Biochemical changes of rat brain membranes with aging

Modification of membrane composition and enzymatic activities both in total brain homogenate and purified synaptic plasma membrane of 3 and 24 month old rats has been investigated. Protein, cholesterol and phospholipid content and (Na+, K+)ATPase and 2',3' cyclic nucleotide phosphohydrolase activities were determined. The major changes occurred in the whole homogenate where a general increase in total protein and cholesterol content with age and a significant increase of the cholesterol/phospholipids molar ratio has been detected. In S.P.M. aging process induced a decrease of protein, cholesterol and phospholipids content associated with an increased membrane viscosity and a decrease of delta E. These data are consistent with a change in the structural organization and in the distribution pattern of different cell population in the aging brain. A possible artifactual effect of freezing on the reported parameter is also discussed.     

Fusicoccin Binding to its Plasma Membrane Receptor and the Activation of the Plasma Membrane H+-ATPase. II. Stimulation of the H+-ATPase in a Plasma Membrane Fraction Purified by Phase-partitioning

The effect of fusicoccin (FC) on the activity of the PM H⁺-ATPase was investigated in a plasma membrane (PM) fraction from radish seedlings purified by the phase-partitioning procedure. FC stimulated the PM H⁺-ATPase activity by up to 100 %; the effect was essentially on V_max with only a slight decrease of the apparent K_M of the enzyme for ATP. FC-induced stimulation of the PM H⁺-ATPase was evident within the first minute and maximal within five minutes of membrane treatment with the toxin indicating that transmission of the signal from the activated receptor to the PM H⁺-ATPase is very rapid. Both FC-induced stimulation of the PM H⁺-ATPase and FC binding to its receptor decreased dramatically upon incubation of the membranes in ATPase assay medium at 33 °C in the absence of FC, due to the lability of the free FC receptor. FC-induced stimulation of the PM H⁺-ATPase was strongly pH dependent: absolute increase of activity was maximal at pH 7, while percent stimulation increased with the increase of pH up to pH 7.5; FC binding was scarcely influenced by pH in the pH range investigated. Taken as a whole, these results indicate that FC binding is a condition necessary, but not sufficient, for FC-induced stimulation of the PM H⁺-ATPase.     

Extracellular Concentrations of Dopamine and Metabolites in the Rat Caudate After Oral Administration of a Novel Catechol-O-Methyltransferase Inhibitor Ro 40–7592

The effect of the systemic administration of a novel, orally active, catechol-O-methyltransferase (COMT) inhibitor, Ro 40-7592, on the in vivo extracellular concentrations of dopamine (DA) and its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), was studied by transcerebral microdialysis in the dorsal caudate of freely moving rats. Ro 40-7592 (at doses of 3.0, 7.5, and 30 mg/kg p.o.) elicited a marked and long-lasting reduction of HVA, and at doses of 7.5 and 30 mg/kg, an increase of DOPAC output, but it failed to increase DA output. The administration of L-beta-3,4-dihydroxyphenylalanine (L-DOPA, 20 and 50 mg/kg p.o.) with a DOPA decarboxylase inhibitor (benserazide) increased both HVA and DOPAC output, but failed to modify significantly extracellular DA concentrations in dialysates; in contrast, combined administration of L-DOPA+benserazide with Ro 40-7592 (30 mg/kg p.o.) resulted in a significant increase in DA output. Ro 40-7592 prevented the L-DOPA-induced increase in HVA output and markedly potentiated the increase in DOPAC output. To investigate to what extent the increase in extracellular DA concentrations was related to an exocitotic release, tetrodotoxin (TTX) sensitivity was tested. Addition of TTX to Ringer, although abolishing DA output in the absence of L-DOPA, partially reduced it in the presence of L-DOPA+Ro 40-7592 and even more so after L-DOPA without the COMT inhibitor. The results of the present study suggest that metabolism through COMT regulates extracellular concentrations of DA formed from exogenously administered L-DOPA but not of endogenous DA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cytochemical Ca2+ Distribution in Activated Discoglossus pictus Eggs: A Gradient in the Predetermined Site of Fertilization

The K-pyroantimonate/OsO₄ (PA) cytochemical method coupled with EGTA and X-ray microanalytical controls has been used to localize Ca²⁺ at egg activation in Discoglossus pictus eggs. The results show that: 1) the PA method is able to selectively localize Ca²⁺ pools mobilized by activating stimuli; 2) the smooth endoplasmic reticulum (SER) elements located in the animal dimple region, i.e. in the predetermined site of fertilization, are the first egg components labeled by precipitates; 3) a decreasing gradient of precipitates is present from the center beyond the boundaries of the dimple region; 4) precipitates are lacking in the remainder of the egg even at late times after activation.
The possibilities are discussed that a) SER is the major Ca²⁺-releasing store at activation in Discoglossus , and b) the observed gradient of pyroantimonate-detected Ca²⁺ reflects an ionic Ca²⁺ gradient.     

Binding of matrix attachment regions to lamin B1.

Eukaryotic chromatin is organized into topologically constrained loops that are attached to the nuclear matrix. The regions of DNA that interact with the matrix are called matrix attachment regions (MARs). We studied the spatial distribution of MAR-binding sites in the nuclear matrix from rat liver cells, following a combined biochemical and ultrastructural approach. We found that MAR-binding sites are distributed equally over the internal fibrogranular network and the peripheral nuclear lamina. Internal and peripheral binding sites have similar binding characteristics: both sets of binding sites show specific and saturable binding of MARs from different organisms. By means of a DNA-binding protein blot assay and in vitro binding studies, we identified lamin B1 as a MAR-binding protein, which provides evidence for a specific interaction of DNA with the nuclear lamina.     

Primary multilobated T-cell lymphoma of the breast diagnosed by fine needle aspiration cytology and immunocytochemistry

The fine needle aspiration (FNA) cytologic, immunocytochemical and ultrastructural findings of a primary multilobated T-cell lymphoma arising in the breast of a 61-year-old woman are described. Large pleomorphic multilobated malignant cells were primarily identified as lymphomatous in origin and phenotypically as T-cells by a selected panel of monoclonal antibodies applied to the original smears obtained by FNA biopsy. This appears to be the second report of a multilobated lymphoma arising in the breast and the first with a T-cell phenotype in this anatomic site.     

Chromosome constitution of in vitro segregated haploid and diploid cells of the mouse

The composition in segregated haploid sets of paternal and maternal chromosomes has been studied in order to verify whether their composition is uniparental of mixed, fixed or variable. Primary cultures where prepared using kidneys from hybrids of strains of Mus musculus in which the parental chromosomes are distinguishable; the maternal set consists of 20 teleocentric chromosomes, the paternal set of 9 metacentric chromosomes, derived by Robertsonian fusion and 2 telocentrics. Applying Seabright's banding technique, an analysis of segregated haploid and diploid cells, which have originated spontaneously through polyploidisation-segregation processes was carried out. It was concluded that the haploid sets have a variable composition of paternal and maternal chromosomes.