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.     

Analysis of the NH2-Terminal 83rd Amino Acid of Escherichia coli GyrA in Quinolone-Resistance

Artificial mutations of Gyrase A protein (GyrA) in Escherichia coli by site-directed mutagenesis were generated to analyze quinolone-resistant mechanisms. By genetic analysis of gyrA genes in a gyrA temperature sensitive (Ts) background, exchange of Ser at the NH₂-terminal 83rd position of GyrA to Trp, Leu, Phe, Tyr, Ala, Val, and Ile caused bacterial resistance to the quinolones, while exchange to Gly, Asn, Lys, Arg and Asp did not confer resistance. These results indicate that it is the most important for the 83rd amino acid residue to be hydrophobic in expressing the phenotype of resistance to the quinolones. These findings also suggest that the hydroxyl group of Ser would not play a major role in the quinolone-gyrase interaction and Ser83 would not interact directly with other amino acid residues.     

One-dimensional food chain equations and their generalization

Two equations describing one-dimensional food chains are known to possess soliton solutions. It is demonstrated that both equations are embraced within another equation, which arises in the theory of chains of enzymic reactions. We find an elliptic function solution to this equation. We obtain a one-soliton solution from it and re-derive the elliptic function solutions of the two ecological equations.     

The role of the single tryptophan residue in the structure and function of ribonuclease T1

The previously reported method for the preparation of Kyn 59-RNase T1 and NFK 59-RNase T1 has been improved, and these two proteins have been obtained in high purity. Kyn 59-RNase T1, fully active for the hydrolysis of GpA and GpC, emitted a 35-fold-enhanced fluorescence of kynurenine relative to acetylnurenine amide with an emission maximum at 455 nm upon excitation at 380 nm. The polarity of the environment of Kyn 59 estimated from the emission maximum corresponded to a dielectric constant of 6. Upon excitation at 325 nm, NFK 59-RNase T1, less active than Kyn 59-RNase T1, exhibited a quenched N'-formylkynurenine fluorescence with an emission maximum at 423 nm, from which the value of 12 was obtained as the dielectric constant of the surroundings of residue 59. In both modified proteins, distinct tyrosine fluorescence appeared on excitation at 280 nm. The detection of an energy transfer from tyrosine to residue 59 suggests that the tertiary structure is very similar in Kyn 59-RNase T1 and native RNase T1. With guanidine hydrochloride, Kyn 59-RNase T1 was less stable than the native protein. Carboxymethylation at Glu 58 was shown to stabilize the active site of the modified enzyme. Based on the information collected for Kyn 59-RNase T1, the local environment and possible roles of the sole tryptophan residue in RNase T1 are discussed.     

Source of prolactin in human follicular fluid.

To analyze whether prolactin (PRL) in human follicular fluid (FF) is synthesized locally or derived from the circulation, PRL concentrations of plasma and FF were determined in the patients after ovarian stimulations. The amounts of PRL messenger ribonucleic acid (mRNA) in the follicular tissues during different menstrual phases were also determined. The FF PRL concentration was correlated positively with plasma PRL and highest estradiol levels during the stimulatory cycle. No PRL mRNA sequence was detected in the RNAs extracted from follicles at any stage in the menstrual cycle, although beta-actin mRNA was detected in all samples. In a comparison with pituitary RNA, the PRL mRNA concentration in ovarian follicular tissues seemed to be 10,000 times less than that in the pituitary. These results suggest that FF PRL may not be synthesized locally, but derived from the pituitary via the circulation through passive diffusion, and thus regulated by estrogen.     

The structure of pyridinoline,a collagen crosslink

Pyridinoline is an amino acid isolated from collagen and probably serves as a crosslink in collagen fiber. This compound was isolated on a large scale from bovine bone and investigated by ¹H-nmr and ¹³C-nmr spectroscopy, mass spectroscopy and chemical degradation. The structure is proposed on the basis of these data.