Angel A. Zaninovich

Effect of melatonin treatment on oxygen consumption by rat liver mitochondria

Summary.The objective of this study was to examine the in vivo effect of melatonin on rat mitochondrial liver respiration. Two experiments were performed: For experiment 1, adult male rats received melatonin in the drinking water (16 or 50 µg/ml) or vehicle during 45 days. For experiment 2, rats received melatonin in the drinking water (50 µg/ml) for 45 days, or the same amount for 30 days followed by a 15 day-withdrawal period. At sacrifice, a liver mitochondrial fraction was prepared and oxygen consumption was measured polarographically in the presence of excess concentration of DL-3-β-hydroxybutyrate or L-succinate. Melatonin treatment decreased Krebs’ cycle substrate-induced respiration significantly at both examined doses. The stimulation of mitochondrial respiration caused by excess concentration of substrate recovered after melatonin withdrawal. Basal state 4 respiration was not modified by melatonin. Melatonin, by curtailing overstimulation of cellular respiration caused by excess Krebs’ cycle substrates, can protect the mitochondria from oxidative damage.

Evidence suggesting that the sympathetic nervous system mediates thyroidal depression in turpentine-induced nonthyroidal illness syndrome.

Acute superior cervical ganglionectomy (SCGx) induces in the rat a supraliminal release of neurotransmitter in the innervated tissues (i.e., thyroid gland). This temporary adrenergic hyperactivity is correlated with a significant depression of the thyroid economy resembling the nonthyroidal illness (NTI) syndrome in the rat, and suggest that the sympathetic nervous system may mediate thyroidal changes in NTI. In order to gain further insight into the thyroidal depression in the NTI syndrome, we studied the thyroidal norepinephrine (NE) turnover in turpentine oil (TURP)-induced NTI syndrome and the role of the cervical ganglia (SCG) in the development of NTI in the rat. TURP administration to sham operated rats induced a rapid and significant fall in plasma T4 and TSH levels, in the thyroidal response to exogenous TSH (TIU) and in the thyroidal NE content compared to controls (sham + saline) (T4: 3.1 +/- 0.3 vs. 5.1 +/- 0.6 micrograms/dl, respectively, mean +/- SE, p less than 0.02; TSH: 1.4 +/- 0.4 vs. 4.7 +/- 1.4 ng/ml, respectively, p less than 0.05; TIU: 92 +/- 14 vs. 201 +/- 20 cpm.microliter thyroid/cpm.mg plasma (T/P ratio), respectively, p less than 0.01; thyroidal NE: 680 +/- 20 vs. 761 +/- 29 pg/mg thyroid, respectively, p less than 0.05). The thyroidal turnover rate of NE, however, was significantly increased in TURP-injected rats compared to controls (122 +/- 13 vs. 86 +/- 10 pg/mg/h, respectively, p less than 0.05). TURP injection to chronic SCGx rats induced a similar fall in plasma TSH compared to controls (SCGx + saline) (1.3 +/- 0.2 vs. 4.3 +/- 1.1 ng/ml, respectively, p less than 0.02); plasma T4 and TIU, however, did not change significantly (T4: 3.4 +/- 0.4 vs. 3.7 +/- 0.3 micrograms/dl, respectively, NS; TIU: 172 +/- 8 vs. 226 +/- 27 T/P ratio, respectively, NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of diabetes, β-hydroxybutyric acid and metabolic acidosis on the pituitary-thyroid axis in the rat

Previous studies demonstrated alterations of thyroidal economy in untreated diabetes mellitus both in man and experimental animals. To test the role of β-hydroxybutyric acid (BHB) and acidosis in generating such changes, we studied the pituitary-thyroid axis of streptozotocin-diabetic rats, BHB or ammonium chloride (NH4CI)-treated normal rats. Serum TSH, pituitary content and pituitary concentration of TSH, serum T4, T3 and free T4 (FT4), were all measured by RIA. In short term (2 days) diabetic rats the pituitary content of TSH was normal whereas the concentration (per mg of protein) was elevated (p< 0.05 versus control group). Serum TSH (p< 0.05), serum T4 (p< 0.05), serum T3 (p<0.01) and serum FT4 (p<0.05) were all significantly decreased. In long term (30 days) untreated diabetic rats serum changes were similar to the short term diabetic group, though the pituitary content of TSH was significantly decreased (p< 0.05). Animals treated with NH4Cl had no variations from controls. However, rats treated with BHB displayed a significant decrease in pituitary content of TSH (p< 0.05), pituitary concentration of TSH (p< 0.05) and in plasmaTSH (p< 0.01 ), and normal thyroid hormones in serum. No significant changes were seen in theTSH response toTRH in 2 or 30 days untreated diabetic and in BHB — treated animals. The data suggest that BHB, although not NH4CI acidosis, may be capable of inducing a moderate depression of pituitary and plasma TSH of a lesser magnitude of that accompanying the full, long term diabetic state in the rat.

Effects of Thyroxine on Rat Brown Fat and Muscle Thermogenesis in the Cold

We studied whether the activation of rat brown adipose tissue (BAT) by cold exposure or by the administration of beta-3-noradrenergic agonist CGP-12177 could be prevented by the inhibition of thyroxine (T4) to triiodothyronine (T3) conversion. Hypothyroid rats were treated with replacement doses of T4, T4 plus iopanoic acid (IA) or T3. Groups of rats were placed at 4°C for 24 h or kept at room temperature. Cold exposure induced a significant increase in guanosine diphosphate (GDP) binding to BAT mitochondrial proteins in T4-treated rats, an effect not abolished by IA. No significant changes were seen in T3-treated rats. In rats maintained at room temperature and injected with CGP-12177, T4 induced a significant rise in GDP binding which was not blocked by IA. T3 also induced a significant increase in binding. The study of mitochondrial oxygen consumption in muscle from cold-exposed rats showed a marked decrease in consumption in T3-treated rats as compared to values in the warm. Normal oxygen consumption was restored with 2-fold doses of T3 replacement, whereas 5-fold doses increased consumption above normal. The data suggest that in states with low or absent T3, T4 can stimulate heat production and preserve normothermia.

Reciprocal effects of an acute load of thyroxine or triiodothyronine on their peripheral metabolism and deiodination in the cold-acclimated rat

Wistar rats acclimated to cold received iv 25μg T4/100g bw, 21 μg T3/100g bw or 2.5 μg T3/100g bw, to observe the changes induced in the peripheral metabolism of 125I-T4 and 125I-T3 relative to cold-adapted untreated animals. Each animal was injected with a tracer dose of either labelled hormone in addition to the T4 or T3 load, and placed in metabolic cages for 24 hour collection of urine and feces. Sequential heparinized blood samples were obtained by cardiac puncture. A T4 load decreased the deiodination of 125I-T4 and 125I-T3 (p < 0.01) as revelated by urinary 125I. A T3 load, in the two dosages employed, decreased the deiodination of 125I-T3(p < 0.01) but had no effect on deiodination of 125I-T4. Similarly, a T4 load increased the fecal excretion of both radioactive iodothyronines (p < 0.01) whereas a T3 load failed to alter the excretion of 125I-T4. In cold-adapted animals plasma TSH was elevated (p < 0.05) and plasma T4 was low (p < 0.001) as compared to rats housed at 22 C. It is concluded that the relative contribution of T4 and T3 to the metabolic state in the rat is not significantly altered by cold exposure.

NEUROPEPTIDE Y SUPPRESSED THE T3-INDUCED RISE IN BROWN FAT MITOCHONDRIAL RESPIRATION IN HYPOTHYROID RATS

It has been postulated that a cold-induced decrease in hypothalamic neuro-peptide Y (NPY) might disinhibit the sympathetic outflow that activates brown adipose tissue (BAT) thermogenesis. The present work has assessed the interscapular BAT NPY and norepinephrine (NE) concentrations 7 days after surgical excision of the sympathetic nerves (Sx) of each BAT lobe in rats exposed to 4°C or 24°C during 24 h. In addition, the effects of NPY treatment on BAT oxygen (O2) consumption in normal and T3-treated hypothyroid rats was determined. Sprague-Dawley rats weighing 220–240 g were used. BAT was removed and homogenized in 0.1 M HCl. BAT NPY content at 24°C was 109 ± 19 pg/mg protein. Sx decreased this value significantly (P < 0.001) and cold increased it by two-fold (P < 0.001). Neither cold nor Sx altered hypothalamic NPY. BAT O2 consumption was depressed by NPY treatment (P < 0.001) and increased by NE (P < 0.001). In euthyroid Sx rats, NPY decreased O2 consumption (P < 0.001). NPY failed to alter O2 consumption in hypothyroid rats and sharply decreased it in T3-treated rats. The results show that NPY blocked the effects of T3 on BAT O2 consumption, and that cold-induced activation of the thermogenic process did not produce measurable changes in hypothalamic NPY.