Involvement of L-Type-Like Amino Acid Transporters in S-Nitrosocysteine-Stimulated Noradrenaline Release in the Rat Hippocampus

Abstract: Nitrogen oxides, such as nitric oxide, have been shown to regulate neuronal functions, including neurotransmitter release. We investigated the effect of S-nitroso-l -cysteine (SNC) on noradrenaline (NA) release in the rat hippocampus in vivo and in vitro. SNC stimulated [³H]NA release from prelabeled hippocampal slices in a dose-dependent manner. SNC stimulated endogenous NA release within 30 min to almost five times the basal level in vivo (microdialysis in freely moving rats). In a Na⁺-containing Tyrode's buffer, SNC-stimulated [³H]NA release was inhibited 30% by the coaddition of l -leucine. In the Na⁺-free, choline-containing buffer, SNC-stimulated [³H]NA release, which was similar to that in the Na⁺-containing buffer, was inhibited markedly by l -leucine, l -alanine, l -methionine, l -phenylalanine, and l -tyrosine. The effects of the other amino acids examined were smaller or very limited. The effect of l -leucine was stronger than that of d -leucine. A specific inhibitor of the L-type amino acid transporter, 2-aminobicyclo[2.2.1]-heptane-2-carboxylate (BCH), inhibited the effects of SNC on [³H]NA release in the Na⁺-free buffer. Uptake of l -[³H]leucine into the slices in the Na⁺-free buffer was inhibited by SNC, BCH, and l -phenylalanine, but not by l -lysine. The effect of SNC on cyclic GMP accumulation was not inhibited by l -leucine, although SNC stimulated cyclic GMP accumulation at concentrations up to 25 µM, much less than the concentration that stimulates NA release. These findings suggest that SNC is incorporated into rat hippocampus via the L-type-like amino acid transporter, at least in Na⁺-free conditions, and that SNC stimulates NA release in vivo and in vitro in a cyclic GMP-independent manner.     

Peroxidation of lipids and growth inhibition induced by UV-B irradiation

Cotyledons excised from dark-grown seedlings of cucumber (Cucumis sativus L.) were cultured in vitro under UV radiation at different wavelengths, obtained by passage of light through cut-off filters with different transmittance properties. Growth and the synthesis of chlorophyll (Chl) in cotyledons were inhibited and malondialdehyde was accumulated upon irradiation at wavelengths below 320 nm. Exogenous application of scavengers of free radicals reversed the growth inhibition induced by UV-B. Measurement of the fluorescence of Chl a suggested that electron transfer in photosystems was affected by UV-B irradiation. On the basis of these results, the involvement is postulated of active species of oxygen in damages to thylakoid membranes and the growth inhibition that are induced by UV-B irradiation.Abbreviations Chl chlorophyll - F_m maximal fluorescence (dark) - F_m maximal fluorescence (light) - F_v variable fluorescence (dark) - F_v variable fluorescence (light) - MDA malondialdehyde - O₂ Superoxide radical - PS photosystem - q_N non-photochemical quenching of fluorescence - q_P photochemical quenching of fluorescence - UV-B_BE biologically effective UV-B radiation - WL(T = 0.5) wavelength at which 50% transmittance occurs     

Biochemical studies on strain differences of mice in the susceptibility to nitrogen dioxide

Strain differences of mice in their susceptibility to nitrogen dioxide (NO₂) were examined by measuring the activities of antioxidative protective enzymes, and the amounts of antioxidants and lipid peroxides in lungs. Four strains of mice: ICR, BALB/c, ddy and C57BL/6 were used in this study and their LC₅₀ values after exposure to NO₂ for 16 hr were: 38, 49, 51 and 64 ppm, respectively (1).Genetic strain differences were observed in the enzyme activities, the antioxidant contents and lipid peroxide contents among these four different strains. The activities of glutathione peroxidase (GP_x), glutathione S-transferase, and superoxide dismutase (SOD), and the contents of non-protein sulfhydryls (NPSH), α-tocopherol (α-Toc) and total lipids in lungs of the four strains were related to their LC₅₀, while TBA reactants in lungs of the four strains were inversely related to their LC₅₀.After exposure to 20 ppm NO₂ for 16 hr, the activities of the protective enzymes and the contents of NPSH decreased, while the level of α-Toc increased markedly. The activities of GP_x, 6-phosphogluconate dehydrogenase, SOD and disulfide reductase, and the contents of NPSH, α-Toc and total lipids were also related to their LC₅₀. On the other hand, TBA reactants increased higher than those of the control groups and were inversely related to their LC₅₀.These results suggest that the protective enzymes and the antioxidants are important factors as defence mechanism in lungs to NO₂ and that the intensity of the protective systems in pigmented strains is generally greater than that in albino strains.     

Degradation of methylmercury by selenium

When methylmercury was incubated in the presence of selenite and reduced glutathione (GSH), the mercury which was extracted into benzene under acidic condition decreased gradually with the elapse of time. This decrease was due to the cleavage of mercury-carbon bond of methylmercury. The reaction did not proceed when selenite or GSH was singly added to the reaction mixture. L-Cysteine, 2-mercaptoethanol and sodium sulfide in place of GSH also were effective for decomposition of methylmercury in combination with selenite, but oxidized glutathione (GSSG) and L-cystine were not. This suggests that reduction of selenite is needed for the degradation of methylmercury. Thus, the effect of reduced metabolites of selenite produced by GSH was investigated. Glutathione selenotrisulfide (GSSeSG) requierd GSH for the degradation of methylmercury, whereas H₂Se possessed a strong activity even in the absence of GSH. This may indicate that H₂Se is involved directly in the conversion of methylmercury to inorganic mercury. This phenomenon found in $\text{in vitro}$ experiments is discussed in relation to the biotransformation of methylmercury.     

Developmental changes in soluble proteins during the early development of Dictyostelium discoideum

Changes in the pattern of soluble proteins that accumulatedat the growth phase, interphase and late-aggregation phase ofthe cellular slime mold Dictyostelium discoideum were studiedby two-dimensional polyacrylamide gel electrophoresis. Amongthe 300 proteins detected during the early development, themost soluble do not change during the growth and aggregationphases, but about 90 proteins show changes in their relativeintensities on staining. During the transition from growth tothe interphase, the predominant changes were the disappearanceof 16 spots, the decrease in 30 spots, the appearance of 13new spots, and the increase in 14 spots. In contrast, from theinterphase to the late-aggregation phase, there were remarkableprogressive increases in 13 spots, an overall increase in 6spots, a decrease in 16 spots, the appearance of 8 new spotsand the disappearance of 4 spots. (Received July 13, 1979; )     

Photosynthetic regeneration of ATP using a strain of thermophilic blue-green algae

Photosynthetic ATP accumulation was shown in the presence of exogenous ADP plus orthophosphate on illumination to the intact cells of a strain of thermophilic blue-green algae isolated from Matsue hot springs, Mastigocladus sp. Kinetic studies of various effectors on the ATP accumulation proved that the ATP synthesis depends mainly on the cyclic photophosphorylation system around photosystem I (PS-I) in the algal cells. The temperature and pH optima for the accumulation were found at 45 degrees C and pH 7.5. Maximum yield was obtained with light intensity higher than 15 mW/cm(2). Borate ion exerted pronounced enhancement on the ATP synthesis. With a continuous reactor at a flow rate of 1 ml/hr at 45 degrees C and pH 7.5, efficient photoconversion of ADP (2mM, at substrate reservoir) to ATP (1mM, at product outlet) has been maintained for a period of 2.5 days, though the efficiency has decreased in a further 2-day period to the level of 0.5mM ATP/9.5 h of residence time.     

Ribosomal resistance of an istamycin producer, Streptomycestenjimariensis, to aminoglycoside antibiotics

$\text{Streptomyces}\text{tenjimariensis}$ SS-939 was resistant to its own aminoglycoside antibiotics, istamycins, as well as kanamycin A, neamine, ribostamycin and butirosin A, but was susceptible to neomycin B, lividomycin A and streptomycin. This resistance to these antibiotics was found to be due to ribosomes of the strain.     

Earthworm (Pheretima communissima and Pheretima hilgendorfi) hemoglobins: giant assemblies and subunits

The molecular architecture of hemoglobins and their subunits of the earthworms Pheretima communissima and Pheretima hilgendorfi was investigated. In both species, their s0.20,w of 60.8 S and D020,w of 1.80 . 10(-7) cm2 . s-1 corresponded to a molecular weight of 3.07 . 10(6). From electron microscopic observations, the overall structure of the hemoglobins was shown to be two superimposed hexagonal discs, each composed of six-membered constituents. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of both hemoglobins revealed the presence of five species of subunits with molecular weights of 13,000-14,000 (subunit 1), 27,000-28,000 (subunit 2), 30,000-31,000 (subunit 3), 33,000-34,000 (subunit 4) and approx. 52,000 (subunit 5), respectively, and the molar ratio of these subunits 1:2, 3, 4:5 was 2:3:3. If we consider this set of the subunits 1 to 5 as one unit, the molecular weight of this unit should be 2.7-2.8 . 10(5). This one unit, therefore, should be considered to represent one-twelfth the whole molecule with molecular weight of 3.07 . 10(6).     

Elongation and shortening of time required for entry into S phase after release from G 1 and G 0 arrests in temperature-sensitive mutants of rat 3Y1 Cells

Temperature-sensitive (ts) mutants of rat 3Y1 fibroblasts representing four separate complementation groups (3Y1tsD123, 3Y1tsF121, 3Y1tsG125, and 3Y1tsH203) are arrested mainly in the G1 phase when cells of randomly proliferating population at 33.8 degrees C are shifted to 39.8 degrees C (temperature arrest). We examined the time lag of the cellular entry into the S phase after release at 33.8 degrees C, both from the temperature arrest and from the arrest at 33.8 degrees C at a confluent cell density (density arrest). In the temperature-arrested cells, as the duration of temperature arrest increased, the time lag of entry into S phase after shift down to 33.8 degrees C was prolonged, in all four mutants. These observations suggest that the four different functional lesions, each causing arrest in the G1 phase, are also responsible for prolongation of the time lag of entry into the S phase in cells arrested in the G1 phase. The prolongation of the time lag in the temperature-arrested cultures was accelerated at a higher cell density, in medium supplemented with a lower concentration of serum, and at a higher restrictive temperature. In the density-arrested cells, as the duration of pre-exposure to 39.8 degrees C was increased, the time lag of entry into S phase at 33.8 degrees C after release from the arrest was drastically prolonged, in all four mutants. In 3Y1tsF121, 3Y1tsG125, and 3Y1tsH203, when the density-arrested cells were prestimulated by serum at 39.8 degrees C for various periods of time, the time lag of entry into S phase after release from the density arrest at 33.8 degrees C was initially shortened, and then, prolonged progressively as the period of prestimulation increased. These findings, taken together with other data, show that all four ts defects affect cells in states ranging from the deeper resting to mid- or late-G1 phase. It is suggested that events represented by these four mutants are required for entry into the S phase and normally operate in parallel but not in sequence in cells in states ranging from the deeper resting to the mid- or late-G1 phases, though they may affect each other.