Jun'ichi Semba

DIFFERENTIAL EXPRESSION OF c-fos mRNA IN RAT PREFRONTAL CORTEX, STRIATUM, N. ACCUMBENS AND LATERAL SEPTUM AFTER TYPICAL AND ATYPICAL ANTIPSYCHOTICS: AN IN SITU HYBRIDIZATION STUDY

The regional difference in the expression of c-fos mRNA induced by typical and atypical antipsychotics was determined in prefrontal cortex, striatum, N. accumbens and lateral septum in rats by in situ hybridization. Two typical antipsychotics, haloperidol (2 mg/kg) and fluphenazine (2 mg/kg), and three atypical antipsychotics, (-)sulpiride (100 mg/kg), clozapine (20 mg/kg) and OPC-14597 (40 mg/kg), were used. Brains were fixed with 4% paraformaldehyde 45 min after drug administration (i.p.). Brain sections of 30 microns-thickness were made in a cryostat and hybridized with 35S-labelled for c-fos oligonucleotide probe. These sections were apposed to X-ray films and the autoradiograms were semi-quantitatively analysed by computer-assisted densitometry. All antipsychotics used increased c-fos mRNA expression in N. accumbens shell, a region of the forebrain associated with limbic systems. On the other hand, two typical antipsychotics (haloperidol and fluphenazine) that cause a high incidence of acute motor side effects increased the expression of c-fos mRNA in the dorsolateral striatum, an extrapyramidal region primarily involved in motor control. Only clozapine induced c-fos mRNA in the medial prefrontal cortex and lateral septum. These results strongly suggest that the shell region of N. accumbens may be a common site of therapeutic action of antipsychotics.

Chronic lithium chloride injection increases glucocorticoid receptor but not mineralocorticoid receptor mRNA expression in rat brain.

Lithium has been used clinically for the treatment of bipolar disorders. However, the brain mechanisms, by which lithium acts, are still unclear. An impaired hypothalamic-pituitary-adrenal (HPA) axis has been implicated in the pathogenesis of mood disorders. In this study, we investigated the effects of chronic lithium on the corticosteroid receptors in the brain. Male Wistar rats were injected with LiCl (1.5 mEq/kg) or saline intraperitoneally (i.p.) once a day for 14 days. Twenty-four hours after the last injection, the expressions of mRNA for glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) in the brain were determined by non-radioactive in situ hybridization. Chronic administration of LiCl increased the expression of GR mRNA in the hippocampus and paraventricular nucleus of the hypothalamus (PVN). However, no significant changes were observed in the expression of either MR mRNA in the hippocampus or GR mRNA in the locus ceruleus. Since the hippocampus and PVN mediate negative feedback regulation of the HPA axis, an increased expression of GR mRNA in these regions may normalize HPA axis activity in mood disorders. Thus, the effect of chronic lithium on GR function may be involved in its antimanic and/or prophylactic activity in bipolar disorders.

The effects of L-threo-dihydroxyphenylserine on norepinephrine metabolism in rat brain

The effects of L-threo-3,4-dihydroxyphenylserine (DOPS), an artificial precursor of norepinephrine (NE), on NE metabolism in rat brain were investigated. DOPS administration resulted in a significant elevation in cerebral NE and 3-methoxy-4-hydroxyphenylglycol contents, while carbidopa pretreatment completely blocked these increases. After brain NE was depleted by either alpha-methyl-p-tyrosine (AMPT), fusaric acid, FLA-63, or reserpine, NE restoration by DOPS was observed in rats treated with either fusaric acid or chronic reserpine. No NE restoration was observed after pretreatment with AMPT, FLA-63, or acute reserpine. The results suggest that NE formed after DOPS administration is mainly localized in the brain capillaries. In some NE-depleting conditions, however, DOPS can penetrate the brain parenchyma.

Effect of monoamine precursors on the forced-swimming test in mice

The antidepressant properties of monoamine precursors were evaluated by the forced-swimming test for mice developed by Porsolt et al. DOPA but not 5-hydroxy-tryptophan (5HTP) shortened immobility at doses that did not increase locomotor activity. Although l-threo-dihydroxyphenylserine (DOPS), an artificial norepinephrine (NE) precursor, did not change immobility in intact mice, DOPS significantly reduced immobility in mice pretreated with the selective NE neurotoxin DSP4. These results suggest possible antidepressant properties of DOPA and DOPS, the latter of which may act as an antidepressant in a certain NE-depleting condition.

Determination of free and total 3-methoxy-4-hydroxyphenylethylene glycol in human plasma by high-performance liquid chromatography with electrochemical detection

A new assay method for the determination of free and total 3-methoxy-4-hydroxyphenylethylene glycol (MHPG) in human plasma is described. MHPG was purified with a Bond Elut PH column followed by ethyl acetate extraction. High-performance liquid chromatography (HPLC) with electrochemical detection was used for separation and detection of MHPG. Total MHPG was measured after enzymatic hydrolysis with sulfatase type H-5. 3-Hydroxy-4-methoxyphenylethylene glycol (iso-MHPG) was used as an internal standard to correct the recovery of extraction. One assay could be completed within 20 min with a short reverse-phase column. This technique is sensitive, reliable and less time-consuming than other HPLC methods. With this method, the plasma values of MHPG in healthy controls were in good agreement with those using gas chromatography-mass spectrometry.

Neonatal phencyclidine treatment selectively attenuates mesolimbic dopamine function in adult rats as revealed by methamphetamine-induced behavior and c-fos mRNA expression in the brain

One of the major hypotheses regarding the pathogenesis of schizophrenia is the implication of neurodevelopmental abnormality. However, the mechanism of delayed onset of schizophrenic symptoms, in which increased dopaminergic activity in mesolimbic or mesocortical dopamine systems plays a pathological role, is not known. In this study, we investigated whether the chronic blockade of N‐methyl‐D‐aspartate (NMDA) receptor by phencyclidine (PCP), an NMDA channel blocker, during development could disrupt the dopamine system during later life. Neonatal rats were injected with PCP subcutaneously daily from postnatal day (PD) 1 to PD 14 and their dopaminergic function was evaluated on PD 42 by rating the methamphetamine (MAP)‐induced behavior. To illustrate the activated brain regions, the expression of c‐fos mRNA in response to a MAP challenge was also studied utilizing in situ hybridization. Chronic neonatal PCP treatment attenuated MAP‐induced oral stereotypy (licking and gnawing) and reduced MAP‐induced expression of c‐fos mRNA in the N. accumbens shell region and VTA but not in the N. accumbens core region, medial striatum, or substantia nigra. These results suggest that neonatal blockade of NMDA receptor, which induces a number of effects in the developing nervous system, may cause long‐lasting functional changes of the mesolimbic dopamine system. Synapse 40:11–18, 2001.

Measurement of glutamate, aspartate and glycine and its potential precursors in human brain using high-performance liquid chromatography by pre-column derivatization with diethylaminoazobenzene sulphonyl chloride

This paper describes a high-performance liquid chromatographic technique, with dimethylaminoazobenzene sulphonyl chloride derivatization, for the measurement of glutamate, aspartate and glycine and its potential precursors in human brain tissue. The derivatization procedure is simple, sensitive and highly reproducible. The derivatized amino acids are stable and can be analysed by reversed-phase chromatography with visible detection at an absorption wavelength of 436 nm. A preliminary application to the determination of the concentrations of several amino acids in several regions of the human brain is described.

Neonatal treatment with L-name (NG-nitro-L-arginine methyl ester) attenuates stereotyped behavior induced by acute methamphetamine but not development of behavioral sensitization to methamphetamine.

1. The neurodevelopmental hypothesis of schizophrenia postulates that disturbed nitric oxide (NO) function during neuronal development is one of premorbid factors for schizophrenia in later life. 2. The aim of present study is to investigate behaviorally whether neonatal inhibition of nitric oxide synthase (NOS) affects dopaminergic function, the abnormality of which may be ascribed to a major pathophysiology of schizophrenia. 3. Male rat pups were injected daily with NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), from postnatal day (PD) 1 to 14. 4. When methamphetamine (MAP) was challenged on PD42, MAP-induced stereotypy was significantly attenuated in the L-NAME treated rats. The development of sensitization to the stereotypy-inducing effect of MAP, however, was not prevented with neonatal L-NAME. 5. These results suggest that decreased NO production during neonatal period may disturb normal maturation of dopaminergic system and result in impaired dopaminergic function in adult period.

A postmortem study of glycine and its potential precursors in chronic schizophrenics

We have measured the concentrations of glycine and its potential precursors, serine and threonine, in 20 areas of the postmortem brains of chronic schizophrenics and controls using high-performance liquid chromatography by pre-column derivatization with dimethyl-amino-azobenzene sulphonyl chloride. The regional distribution pattern of glycine in the postmortem brains with and without the disease was more similar to that of serine (r = 0.874, P < 0.0001) than to that of threonine (r = 0.476, P < 0.01). A multiple regression analysis with regressor variables including diagnosis, age at death and interval between death and freezing revealed that there is a significant difference between schizophrenics and controls in the contents of these amino acids in a number of brain areas. The level of glycine in the orbitofrontal cortex of schizophrenics was found to be significantly increased in schizophrenics, with a tendency to an increase in that of serine. The increase in glycine was also significantly high in the off-drug group of schizophrenics who had not taken antipsychotics more than 40 days before death. Prominent decreases in both glycine and serine were observed in the somesthetic cortex of the on-drug schizophrenics. Serine was found to be significantly decreased in the putamen of the off-drug schizophrenics. A marked decrease in threonine was also observed in the supramarginal cortex and posterior portion of the lateral occipitotemporal cortex of the off-drug group of schizophrenics and in the putamen of all schizophrenics. The highly similar distribution pattern of glycine and serine in the postmortem brains supports the close coupling of synthesis and metabolism between these chemicals in human brains. The increased content of glycine in the orbitofrontal cortex, the reduced level of serine in the putamen and the decrease in threonine in the cerebral cortices, which were prominent in the off-drug schizophrenics, may be involved in the pathophysiology of schizophrenia.

Neuroendocrinological Effects of L‐Threo‐3, 4‐Dihydroxyphenylserine (DOPS), a Putative Norepinephrine Precursor, on Healthy Volunteers

Abstract: The effect of L‐threo‐3, 4‐dihydroxyphenylserine (DOPS) on plasma Cortisol, prolactin, thyrotropin‐stimulating hormone (TSH) and growth hormone concentrations was studied in nine healthy male volunteers. The drug was administered orally (300 mg or 600 mg DOPS) using a multiple crossover placebo‐controlled study design. Plasma hormone concentrations were measured at 30 minute intervals for 3 hours after dosing. Plasma DOPS peak concentrations were observed between 2 and 3 hours after dosing. DOPS, however, had no effect on plasma hormone concentrations and this may be attributed to the known low brain permeability of DOPS in healthy subjects.