Neurobiology and pharmacology of Huntington's disease.

Reverse triiodothyronine (rT₃) which is apparently a product of 5-monodeiodination of thyroxine (T₄) at the periphery, was measured in 199 chickens of various strains. rT₃ was virtually absent in young birds (less than 1 week to 8 weeks of age) in marked contrast to the elevated rT₃ levels found in human and other mammalian neonates. At one to two years of age there was a significant increase in the number of birds with detectable rT₃. However, rT₃ concentrations were low and often close to the detection limits of the assay in contrast to significant rT₃ levels found in mammals (man, monkey, sheep, rat and dog). An apparent sex difference in relation to rT₃ formation was noted; 46.4% of 97 females and 9.3% of 54 males showed detectable rT₃ levels. The observations described suggest a species difference in regard to peripheral T₄ monodeiodination between birds and mammals.     

Regulation of thyroid hormone metabolism in rat liver fractions

The nature of the conversion of thyroxine (T₄) to triiodothyronine (T₃) and reverse triiodothyronine (rT₃) was investigated in rat liver homogenate and microsomes. A 6-fold rise of T₃ and 2.5-fold rise of rT₃ levels determined by specific radioimmunoassays was observed over 6 h after the addition of T₄. An enzymic process is suggested that converts T₄ to T₃ and rT₃. For T₃ the optimal pH is 6 and for rT₃, 9.5. The converting activity for both T₃ and rT₃ is temperature dependent and can be suppressed by heat, H₂O₂, merthiolate and by 5-propyl-2-thiouracil. rT₃ and to a lesser degree iodide, were able to inhibit the production of T₃ in a dose related fashion. Therefore the pH dependendy, rT₃ and iodide may regulate the availability of T₃ or rT₃ depending on the metabolic requirements of thyroid hormones.     

Different pathways of iodothyronine 5′-deiodination in rat cerebral cortex

Microsomal fractions of rat cerebral cortex catalyze the 5′-deiodination of 3,3′,5′-triiodothyronine (rT₃) in the presence of thiols such as dithiothreitol. Evidence is presented that two different enzymatic pathways are involved. One of these has a low apparent K_m (2.7 nM) for rT₃, is inhibited by nanomolar concentrations of thyroxine (T₄), but not by up to 1 mM 6-propyl-2-thiouracil (PTU). The other pathway has a high apparent K_m (31 nM) for rT₃, is inhibited by PTU, but not by <1 μM T₄. The relative proportion of rT₃ 5′-deiodination via either pathway depends on thyroid status, with increased contributions from the low-K_m system especially in short-term hypothyroidism.     

Serum principal iodothyronines and the influence of cold exposure associated with shearing in the sheep

The effect of cold exposure caused by shearing on serum thyroid hormone (TH) concentrations in sheep kept at an ambient temperature of 8.5°C was studied. While the deep body temperature fell to the lowest level 4 h after shearing the concentration of triiodothyronine (T₃) increased to a peak value at that time. Thyroxine (T₄) and metabolically inactive reverse triiodothyronine (rT₃) levels reached their peak value after 24 h. The $\text{T}{}_3\text{T}{}_4$ ratio reached a maximum at about 4 h and $\text{rT}{}_3\text{T}{}_4$ and $\text{rT}{}_3\text{T}{}_3$ ratios rose to maximum values about 24 h after shearing. This sequence of events suggest a biphasic response to cold—an immediate secretion of TH from the thyroid gland, followed by adaptive alteration in T₃ and rT₃ generation in the extrathyroidal tissues.     

The effect of propylthiouracil on thyroid-stimulating hormone-induced alterations in iodothyronine secretion from perfused dog thyroids

The aim of this study was to see whether the inhibitory effect of propylthiouracil on thyroidal secretion of 3,5,3′-triiodothyronine (T₃) and 3,3′,5′-triiodothyronine (rT₃) could be reproduced in intensively stimulated thyroids, and to elucidate whether an increase in the fractional deiodination of thyroxine (T₄) to T₃ and rT₃ during iodothyronine secretion might be responsible for the transient fall in the T₄/T₃ and T₄/rT₃ ratios in thyroid secretion seen in the early phase after stimulation of thyroid secretion.For this purpose T₄, T₃ and rT₃ were measured in effluent from isolated dog thyroid lobes perfused in a non-recirculation system using a synthetic hormone free medium. 1 mmol/l propylthiouracil induced a significant reduction in thyroid-stimulating hormone (TSH) stimulated T₃ and rT₃ release while the release of T₄ was unaffected. This supports our previous conclusion that T₄ is partially monodeiodinated to T₃ and rT₃ during thyroid secretion. Infusion of 1 mmol/l propylthiouracil for 30 min or 3 mmol/l propylthiouracil for 120 min did not abolish the transient fall in effluent T₄/T₃ and T₄/rT₃ induced by TSH stimulation. Thus, this phenomenon seems not to depend on intrathyroidal iodothyromine deiodinating processes.     

Serum T4, T3 and reverse T3 in ethanol fed rats.

To investigate the influence of chronic ethanol consumption on circulating thyroid hormone levels, male and female rats were given 20% ethanol as the only drinking solution daily for 8 weeks. Blood ethanol levels ranged 30–45 mg/L. In male rats serum T₄ decreased from the initial mean ± SD value of 5.2±1.4 to3.0 ±0.7 μg/dl; T₃ decreased from initial value of 97±14 to 66±11 ng/dl and rT₃ decreased from initial value of 19±9 to 10±1 ng/dl after 8 weeks of ethanol ingestion. Under similar experimental conditions, female rats showed a significant decrease in serum T₄ and rT₃ levels; however, T₃ levels decreased slightly but not significantly as compared to initial values. The results indicate adverse effect of chronic ethanol intake on serum thyroid hormone levels in rats.     

The crystal structure of a major metabolite of thyroid hormone: 3,3′,5′-triiodo-L-thyronine

Two independent conformations of the thyroinactive thyroid hormone metabolite, 3,3′,5′-triido-L-thyronine (rT₃) were determined by X-ray diffraction methods. The conformations show significant difference in the lettering geometry when compared with those of the thyroactive thyroxine (T₄) and 3,5,3′-triido-L-thyronine (T₃). The diphenyl ether conformation of the two conformers of rT₃ is an anti-skewed one, in which the torsion angels, φ (C5-C4-O4-Cl′) are 8° and ?6°, and φ′ are 86° and 87°. This conformation is in contrast to a twist-skewed one of T₄ and T₃. The difference in the binding abilities between T₄, T₃ and rT₃ to thyroxine binding carrier proteins in serum or to a nuclear receptor protein may be explained by the characteristics solid-state conformations of these metabolites.     

Effect of insulin-induced hypoglycemia on the serum concentrations of thyroxine,triiodothyronine and reverse triiodothyronine

The effect of insulin-induced hypoglycemia on serum thyroid hormone concentrations was studied in nine healthy individuals. Before, during and after the hypoglycemia blood samples were taken for measurement of the concentrations of glucose, thyroxine (T₄), triiodothyronine (T₃), reverse triiodothyronine (rT₃), catecholamines and pituitary hormones.There was no change in the mean serum T₄ level (± the standard error of the mean) of 67 ± 2 μg/l. However, the T₃ concentrations rose from a mean basal level of 1.86 ± 0.06 μg/l to a mean peak of 2.51 ± 0.21 μg/l (P < 0.01) at 45 minutes after the insulin injection, and the rT₃ concentrations fell from a mean basal level of 0.184 ± 0.008 μg/l to a mean nadir of 0.171 ± 0.022 μg/l (not a significant change). The mean peak epinephrine level was 545 ± 103 ng/l and it occurred between 30 and 45 minutes after the insulin injection; the mean peak norepinephrine level was 584 ± 114 ng/l and it occurred between 30 and 90 minutes after the injection. The growth hormone levels reached a mean peak of 26.1 ± 4.8 μg/l and the plasma cortisol levels rose to 215 ± 9 μg/l. The mean basal prolactin level was 8.5 ± 0.9 μg/l; in five subjects there was a rise to a mean peak of 50.6 ± 14.6 μg/l, whereas in the remaining four no significant increase occurred. No correlation was found between the changes in the serum T₃ concentration and any of the other factors studied.It was concluded that acute hypoglycemia is associated with a rapid increase in the serum T₃ concentration.     

Binding of thyroid hormone by erythrocyte cytoplasmic proteins.

Characteristics of iodothyronine-binding to dog erythrocyte cytosol proteins are described. Half-time of association of both thyroxine (T₄) and triiodothyronine (T₃) is 60 min and equilibrium is achieved at 120 min (20°C). Binding is enhanced at 37°C compared to 20°C. T₄ and T₃ binding capacities of the cytosol are 10 and 5 picomoles/mg cytosol protein, respectively. Gel filtration (G-100) reveals 3 protein fractions that dissociably bind both T₄ and T₃. Quantitative and qualitative differences distinguish the erythrocyte cytosol “receptor” proteins from those previously described in other dog tissues. The erythrocyte is a model for studying functions of cytosol “receptors” for iodothyronines.     

Subcellular distribution of iodothyronine 5′-deiodinase in cerebral cortex from hypothyroid rats

We have examined iodothyronine deiodination in subcellular fractions of cerebral cortex obtained from hypothyroid rats. Enzymatic activities were measured at 37°C in the presence of 20 mM dithiothreitol with ¹²⁵I-labeled T₄ and ¹²⁵I-labeled rT₃ as substrate for 5′-deiodination and ¹³¹I-labeled T₃ as the substrate for the 5-deiodinase. Reaction products were separated by descending paper and/or ion-exchange chromatography. Cerebral cortex subcellular fractions were also characterized by marker enzyme analysis and electron microscopy. Under optimal reaction conditions more than 80% of the 5′-deiodinase was recovered after fractionation. Both 5′-deiodinase and (Na⁺ +K⁺-ATPase showed similar subcellular distributions and were enriched approx. 3-fold in the easily sedimenting membrane fraction and nerve terminal plasma membranes. Crude microsomal membranes (6·10⁶g·min pellet) also showed 2-fold enrichment of these enzymes. Nuclei and isolated mitochondria were devoid of deiodinating activity. T₄ and T₃ 5-deiodinating activity was absent in the easily sedimenting membranes and present but not enriched in particulate fractions containing microsomal membranes. These data suggest that iodothyronine 5′-deiodinase is associated with plasma membrane fractions in the cerebral cortex.     

Rapid Responses to Reverse T3 Hormone in Immature Rat Sertoli Cells: Calcium Uptake and Exocytosis Mediated by Integrin

There is increasing experimental evidence of the nongenomic action of thyroid hormones mediated by receptors located in the plasma membrane or inside cells. The aim of this work was to characterize the reverse T₃ (rT₃) action on calcium uptake and its involvement in immature rat Sertoli cell secretion. The results presented herein show that very low concentrations of rT₃ are able to increase calcium uptake after 1 min of exposure. The implication of T-type voltage-dependent calcium channels and chloride channels in the effect of rT₃ was evidenced using flunarizine and 9-anthracene, respectively. Also, the rT₃-induced calcium uptake was blocked in the presence of the RGD peptide (an inhibitor of integrin-ligand interactions). Therefore, our findings suggest that calcium uptake stimulated by rT₃ may be mediated by integrin α_vβ₃. In addition, it was demonstrated that calcium uptake stimulated by rT₃ is PKC and ERK-dependent. Furthermore, the outcomes indicate that rT₃ also stimulates cellular secretion since the cells manifested a loss of fluorescence after 4 min incubation, indicating an exocytic quinacrine release that seems to be mediated by the integrin receptor. These findings indicate that rT₃ modulates the calcium entry and cellular secretion, which might play a role in the regulation of a plethora of intracellular processes involved in male reproductive physiology.     

Phenolic and nonphenolic ring deiodinations of iodothyronines in cultured hepatocarcinoma cell hemogenate from monkey

Iodothyronine monodeiodinase activities in homogenates of cultured monkey hepatocarcinoma cells were measured by the deiodination of [3,5-¹²⁵I]triido-l-thyronine or 3-[3′5′-¹²⁵I]triido-l-thyronine (phenolic ring-labeled ‘reverse’ triiodothyronine). The assay system utilized a small ion-exchange column (AG50W-X4, 0.9×～1 cm) to measure ¹²⁵I^?. Both deiodinases were destroyed by boiling for 1 min.Maximal nonphenolic ring deiodination was observed at pH 7.9 whereas maximal phenolic ring deiodination was at pH 6.3. Both reactions were enhanced strongly by dithiothreitol (0.1–5 mM), and slightly by 5 mM β-mercaptoethanol. Phenolic ring deiodination was strongly inhibited by 0.1 mM propylthiouracil. Nonphenolic ring deiodination was accelerated by EDTA (1.2 mM) and inhibited by Mg²⁺ (5 mM). Methylmercaptoimidazol and Mg²⁺, Ca²⁺ and Mn²⁺ (0.1–1.0 mM) had little or no effect on either reaction, but Zn²⁺ (0.1 mM) strongly inhibited both.Both reactions were inhibited by excess iodothyronine analogues at 10 mM to 10μM, and thyroxine was shown to be a competitive inhibitor in both cases. On the basis of relative affinities and inhibitory effects, it appears that the order of affinity for the phenolic ring deiodinase is 3,3′,5′-triiodo-l-thyronine-(rT₃) > l-thyroxine(T₄) > 3,4,3′-triido-l-thyronine(T₃), whereas for the nonphenolic ring deiodinase the order is T₃ > T₄ > rT₃. Diiodotyrosine did not affect their deiodination.     

Limitations in energy intake affect the ability of young turkeys to cope with low ambient temperatures

Young turkeys exposed to low ambient temperature (T_a) showed significantly reduced body weight, which coincided with a reduction in energy intake and with changes in the circulatory system to accommodate higher oxygen demand. These changes included a significant increase in hematocrit, hemoglobin concentration, plasma triiodothyronine (T₃) concentration, blood volume, and blood oxygen capacity. At the relatively high T_a, changes to accommodate heat dissipation included significant increases in plasma volume and panting rate. These compensations were sufficient to control body temperature (T_b). However, the higher energy expenditure for maintenance followed by significantly higher plasma triiodothyronine (T₃) concentration, but with lower energy intake at low T_a, suggest a physical limitation in the ability to further increase energy intake as T_a declines.     

Triiodothyronine bound to red blood cells is not available for transport through the blood-brain barrier

Many steroid and thyroid hormones and some drugs are bound by circulating red cells. Red cell-bound ligand may not be physiologically inert, as recent studies show that red cell-bound drug is available for uptake by brain. To investigate whether triiodothyronine (T₃) is available for uptake by brain in vivo from the circulating red cell pool, the present investigations measure the effects of human erythrocytes on rat brain uptake of [¹²⁵I]T₃ in vivo. The fraction of circulating T₃ available for uptake in vivo in the presence of 0, 2, 5, 10, 22, or 44% red cells was essentially identical to the fraction of [¹²⁵I]T₃ unbound in vitro. Therefore, [¹²⁵I]T₃ bound to red cells obtained from normal volunteers is not available for uptake by brain in vivo.     

The influence of natural short photoperiodic and temperature conditions on plasma thyroid hormones and cholesterol in male Syrian hamsters

Adult male Syrian hamsters were subjected to 1, 3, 5, 7 or 11 weeks of either natural winter conditions or rigorously controlled laboratory conditions (LD 1014; 22 ± 2C). Although both groups of hamsters gained weight over the course of the experiment, hamsters housed indoors were significantly heavier after 5 weeks of treatment compared to their outdoors counterparts. Animals housed under natural conditions exhibited a significant decrease in circulating levels of thyroxine (T₄) and a rapid rise in triiodothyronine (T₃) levels; the free T₄ and free T₃ index (FT₄I and FT₃I) mirrored the changes in circulating levels of the respective hormones. Laboratory-housed animals had a slight rise in T₄ and FT₄I at 3 weeks followed by a slow steady decline in these values; T₃ and FT₃I values did not change remarkably in these animals. Plasma cholesterol declined steadily over the course of the experiment in laboratory-maintained animals but increased slightly during the first 5 weeks in animals under natural conditions. Since the photoperiodic conditions were approximately of the same duration in these 2 groups, it is concluded that the major differences in body weight, thyroid hormone values and plasma cholesterol are due to some component (possibly temperature) in the natural environment.     

Superiority of the use of a pre-incubated complex of primary and second antibody in the radioimmunoassay of thyroid hormones

We report on the superiority of a radioimmunoassay (RIA) system wherein the second antibody (Ab₂) is incorporated as a pre-incubated complex with the primary antibody (Ab₁) for the assay of haptens like triiodothyronine (T₃) and thyroxine (T₄). Separation of the antibody bound and free antigen (Ag) was accomplished by 8% polyethylene glycol (PEG) (final concentration) following a single incubation of less than 1 h. The other advantages of this system are a 15-fold reduction in the quantity of Ab₂ (without any need for increasing the concentration of PEG) and the consequent savings in cost.     

Harderian glands of golden hamsters: morphological and biochemical responses to thyroid hormones

Summary Manipulation of circulating levels of thyroid hormones modifies Harderian gland structure and porphyrin concentrations in male and female golden hamsters. Specifically, thyroxine (T₄) and triiodothyronine (T₃) induce the morphological conversion of the Harderian glands of females to approximate those of the male. Further, porphyrin concentrations are markedly decreased by this treatment. This effect occurs in ovariectomized animals as well, indicating that the gonads are not involved. Suppression of thyroid function by potassium perchlorate (KClO₄) drastically reduces Harderian gland weight in both males and females. However, KClO₄ decreases porphyrin levels in the Harderian glands of females and increases it in the male. Concurrently, KClO₄ also induces a morphological conversion of the Harderian glands of males to the female type. This effect is evident in photoperiods of either 14:10 (h) or 8:16 (h).     

Hypohydration increases the plasma catecholamine response to moderate exercise in the dog (Canis)

1. The effects of hypohydration produced by 48 hr water deprivation were examined in dogs during moderate treadmill exercise at an ambient temperature (T_a) of 21°C.2. Hypohydration caused a significant elevation in plasma levels of adrenaline (A), proteins (pp) and osmolality (pOsm).3. During 1 hr of running, plasma concentrations of adrenaline (A) and noradrenaline (NA) rose significantly, whilst no change in these hormones occurred in dogs hydrated ad libitum.4. The results suggest that hypovolemia in the dog may be a sufficient stimulus to intensify the sympatho-adrenal response to moderate exercise performed at a room T_a.