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Biomedical subjects

A Pascual

Publications and source records attributed to A Pascual.

At least 199 records · Page 11Linked to original sources
PubMed1986

Regulation of glycerol phosphate dehydrogenase and lactate dehydrogenase activity by forskolin and dibutyryl cyclic AMP in the C6 glial cells.

We have compared the effects of norepinephrine, forskolin, and dibutyryl cyclic AMP (Bt2cAMP) on the regulation of the cytosolic enzyme glycerol phosphate dehydrogenase (GPDH) in the C6 rat glioma cell line. Forskolin and Bt2cAMP elicit a dose-dependent increase in the levels of the enzyme that was, however, unaffected by norepinephrine. The half-maximal effect of forskolin was obtained at 7-8 microM, and the effect was maximal at 30 microM. Dexamethasone at a 50 nM concentration produced a two- to sixfold induction of GPDH after 48 h. The combination of dexamethasone with forskolin or Bt2cAMP leads to an elevation in GPDH levels that is higher than that produced by one of the compounds alone. This potentiation is found when both agents are added together with or after the glucocorticoid. The increase in uninduced and dexamethasone-induced GPDH activity was blocked by cycloheximide and actinomycin D, indicating that de novo protein and RNA synthesis are required. The activity of cytosolic lactate dehydrogenase activity did not change after incubation with dexamethasone, but increased with forskolin or Bt2cAMP.

Animals↗
PubMed1986

Effect of medium, pH, and inoculum size on activity of ceftizoxime and Sch-34343 against anaerobic bacteria.

The effect of two media, three pH values, and three inoculum densities on the activity of ceftizoxime and Sch-34343 against anaerobic bacteria was evaluated. The activities of both antimicrobial agents were affected by medium composition, especially against Bacteroides fragilis. Changes in pH and inoculum size affected only the activity of ceftizoxime against anaerobic bacteria.

Anti-Bacterial Agents↗
PubMed1986

Triiodothyronine receptors in developing mouse neuronal and glial cell cultures and in chick-cultured neurones and astrocytes.

The evolution of L-triiodothyronine (T3) receptors was studied in developing cultures of cells dissociated from cerebral hemispheres of 14-day-old mouse embryos, which present successive distinct periods of cell proliferation and/or maturation. These periods are characterized essentially as neuronal from 1 to 12 days in vitro (DIV) and glial between 12 and 60 DIV. Furthermore myelin-related membranes are produced in this culture system. Binding capacities of the T3 nuclear receptors increased from 1 to 6 DIV, when it reached a maximum (16 fmol/100 micrograms DNA). A similar increase of the DNA content of the cell was observed until 8 DIV. Thereafter a sharp fall of receptor concentration leading to a 5-fold decrease in the binding capacity occurred until day 15, a period at which neurones disappeared from the cultures. From 25 to 50 DIV (coinciding with the glial period), the concentration of receptor remained more or less constant (1-2 fmol/100 micrograms DNA). In parallel, the DNA content did not vary greatly between 30 and 50 DIV. Scatchard analysis revealed the presence of a single class of receptors at 6 and 20 DIV, representative of 'neuronal' and 'glial' periods, respectively. The equilibrium dissociation constant (Kd) of the nuclear receptor from cells at 6 DIV (2 X 10(-10) M) was similar to that found at 20 DIV. These results were confirmed using pure cultured neurones and astrocytes prepared from embryonic chick brain. The effect of T3 on the cellular gangliosides used as an index of neuronal cell maturation, and on cerebroside sulfotransferase (CST), an enzyme involved in the production of myelin sulfatides, was studied to determine a possible correlation between the binding capacity of the T3 nuclear receptor and the response of the cultured cells to thyroid hormone. Our data demonstrate that T3 had no significant effect either on the content of gangliosides or on their developmental pattern, while it increased the level of CST activity by 75% between 18 and 25 DIV. These results show that, although the concentration of T3 receptors per 100 micrograms DNA in glial cells was lower than that in neurones, it was nevertheless sufficient to elicit a response in oligodendrocytes.

Animals↗
PubMed1986

Identification and characterization of L-triiodothyronine receptors in cells of glial and neuronal origin.

High affinity-low capacity binding sites for thyroid hormone have been identified in the nuclei of glial (C6) and neuronal (Neuro 2A) cultured cells. Equilibrium dissociation constants, determined by Scatchard analysis, were very similar in both types of cells (0.2-0.3 nM). The relative affinity of hormonal analogs was also similar: the affinity for T3 was lower than for triiodothyroacetic acid and higher than for T4 or tetraiodothyroacetic acid. The sedimentation coefficients obtained by gradient centrifugation of nuclear receptor extracted with 0.4 M KCl or excised by micrococcal nuclease digestion were 3.5 S and 6.5 S, respectively. These results suggest that the thyroid hormone receptor is not restricted to neuronal cells, but also appears in cells of glial origin.

Animals↗
PubMed1984

Photoaffinity labeling of thyroid hormone nuclear receptors. Influence of n-butyrate and analysis of the half-lives of the 57,000 and 47,000 molecular weight receptor forms.

The thyroid hormone receptor is a nuclear-associated protein which appears to mediate the actions of 3,5,3'-triiodo-L-thyronine (L-T3) and 3,5,3',5'-tetraiodo-L-thyronine (L-T4) in mammalian cells. In a previous study we reported that N-2-diazo-3,3,3-trifluoropropionyl-3,5,3'-triiodo-L-thyronine (L-T3-PAL) serves as an effective photoaffinity label probe of the receptor in GH1 cells, a growth hormone producting rat pituitary cell line. Irradiation of cells at 254 nm covalently cross-links L-[125I]T3-PAL to two molecular weight (Mr) nuclear receptor forms, an abundant 47,000 Mr component and a less abundant 57,000 Mr species (Pascual, A., Casanova, J., and Samuels, H. H. (1982) J. Biol. Chem. 257, 9640-9647). In this study we have explored a number of possible interrelationships of the different Mr receptor forms. Denaturing gel electrophoresis and autoradiography indicates that the 57,000 Mr form is a doublet species which differ in Mr by 1,000 to 2,000. The various receptor forms are not an artifact of the L-[125I]T3-PAL probe, and identical forms can be labeled at 310 nm using underivatized L-[125I]T4 with a 15-fold lower coupling efficiency. The 57,000 and 47,000 Mr receptor forms are not generated by indiscriminate proteolysis, UV peptide cleavage, or zero length protein-protein cross-linking by irradiation at 254 nm. Micrococcal nuclease excises both the 57,000 and 47,000 Mr forms, and receptor is not identified in the residual nuclear matrix fraction. Receptor is also not detected in the cytoplasmic fraction. By coupling dense amino acid labeling and photoaffinity labeling of receptor we determined a half-life of 2.4 h for the 57,000 Mr species and 5.6 h for the 47,000 Mr form while both species have similar relative synthetic rates. n-Butyrate has been previously shown to decrease receptor levels in GH1 cells. We demonstrate that n-butyrate decreases receptor levels primarily by shortening the half-life of the 47,000 Mr form.

Affinity Labels↗
PubMed1984

Cerebral hypothyroidism in rats with adult-onset iodine deficiency.

Rats fed chronically a low iodine diet may have low serum T4 and high circulating TSH, despite normal serum T3. As the brain depends to a great extent on intracellular generation of T3 from T4 for its total and nuclear T3, we have carried out two experiments to determine whether the brain of iodine-deficient rats may become hypothyroid, despite normal serum T3 levels. In both experiments we confirmed previous data, showing that the pituitary and liver of iodine-deficient rats with very low plasma T4 levels are hypothyroid as compared to those of animals receiving the same diet supplemented with KI, though not as markedly as animals which had undetectable circulating levels of both T4 and T3 as a consequence of chronic ingestion of KC1O-4, or of surgical thyroidectomy. We have further found that the nuclear T3 content was decreased in the brain of iodine-deficient rats, as compared with the animals on the iodine-supplemented diet. The nuclear to plasma ratios of labeled T3 showed that the uptake of this hormone into liver and brain nuclei is not decreased in the iodine-deficient rats as compared with those on the iodine-supplemented diet. This finding indicates that the decreased liver and brain nuclear T3 contents of iodine-deficient rats are likely to be a consequence of the marked reduction of their T4 pool, leading to decreased amounts of intracellularly generated T3. The number of spines on shafts of pyramidal neurons from the visual cortex of iodine-deficient rats was lower than that of rats fed the same diet supplemented with KI. Their distributions along the shaft were also not the same. Such changes might well be an index of cerebral hypothyroidism, as they are similar to those found after thyroidectomy of adult rats. It is concluded from the present findings that normal circulating T3 levels may not be sufficient to maintain brain euthyroidism in rats fed a diet iodine deficient enough to result in very low circulating T4 levels.

Animals↗
PubMed1984

Decreased T4 to T3 conversion in tissues of streptozotocin-diabetic rats.

The effect of experimental diabetes on T4 to T3 conversion, T3-deiodination, and the pituitary response to a dose of T4 and T3 was studied. Pituitary GH and plasma TSH were determined as a measure of the biological response to thyroid hormones. Thyroidectomized rats, 5 days after injection with saline or streptozotocin (thyroidectomized-control (Th.C) and thyroidectomized-diabetic (Th.D) rats, respectively) received an ip dose of T4 + [125I]T4 or T3 + [125I]T3. Rats from each group were sacrificed at varying intervals after thyroid hormone injection. Th.D rats had hyperglycaemia, glycosuria, and a body weight of about 80% of that of Th.C rats. The concentrations of [125I]T4 and [125I]T3 were measured in several tissues after ethanol extraction, separation by thin-layer chromatography, and identification with markers. Plasma TSH and pituitary GH were measured by specific RIAs. Diabetes decreased the conversion of T4 to T3 in several tissues, including the pituitary, but did not affect the deiodination of T3. The decrease in pituitary T3 content after a dose of T4 was accompanied by a diminution of the biological effect of the T4 dose on pituitary GH. Since diabetes also interferes with this biological response to a T3 dose, it seems likely that the reduced biological effect of thyroid hormones on pituitary GH may be related to an alteration in the somatotrophin T3 receptors, or in post-receptor events. Moreover, the data indicate that although T3 generation in the pituitary was reduced, the same dose of T4 had a greater inhibitory effect on TSH secretion in Th.D rats than in Th.C rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1980

Evidence against a major role of L-thyroxine at the pituitary level: studies in rats treated with iopanoic acid (telepaque).

To determine whether T4 has an intrinsic effect at the pituitary level, it would be important to block conversion of T4 to T3 completely. We have attempted to achieve this with iopanoic acid (IOP), a radiographic contrast agent. We have then measured in the same animals the effects of such treatment on the conversion of T4 to T3 or on the deiodination of T3 and on the pituitary response to a dose of T4 or T3. Plasma TSH levels and pituitary GH content were measured as biological end points. Thyroidectomized rats were injected with a single dose of T4 (1.7 micrograms/100 g BW) labeled with [125I]T4 (Exp A) or with a single dose of T3 (0.33 microgram/100 g BW) labeled with [125I]T3 (Exp B) and treated with IOP or solvent. Animals of Exp A were killed 24 h after iodothyronine injection and those of Exp B were killed 4, 12, and 24 h after injection of the iodothyronine. The concentrations of [125I]T4 and [125I]T3 were measured in several tissues, including the anterior pituitary, after extraction and paper chromatography and quantified with the aid of 131I-labeled markers added in vitro. Plasma and pituitary T3 and T4 plasma TSH, and pituitary GH were measured by specific RIAs. Results show that treatment with IOP markedly inhibits the conversion of T4 to T3 and the deiodination of T3. In IOP-treated thyroidectomized rats, the injection of T4 results in little, if any, effect at the pituitary level, despite an almost 3-fold increase in the percentage of injected T4 found in the gland. Treatment with IOP does not inhibit the effects of a T3 dose; if anything, they appear to be enhanced. It is concluded that, as assessed from biological responses involving the anterior pituitary, a dose of T4 has little if any effect other than that which can be attributed to the T3 generated from it.

Animals↗
PubMed1979

Tyrosine hydroxylase and the conversion of L-thyroxine into 3',3,5-triiodo-L-thyronone in the rat.

We have studied the effects of alpha-methyl-p-tyrosine (alpha-MPT), an inhibitor of tyrosine hydroxylase, on the in vivo conversion of L-T4 (T4) to 3',3,5-triiodo-L-thyronine (T3), and on the biological effectiveness of T4. Thyroidectomized rats were used and were injected daily with T4 maintenance doses. Three different types of experiments were carred out. The first involved isotopic equilibration with 125I-labeled T4 and measurement of urinary 125I excretion. The second series involved the injection of a single dose of [125I]T4, with the amounts of [125I]T3 in different tissues being studied 7 or 20 h later. The third series involved daily treatment for 13 days with T4 and alpha-MPT, at the end of ehich the liver alpha-glycerophosphate dehydrogenase activity was measured as a parameter of the biological effects of the hormone. Though the experimental approaches used clearly disclosed the well known effects of 6-propyl-2-thiouracil, no clear-cut effects of alpha-MPT were observed. It is concluded that alpha-MPT neither inhibits the conversion of T4 to T3 in vivo in rats nor affects the biological potency of a given dose of T4, at least to an extent compararble to that observed when 6-propyl-2-thiouracil is used. Thus, present results do not support the hypothesis that tyrosine hydroxylase is involved in the extrathyroidal deiodination of T4 to T3.

Animals↗