Yousuke Tsuneyoshi

Intracerebroventricular injection of L-arginine induces sedative and hypnotic effects under an acute stress in neonatal chicks.

Summary.l-Arginine participates in many important and diverse biochemical reactions associated with the normal physiology of the organism. In the present study, we investigated the effect of central administration of l-arginine on the stress response and its mechanism in neonatal chicks. Intracerebroventricular (i.c.v.) injection of l-arginine clearly attenuated the stress response in a dose-dependent manner, and induced sleep-like behavior during 10 min. To clarify the mechanism by which l-arginine induces sedative and hypnotic effects in chicks, we investigated the effects of nitric oxide (NO) synthase (NOS) inhibitors on l-arginine-induced sedative and hypnotic effects, and as well as the effects of a NO donor. l-Arginine-induced (1.9 µmol) sedative and hypnotic effects were attenuated by i.c.v. co-injection with a non-selective NOS inhibitor NG-nitro-l-arginine methyl ester HCl (400 nmol). In addition, the effects of l-arginine were slightly attenuated by the inactive isomer of the NOS inhibitor NG-nitro-d-arginine methyl ester HCl (400 nmol). The i.c.v. injection of 3-morpholinosylnomine hydrochloride, a spontaneous NO donor, had little effect on postures. The i.c.v. injection of l-arginine had no effect on NOx concentration at various brain sites. These results suggested that the contribution of NO generation via NOS may be low in the sedative and hypnotic actions of l-arginine. Therefore, l-arginine and/or its metabolites, excluding NO, may be necessary for these actions.

Central l-arginine reduced stress responses are mediated by l-ornithine in neonatal chicks

Summary.Recently, we observed that central administration of l-arginine attenuated stress responses in neonatal chicks, but the contribution of nitric oxide (NO) to this response was minimal. The sedative and hypnotic effects of l-arginine may be due to l-arginine itself and/or its metabolites, excluding NO. To clarify the mechanism, the effect of intracerebroventricular (i.c.v.) injection of l-arginine metabolites on behavior under social separation stress was investigated. The i.c.v. injection of agmatine, a guanidino metabolite of l-arginine, had no effect during a 10 min behavioral test. In contrast, the i.c.v. injection of l-ornithine clearly attenuated the stress response in a dose-dependent manner, and induced sleep-like behavior. The l-ornithine concentration in the telencephalon and diencephalon increased following the i.c.v. injection of l-arginine. In addition, several free amino acids including L-alanine, glycine, l-proline and l-glutamic acid concentrations increased in the telencephalon. In conclusion, it appears that l-ornithine, produced by arginase from l-arginine in the brain, plays an important role in the sedative and hypnotic effects of l-arginine observed during a stress response. In addition, several other amino acids having a sedative effect might partly participate in the sedative and hypnotic effects of l-arginine.

Intracerebroventricular injection of L-proline and D-proline induces sedative and hypnotic effects by different mechanisms under an acute stressful condition in chicks

The central effects of L-proline, D-proline and trans-4-hydroxy-L-proline were investigated by using the acute stressful model with neonatal chicks in Experiment 1. Sedative and hypnotic effects were induced by all compounds, while plasma corticosterone release under isolation stress was only attenuated by L-proline. To clarify the mechanism by which L-proline and D-proline induce sedative and hypnotic effects, the contribution of the strychnine-sensitive glycine receptor (glycine receptor) and N-methyl-D-aspartate glutamate receptor (NMDA receptor) were further investigated. In Experiments 2–3, the glycine receptor antagonist strychnine was co-injected intracerebroventricular (i.c.v.) with L-proline or D-proline. The suppression of isolation-induced stress behavior by D-proline was attenuated by strychnine. However, the suppression of stress behavior by L-proline was not attenuated. In Experiment 4, the NMDA receptor antagonist (+)-MK-801 was co-injected i.c.v. with L-proline. The suppression of stress behavior by L-proline was attenuated by (+)-MK-801. These results indicate that L-proline and D-proline differentially induce sedative and hypnotic effects through NMDA and glycine receptors, respectively.

N-Methyl-d-aspartate and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors involved in the induction of sedative effects under an acute stress in neonatal chicks

Glutamate, an excitatory amino acid, acts at several glutamate receptor subtypes. Recently, we reported that central administration of glutathione induced hypnosis under stressful conditions in neonatal chicks. Glutathione appears to bind to the N-methyl-d-aspartate (NMDA) receptor. To clarify the involvement of each glutamate receptor subtype during stressful conditions, intracerebroventricular (i.c.v.) injection of several glutamate receptor agonists was given to chicks under social separation stress. Glutamate dose-dependently induced a hypnotic effect. NMDA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) and kainate are characterized as ionotropic glutamate receptors (iGluRs). Although NMDA also induced sleep-like behavior or sedative effects, the potency of NMDA was less than that of glutamate. AMPA tended to decrease distress vocalizations induced by acute stress and brought about a sedative effect. Kainate and (S)-3, 5-dehydroxyphenylglycine, which is a metabotropic glutamate receptor agonist, had no influence on chick behavior. Thus, it is suggested that the iGluRs, NMDA and AMPA, are important in inducing hypnosis and sedation under acute stress in chicks.

L-Serine induces sedative and hypnotic effects acting at GABAA receptors in neonatal chicks

Intracerebroventricular injection of l-serine has been shown to have sedative and hypnotic effects on neonatal chicks exposed to acute stressful conditions. However, the mechanism by which l-serine induces these effects is unclear. The present study was conducted to clarify the mechanism by l-serine. The involvement of gamma-aminobutyric acid (GABA)(A) receptors on the effect of l-serine was investigated using the GABA(A) receptor antagonist picrotoxin. Co-administration of picrotoxin attenuated the sedative and hypnotic effect of l-serine. Further, we also investigated the involvement of glycine receptors since l-serine is suggested to act as the alpha-homomeric glycine receptor agonist. Glycine similarly induced sedative and hypnotic effects in chicks, but its effect was attenuated by the glycine receptor antagonist strychnine. Therefore, whether the effect of l-serine was mediated through the glycine receptor was investigated using l-serine and strychnine. The effect of l-serine was inhibited by picrotoxin, but not strychnine. It appears that l-serine induces sedative and hypnotic effects by enhancing inhibitory neurotransmission via GABA(A) receptors.

Relationships between the sedative and hypnotic effects of intracerebroventricular administration of L-serine and its metabolites, pyruvate and the derivative amino acids contents in the neonatal chicks under acute stressful conditions

Summary.Intracerebroventricular (i.c.v.) injection of L-serine was shown to have sedative and hypnotic effects on neonatal chicks under acute stressful conditions. To clarify the central mechanism of these effects of L-serine, two experiments were done. First, we focused on the glycogenic pathway in which L-serine is converted into pyruvate and finally glucose. I.c.v. administration of pyruvate (0.84 µmol) did not induce any behavioral and endocrinological changes, while L-serine and glucose triggered sedative and hypnotic effects. Secondly, the relationship between the sedation by L-serine and the metabolism into other amino acids which have sedative effects was investigated in the telencephalon and diencephalon. In both brain areas, a dose-dependent increase was seen in L-serine, although other amino acids were not changed. In the present study, it was concluded that the sedative action of L-serine was not due to the action of its metabolite pyruvate, or to the action of other amino acids.

Oral administration of l-serine reduces the locomotor activity of socially isolated rats

L-Serine is considered a functional amino acid in the central nervous system, since intracerebroventricular injection of L-serine induced sedative and hypnotic effects in neonatal chicks exposed to acute stressful conditions. Accordingly, L-serine is a candidate anti-stress factor, but the effect of daily intake of L-serine on behavior of animals exposed to chronic stress has not been investigated. In the present study, we exposed rats to social isolation stress for 4 weeks, and home cage test and open field test were concluded to evaluate the effect of L-serine on behavior. To investigate L-serine supplementation modifies the brain L-serine and its metabolite contents, free amino acid contents were measured by a high performance liquid chromatography. L-Serine in the drinking water increased L-serine levels in some brain areas, but changes in its metabolites were almost negligible. L-Serine decreased locomotor activity in rats exposed to a familiar environment. In addition, L-serine decreased exploratory behavior of isolated rats, even in a novel environment. Our results could suggest that daily intake of L-serine can attenuate symptoms induced by chronic stress.

Intracerebroventricular injection of L-aspartic acid and L-asparagine induces sedative effects under an acute stressful condition in neonatal chicks.

The present study was conducted to clarify the central functions of L-aspartic acid (Asp) and L-asparagine (Asn) during an acute stressful condition in chicks. Intracerebroventricular (i.c.v.) injection of Asp and Asn (0.84 micromol) attenuated the vocalization that normally occurs during social separation stress. Asp decreased the time spent in active wakefulness and induced sedation. Asn had a similar effect to Asp, although somewhat weaker. However, i.c.v. injection of Asp and Asn further enhanced plasma corticosterone release under social separation stress. Taken together, the i.c.v. injection of Asp and Asn has sedative effects under an acute stressful condition, which does not involve the hypothalamic-pituitary-adrenal axis.

Central administration of dipeptides, beta-alanyl-BCAAs, induces hyperactivity in chicks

BackgroundCarnosine (β-alanyl-L-histidine) is a putative neurotransmitter and has a possible role in neuron-glia cell interactions. Previously, we reported that carnosine induced hyperactivity in chicks when intracerebroventricularly (i.c.v.) administered. In the present study, we focused on other β-alanyl dipeptides to determine if they have novel functions.ResultsIn Experiment 1, i.c.v. injection of β-alanyl-L-leucine, but not β-alanyl-glycine, induced hyperactivity behavior as observed with carnosine. Both carnosine and β-alanyl-L-leucine stimulated corticosterone release. Thus, dipeptides of β-alanyl-branched chain amino acids were compared in Experiment 2. The i.c.v. injection of β-alanyl-L-isoleucine caused a similar response as β-alanyl-L-leucine, but β-alanyl-L-valine was somewhat less effective than the other two dipeptides. β-Alanyl-L-leucine strongly stimulated, and the other two dipeptides tended to stimulate, corticosterone release.ConclusionThese results suggest that central β-alanyl-branched chain amino acid stimulates activity in chicks through the hypothalamus-pituitary-adrenal axis. We named β-alanyl-L-leucine, β-alanyl-L-isoleucine and β-alanyl-L-valine as Excitin-1, Excitin-2 and Excitin-3, respectively.

Central L-cysteine induces sleep, and D-cysteine induces sleep and abnormal behavior during acute stress in neonatal chicks.

L-cysteine (L-Cys) is a non-essential and glycogenic amino acid. Previously, we reported that the intracerebroventricular (i.c.v.) injection of L-Cys induced sedative effects under isolation-induced stress in neonatal chicks. L-Cys has an optical isomer, D-Cys. The aim of this study was to clarify the effect of L-Cys and D-Cys during a stressful condition in chicks. The i.c.v. injection of L-Cys and D-Cys (0.84 micromol) decreased both distress vocalization and spontaneous activity induced by isolation. However, the two cysteine isomers induced different behaviors. L-Cys increased sleep-like behavior while D-Cys caused abnormal behavior including syncope as well as sleep-like behavior. In conclusion, while both L-Cys and D-Cys caused a sedative effect when injected i.c.v, D-Cys caused abnormal behavior and may be detrimental to neonatal chicks.