorphosis represents a promising model for the identification of thyroid system-disrupting chemicals due to the pivotal role played by thyroid hormones for the initiation and regulation of metamorphosis. An important aspect of bioassay development is the identification and evaluation of sensitive and diagnostic endpoints. In this study, several morphological, histological, and molecular endpoints were evaluated for their utility to detect alterations in thyroid system function after exposure of stage 51 Xenopus laevis tadpoles to various concentrations (1.0, 2.5, 10, 25, and 50 mg/l) of the anti-thyroidal compound ethylenethiourea (ETU). Analysis of developmental stages on exposure day 20 and monitoring of time to fore limb emergence (FLE) revealed retardation and complete arrest of tadpole development at 25 mg/l and 50 mg/l ETU, respectively. Development was not affected by 1.0, 2.5, and 10 mg/l ETU. Histological alterations in the thyroid gland were observed in FLE-displaying tadpoles after exposure to 2.5, 10, and 25 mg/l ETU, as well as in developmentally arrested tadpoles exposed to 50 mg/l ETU. Prevalence and severity of histological changes increased in a concentration-dependent manner. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) showed increased mRNA expression of the alpha- and beta-subunits of thyroid-stimulating hormone (TSHalpha, TSHbeta) in pituitary tissue of tadpoles exposed to 25 and 50 mg/l ETU. Results demonstrate the successful detection of anti-thyroidal effects of ETU in Xenopus laevis tadpoles using various endpoints and highlight the particular sensitivity of thyroid gland histology to detect thyroid system disruption in tadpoles.

The uptake of iodide represents the first step in thyroid hormone synthesis by thyroid follicular cells and is mediated by the sodium-iodide symporter (NIS). In mammals, expression of NIS is stimulated by TSH and transcription of the NIS gene involves regulation by the thyroid-specific transcription factors Pax8 and Nkx2$1. In this study, we examined the mRNA expression of NIS, Pax8 and Nkx2$1 in the thyroid gland of Xenopus laevis tadpoles by semiquantitative reverse transcriptase (RT)-PCR. During spontaneous metamorphosis, NIS mRNA expression was low in premetamorphic tadpoles, increased throughout prometamorphosis, and peaked at climax stage 60. Analysis of TSH bsubunit (TSHb) mRNA in the pituitary of the same tadpoles revealed a close temporal relationship in the expression of the two genes during metamorphosis, suggesting a regulatory role of TSH in the developmental expression of NIS. Treatment of tadpoles with goitrogenic compounds (sodium perchlorate and ethylenethiourea) increased TSHb mRNA expression (approximately twofold) and caused thyroid gland hyperplasia, confirming that feedback along the pituitary-thyroid axis was operative. Analysis of gene expression in the thyroid gland revealed that goitrogen treatment was correlated with increased expression of NIS mRNA (w20-fold). In the thyroid gland organ culture experiments, bovine TSH (bTSH; 1 mU/ml) strongly induced NIS mRNA expression. This effect was mimicked by co-culture of thyroid glands with pituitaries from stage 58 tadpoles and by agents that increase intracellular cAMP (forskolin, dibutyryl-cAMP). In addition, it could be shown that thyroid glands of X. laevis tadpoles express Pax8 and Nkx2$1 mRNA in a developmentally regulated manner and that ex vivo treatment of thyroid glands with bTSH, forskolin, and cAMP analogs increased the expression of Pax8 and Nkx2$1 mRNA. This is the first report on developmental profiles and hormonal regulation of thyroid gland gene expression in amphibian tadpoles and, together, results reveal a critical role of TSH in the regulation of NIS mRNA expression in the thyroid gland of X. laevis tadpoles.