Influence of thyroparathyroidectomy and thyroxine replacement on CU and ZN cellular distribution and on the metallothionein level and induction in rats

Thyroid hormones are involved in copper and zinc distribution in rat tissues. We examined the influence of thyroparathyroidectomy (TPTY) and of a replacement therapy by T4 on Cu and Zn organ distribution. MT levels were also measured both in basal conditions and after induction by cadmium. The results confirm that a lack of T4 modified Cu and Zn in serum and tissues. In serum, TPTY increased Cu (+15%) and ceruloplasmin (+18%), and decreased Zn (−18%). In tissues, Cu was altered in liver (+13%), kidney (−24%), heart (−16%) duodenum (−18%), and Zn in liver (+25%) and kidney (−10%). The soluble fractions (100,000 g supernatant) were mainly affected in liver and kidney, and the subcellular fractions in heart and duodenum. MT levels were modified in basal conditions only in liver (+57%) and kidney (−36%). T4 administration partially prevented the effect of TPTY on both elements and MT concentrations. Therefore, no evidence is provided for a direct role of T4 in the metabolism of MT in a way comparable to the effects of glucocorticoids. However, MT could mediate the consequences of TPTY on metal distribution in certain organs, such as liver and kidney.     

Characterization of a major brain tubulin variant which cannot be tyrosinated

Brain tubulin preparations contain an abundant type of tubulin which does not undergo the normal cycle of tyrosination-detyrosination, and whose nature is still unknown. We have used peptide sequence analysis and mass spectrometry combined with immunological procedures to show that this non-tyrosinatable tubulin has a specific primary structure. It differs from the tyrosinated isotype in that it lacks a carboxy-terminal glutamyl-tyrosine group on its alpha-subunit. Thus, non-tyrosinatable tubulin originates from a well-defined posttranslational modification of the tubulin primary structure which is located at the expected site of activity of tubulin tyrosine ligase. This probably accounts for the reason why it cannot be tyrosinated. The significance of this abundant brain isotubulin and the metabolic pathway involved in its formation remain to be elucidated. This should shed light on the relation between the structural diversity of the carboxy terminus of alpha-tubulin and the regulation of functional properties of microtubules.     

A METHOD FOR ESTIMATING MASS OF LARGE PINNIPEDS

Fifty-two male elephant seals were weighed and photographed at Año Nuevo State Reserve, California, to establish a predictive relationship between photographically measured morphological variables (length, side area, and girth area) and body mass. Regression of mass on these variables revealed that side area, roughly equivalent to a longitudinal cross-section, was the most useful single variable for predicting mass, and that adding the other two variables to side area slightly improved the accuracy of the photogrammetric technique. Curvilinear regressions based on a power model provided the best predictive relationships. This technique may prove useful for estimating body mass of other pinnipeds.     

High-performance liquid chromatographic determination of pipotiazine in human plasma and urine

A high-performance liquid chromatographic method has been developed for the determination of pipotiazine in human plasma and urine. After selective extraction, pipotiazine and the internal standard (7-methoxypipotiazine) are chromatographed on a column packed with Spherosil XOA 600 (5 μm) using a 7:3 (v/v) mixture of diisopropyl ether—isooctane (1:1, v/v) + 0.2% triethylamine and diisopropyl ether—methanol (1:1, v/v) + 0.2% triethylamine + 2.6% water. The eluted compounds are measured by fluorescence detection. The sensitivity of the method was established at 0.25 ng/ml pipotiazine in plasma and 2 ng/ml pipotiazine in urine (C.V. < 5%). The method has been successfully applied to a pharmacokinetic study following a single oral administration of 10 mg of pipotiazine.     

Decrease in lymphocyte [3H]spiroperidol binding sites in parkinsonism

[³H]spiroperidol binding has been measured in lymphocytes from patients with Parkinson's disease and age matched healthy volunteers. A dramatic decrease (73%) in the number of binding sites (^Bmax) without any variation of the affinity (^KD) has been observed in Parkinsonian patients. This decrease in ^Bmax is linearly correlated with the degree of disability of the Parkinsonian patients (r = 0.891, p <0.001). This decrease appeared to be relatively selective since no variation was observed with patients suffering of other neurological disorders (vascular hemiplegia, Alzeihmer's disease, olivopontocerebellar degeneration, Huntington's chorea).     

Freeze-fracture study of water-soluble, standard proteins and of detergent-solubilized forms of sarcoplasmic reticulum Ca2+-ATPase

Conventional freeze-fracturing electron microscopy was used to study water-soluble proteins and different forms of Ca2+-ATPase-detergent complexes. Freeze-fracture images of solutions containing proteins larger than myoglobin showed the presence of distinct, randomly dispersed particles on smooth fracture surfaces. The distribution of sizes of these particles was closely to Gaussian, with a mean size which was correlated to the Stokes diameter. Monomeric Ca2+-ATPase from sarcoplasmic reticulum, solubilized by deoxycholate or a non-ionic detergent, showed a bimodal distribution of particle sizes. Even more complex distributions were found for dimeric and trimeric preparations of Ca2+-ATPase. The results can be interpreted on the assumption that the Ca2+-ATPase molecule is elongated, with an overall length of about 110 A and a width in its largest part of about 75 A. It is concluded on the basis of the presented results that freeze-fracture electron microscopy can be successfully used for morphological studies of protein molecules in solution.     

Substrate specificity and adenosine triphosphatase activity of the ATP-dependent deoxyribonuclease of Bacillus subtilis

Studies on the specificity of the ATP-dependent DNase of Bacillus subtilis 168, carried out with pure enzyme at the optimal conditions for its action, have shown that the substrate is double-stranded linear DNA. Linear single-stranded DNA (separated strands of B. subtilis DNA and linear phage fd DNA) is not attacked, neither are there any circular forms (supercoiled or nicked simian virus 40 and circular single-stranded fd DNAs). The double-stranded DNA can be completely hydrolysed, the limit products being, almost exclusively, mononucleotides. The presence of terminal phosphate residues in the substrate (either at the 3' or the 5' end) is not necessary for enzyme action. This DNase appears therefore to be an exonuclease processively liberating mononucleotides from both strands of the native linear DNA. ATP (indispensable for the DNase reaction) is also hydrolysed by the enzyme, to ADP and inorganic orthophosphate (Pi) in the presence of DNA. The apparent Km for ATP, in the ATPase reaction, is 0.15 mM. At high ATP concentrations, which inhibit the DNase activity, there is activation of the ATPase reaction. Three molecules of ATP are consumed for each DNA phosphodiester bond split, at optimal conditions for DNase activity.     

Active K+ transport in Mycoplasma mycoides var. Capri. Net and unidirectional K+ movements.

Analysis of the cation composition of growing Mycoplasma mycoides var. Capri indicates that these organisms have a high intracellular K+ concentration (Ki: 200--300 mM) which greatly exceeds that of the growth medium, and a low Na+ concentration (Na+i: 20 mM). Unlike Na+i,K+i varies with cell aging. The K+ transport properties studied in washed organisms resuspended in buffered saline solution show that cells maintain a steady and large K+ concentration gradient across their membrane at the expense of metabolic energy mainly derived from glycolysis. In starved cells, K+i decreases and is partially compensated by a gain in Na+. This substitution completely reverses when metabolic substrate is added (K+ reaccumulation process). Kinetic analysis of K+ movement in cells with steady K+ level shows that most of K+ influx is mediated by an autologous K+-K+ exchange mechanism. On the other hand, during K+ reaccumulation by K+-depleted cells, a different mechanism (a K+ uptake mechanism) with higher transport capacity and affinity drives the net K+ influx. Both mechanisms are energy-dependent. Ouabain and anoxia have no effect on K+ transport mechanisms; in contrast, both processes are completely blocked by dicyclohexylcarbodiimide, an inhibitor of the Mg2+ -dependent ATPase activity.