Metyrapone - reduced cytochrome P-450 complex: A specific method for the determination of the phenobarbital inducible form of rat hepatic microsomal cytochrome P-450

Using homogeneous cytochrome P-450, we have shown that the well-known metyrapone-dithionite reduced cytochrome P-450 complex is specific for the cytochrome P-450b induced by phenobarbital. A linear relationship was observed between the absorbance of metyrapone-reduced cytochrome P-450 complex and the one of CO-reduced cytochrome P-450 complex, the usual method for the determination of cytochrome P-450. A method has been proposed for the specific determination of the cytochrome P-450b.     

Iodide-induced inhibition of adenylate cyclase activity in horse and dog thyroid

The characteristics of the iodide-induced inhibition of cyclic AMP accumulation in dog thyroid slices have been previously described [Van Sande, J., Cochaux, P. and Dumont, J. E. (1985) Mol. Cell. Endocrinol. 40, 181-192]. In the present study we investigated the characteristics of the iodide-induced inhibition of adenylate cyclase activity in dog and horse thyroid. The inhibition of cyclic AMP accumulation by iodide in stimulated horse thyroid slices was similar to that observed in dog thyroid slices. The inhibition was observed in slices stimulated by thyroid-stimulating hormone, cholera toxin and forskolin. Increasing the concentration of the stimulators did not overcome the iodide-induced inhibition. Adenylate cyclase activity, assayed in crude homogenates or in plasma-membrane-containing particulates (100,000 x g pellets), was lower in homogenates or in particulates prepared from iodide-treated slices than from control slices. This inhibition was observed on the cyclase activity stimulated by forskolin, fluoride or guanosine 5'-[beta, gamma-imino]triphosphate, but also on the basal activity. It was relieved when the homogenate was prepared from slices incubated with iodide and methimazole. Similar results were obtained with dog thyroid. The inhibition persisted when the particulate fraction was washed three times during 1 h at 100,000 x g, in the presence of bovine serum albumin or increasing concentration of KCl. It was similar whatever the duration of the cyclase assay, in a large range of protein concentration. These results indicate that a stable modification of adenylate cyclase activity, closely related to the plasma membrane, was induced when slices were incubated with iodide. Iodide inhibition did not modify the affinity of adenylate cyclase for its substrate (MgATP), but induced a decrease of the maximal velocity of the enzyme. The percentage inhibition was slightly decreased when Mg2+ concentration increased, and markedly decreased when Mn2+ concentration increased. A detectable adenylate cyclase activity was demonstrated when intact slices were incubated in the presence of [alpha-32P]ATP, probably because of the presence of broken cells produced during the slicing. Iodide had no direct effect on this cyclase system, which confirms that iodide needs the integrity of the cell to induce the inhibition and suggests that the inhibition is not transmitted between cells.     

Comparison of the teratogenic properties manifested in BALB c mice, by diphenylhydantoin and deuterated diphenylhydantoin

The aim of this study was to investigate the role of an arene oxide pathway in the teratogenicity displayed by DPH, a highly effective antiepileptic agent. This approach was carried out by comparing the teratogene potential of DPH and of p2-H-DPH administrated at days 8, 9 and 12 of gestation. The present findings support the hypothesis that substitution of protium by deuterium at the para position of one of the phenyl rings, which favours an arene oxide pathway, causes an increase in some to the teratogenic effects of DPH.     

Increased preproenkephalin A gene expression in the rat heart after induction of a myocardial infarction.

The expression of preproenkephalin A (ppENK) gene was investigated in the rat heart, following the onset of myocardial infarction induced by ligation of the left anterior descending coronary artery. The relative abundance of ppENK mRNA and the level of enkephalins were measured by Northern blot analysis and radioimmunoassay, respectively, in the ventricles from control-unoperated, sham-operated, and operated rats. Three hours after the surgery, a comparison between rats with infarction and sham-operated rats revealed that the relative abundance of ppENK mRNA and the level of enkephalins were increased three- to four- and two- to three-fold, respectively, in the ventricles of rats with infarction. No difference was observed between rats with infarction and sham-operated rats 24 h after the surgery, or between rats with infarction compared at time intervals of 3 and 24 h following the surgery. The abundance of the ppENK mRNA in the polysomal fraction of the ventricular septum was also measured 3 h after the surgery and found to be threefold higher in rats with infarction as compared with sham-operated rats. These results indicate that the level of enkephalins rapidly increases in the ventricles of rats following myocardial infarction, and that this higher level may be ascribed to a stimulation of the local synthesis of enkephalins.     

Negative effect of iodide on the survival of newly divided epithelial cells in chronically stimulated rat thyroid

The aim of this work was to investigate some aspects of the thyroid epithelial cell kinetics during the iodide-induced involution of a hyperplastic goitre in the rat. Rats were made iodine-deficient for 6 months, and propylthiouracil (PTU) (0.15%) was added to the diet during the last 2 months. Thereafter, rats were refed with iodide and PTU was removed (day 0). Forty-eight hours previously, all the rats were injected with tritiated thymidine ([3H]TdR) (1 microCi/g). Some animals were killed 1 hr or 24 hr after [3H]TdR injection (i.e. on day -2 and -1, day 0 corresponding to the restoration of a normal iodine diet); the other animals were killed after different delay periods and following [3H]TdR injection. Autoradiography of thyroid sections, iodine determination of plasma iodide and protein-bound iodine (PBI), and RIA of plasma thyroid stimulatory hormone (TSH) were performed. Plasma TSH concentration was very high on day 0 of iodide refeeding (3000 +/- 330 ng/ml) and remained at this level until day 8. Plasma PBI was very low on day 0, remained so until day 4 and greatly increased on day 8. Plasma iodide was also very low on day 0, but markedly increased on day 1, then did not vary significantly until day 43 of iodine refeeding. Thyroid weight, elevated on day 0, decreased relatively quickly until day 30, then more slowly until day 73. The [3H]TdR labelling index (LI) of the thyroid epithelial cells (TEC) was high on day 0 (56 +/- 3 labelled cells/10,000 cells), and 24 hr thereafter increased to 104 +/- 3, by division of the labelled cells. On day 1 of iodine refeeding, the LI had abruptly decreased to about half this value and then remained stable for 3 more days. Between day 4 and day 16, a progressive decline in the LI, (by about 3-4 per day), was observed. The LI showed no further modification, up to day 73, the longest period investigated. The decrease in LI occurred without any significant changes in the labelling intensity (grain count) of the remaining labelled cells between day 1 and 16, this indicates that no cell division took place during this period. The data are therefore interpreted as showing a biphasic elimination after iodide refeeding, of cells that were actively proliferating during the goitrous state.(ABSTRACT TRUNCATED AT 400 WORDS)     

Carbamylcholine, TRH, PGF2 alpha and fluoride enhance free intracellular Ca++ and Ca++ translocation in dog thyroid cells

Effects on Ca++ translocation and [Ca++]i were studied in dog thyro?d cell monolayers using both 45Ca++ efflux and the indicator quin-2. Carbamylcholine, a non hydrolysable analog of acetylcholine, through muscarinic receptors, and to a lesser extent TRH and PGF2 alpha increased both these parameters. [Ca++]i increased by 171, 100 and 75% respectively over a basal level of 66 +/- 17 nM (mean +/- SD). The response to carbamylcholine was biphasic. A transient increase in [Ca++]i was followed by a more sustained phase where the [Ca++]i was slightly higher than the basal level. Only the first phase was insensitive to extracellular Ca++ depletion. This phase is probably due to a release of Ca++ from an intracellular store. NaF also induced a sustained rise in [Ca++]i dependent on extracellular Ca++ and affected 45Ca++ efflux. Our data provide direct evidence of an implication of intracellular Ca++ in the response of dog thyro?d cells to all these agents.