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The metallothionein (MT) family is a class of low molecular, intracellular, and cysteine-rich proteins with a high affinity for metals. Although the first of these proteins was discovered nearly 40 years ago, their functional significance remains obscure. Four major isoforms (MT-I, MT-II, MT-III, and MT-IV) have been identified in mammals. MT-I and MT-II are ubiquitously expressed in various organs including the brain, while expression of MT-III and MT-IV is restricted in specific organs. MT-III was detected predominantly in the brain, and characterized as a central nervous system-specific isomer. The role of MTs in the central nervous system has become an intense focus of scientific research. An isomer of MTs, MT-III, of particular interest, was originally discovered as a growth inhibitory factor, and has been found to be markedly reduced in the brain of patients with Alzheimer's disease and several other neurodegenerative diseases. MT-III fulfills unique biological roles in homeostasis of the central nervous system and in the etiology of neuropathological disorders.

We examined the effects of FK506 and its non-immunosuppressive derivative, GPI1046, on H2O2-induced reduction of cell viability and apoptotic cell death in Neuro 2A cells. Our results suggest that the protective properties of GPI1046 against H2O2-induced reduction of cell viability are equipotent with those of FK506 and may be mediated by increased intracellular concentrations of glutathione (GSH). In addition, both FK506 and GPI1046 prevented apoptotic cell death in Neuro 2A cells, although the antiapoptotic effect of FK506 was somewhat stronger than that of GPI1046. These findings suggest that non-immunosuppressive immunophilin ligands such as GPI1046 might be potentially useful in treatment of neurodegenerative diseases without serious side effects such as immune deficiency.

Changes in the distribution and densities of muscarinic acetylcholine receptors (mACh-R) in young adult and aged rat brain, and the effects of chronic administration of lisuride hydrogen maleate (lisuride) on these changes were studied by in vitro quantitative autoradiography of [3H] quinuclidinyl benzilate binding. mACh-R was relatively higher in the striatum, hippocampus and cerebral cortex in the young adult rats. In contrast, mACh-R binding was markedly reduced in the striatum, accumbens nucleus, amygdaloid nucleus and frontal cerebral cortex of aged rats compared to young adult rats. However, chronic administration of lisuride significantly increased mACh-R binding in the parietal cerebral cortex as well as in the above-mentioned regions in aged rats. This lisuride-induced increase in mACh-R of aged rat brain is considered to have important implications concerning the mechanism of therapeutic efficacy of lisuride.

To gain further insight into the central nervous system (CNS)-action of beta-adrenergic blocking agents (beta-blockers), we examined the effects of various kinds of beta-blockers on opioid receptors (Op-Rs) using radiolabeled receptor assay (RRA). We demonstrated that beta-blockers are competitively bound to Op-Rs in the CNS. Sodium index of beta-blockers in [3H]naloxone binding study indicated that beta-blockers had the mixed agonist-antagonist activity of opiates. The relative potency of beta-blockers in opioid RRA was negatively correlated with their membrane stabilizing activity. Neither beta-blocking activity nor intrinsic sympathomimetic activity was correlated with IC50 values of beta-blockers in opioid RRA. While it is widely accepted that beta-blockers have a tranquilizing activity, a part of the tranquilizing action of beta-blockers may be mediated through Op-Rs in the CNS. Although beta-blockers may have effects on their own receptors (beta-receptors) in the CNS, the more precise mechanisms of central action of these drugs must be further investigated.

We have previously reported that ropinirole, a non-ergot dopamine agonist, has neuroprotective effects against 6-hydroxydopamine in mice based on in vivo antioxidant properties such as the glutathione (GSH)-activating effect. In the present study, we determined that the effects of ropinirole on the level of expression of GSH-related enzyme mRNA, these enzymes were shown to regulate GSH contents in the brain. This study focused on the mechanism of GSH enhancement by ropinirole. Striatal GSH contents were significantly increased by 7-day daily administration of ropinirole. Furthermore, the expression levels of gamma-glutamylcysteine synthetase (gamma-GCS), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione S-transferase (GST) mRNA increased following daily injections of ropinirole for 7 days. In addition, ropinirole treatment for 7 days suppressed auto-oxidation in mouse striatal homogenates, in contrast to the vehicle treatment. In conclusion, ropinirole was able to suppress auto-oxidation, most probably by increasing GSH levels due to an increase of GSH synthesis. In addition, it is likely that auto-oxidation was also suppressed by the activation of GSH-regulating enzymes such as GPx, GR, and GST in the mouse striatum. Thus, our results indicate that the GSH-activating effect of ropinirole may render this dopamine agonist beneficial as a neuroprotective drug.

The delta opioid peptide [D-Ala2, D-Leu5]enkephalin (DADLE) has been reported to block the neurotoxicity induced by multiple administrations of a moderate dose of methamphetamine (METH). We examined in this study if DADLE might block the neurotoxicity caused by a single high dose of METH in CD-1 mice. The levels of dopamine transporter (DAT), tyrosine hydroxylase (TH), major biogenic amines including DA, 5-hydroxytryptamine (5-HT), and their metabolites were examined. In addition, since the tumor suppressor p53 has been implicated in the neurotoxicity of METH, this study also examined the levels of p53 mRNA and protein affected by METH and DADLE. METH (25 mg/kg, i.p.) caused significant losses of DAT, TH, DA, 3,4-dihydroxyphenylacetic acid (DOPAC), and 5-HT in the striatum within 72 h. The administration of a single dose of DADLE (20 mg/kg, i.p., 30 min before METH) caused a complete blockade of all losses induced by METH except for that of the DA content (a approximately 50% blockade). DADLE did not affect the changes of rectal temperature induced by the administration of the high dose of METH. METH increased p53 mRNA in the striatum and the hippocampus of CD-1 mouse. DADLE abolished the p53 mRNA increase caused by METH. METH tended to increase the p53 protein level at earlier time points. However, METH significantly decreased the p53 protein level by about 30% at the 72-h time point. DADLE blocked both the increase of p53 mRNA and the decrease of p53 protein caused by METH. These results demonstrate a neuroprotective effect of DADLE against the neuronal damage and the alteration of p53 gene expression caused by a single high dose of METH. The results also indicate an apparent discordance between the protein level of p53 and the neurotoxicity caused by a high dose of METH. Synapse 39:305-312, 2001. Published 2001 Wiley-Liss, Inc.

Bromocriptine, a dopamine agonist, is commonly used in combination with levodopa for the treatment of Parkinson's disease (PD). To investigate the theoretical basis of such combination therapy, we examined the effects of bromocriptine administered alone or in combination with levodopa on dopamine turnover in the striatum of hemi-parkinsonism rats. The parkinsonian striatum showed a 3.4-fold increase of dopamine turnover relative to the control striatum, as often observed in the brain of PD patients. A 7-day course of levodopa therapy markedly increased dopamine turnover in the parkinsonian striatum (53-fold of control level) than in the control striatum (5-fold of the control level). However, bromocriptine specifically and markedly suppressed the levodopa-induced abnormal activation of dopamine turnover in the parkinsonian striatum. Our findings explain the pharmacological basis for the introduction of bromocriptine during long-term levodopa therapy.

We examined the neuroprotective effects of oren-gedoku-to (TJ15), a herbal medicine, after transient forebrain ischemia. Transient forebrain ischemia was induced by occlusion of both common carotid arteries for 15 min in C57BL/6 mice treated with TJ15. In the control ischemic group without TJ15 treatment, histologic examination of brain tissue collected seven days after reperfusion showed death of pyramidal cells in CA2-3 area of the hippocampus, unilaterally or bilaterally. In mice treated with oral TJ15 (845 mg/kg/day) for five weeks, the frequency of ischemic neuronal death was significantly lower. Immunohistochemistry for Cu/Zn-superoxide dismutase (Cu/Zn-SOD) showed strongly reactive astrocytes in the hippocampus of ischemic mice treated with TJ15. Damage to nerve cells by free radicals plays an important role in the induction of neuronal death by ischemia-reperfusion injury. Our results suggest that TJ15 protects against ischemic neuronal death by increasing the expression of Cu/Zn-SOD and suggest that oren-gedoku-to reduces the exposure of hippocampal neurons to oxidative stress.

Free radicals are involved in neuronal damage. Bifemelane hydrochloride has been reported to protect neural tissues against ischemic damage and age-related neurodegeneration. We examined the protective effects of bifemelane HCl and the relation between its effectiveness and free radical formation in hydrogen peroxide (H2O2)-induced cytotoxicity using cultured rat neuroblastoma cell line (B50). Cytotoxicity was examined by using the lactate dehydrogenase (LDH) assay and cell viability by the WST-1 assay. H2O2 reduced the survival of B50 cells in a dose-dependent manner, and treatment of these cells with 75 microM or 100 microM H2O2 reduced their viability by 50% relative to the control group. B50 cells were treated with 5 or 10 microM bifemelane for 2 days followed by treatment with 75 microM or 100 microM H2O2. H2O2 cytotoxicity was reduced by pretreatment with bifemelane. We also examined the effect of bifemelane on lipid peroxide formation in B50 cells using thiobarbituric acid reactive substances assay. Pretreatment of B50 cells with 10 microM bifemelane for 2 days reduced lipid peroxide formation to approximately 54% of the control group. Our results suggest that bifemelane hydrochloride provides a protective effect against H2O2 cytotoxicity partly due to its anti-oxidative properties.

To determine whether nitric oxide (NO) acts as a modulator of muscarinic acetylcholine receptor (mACh-R) function, we performed a radioligand receptor assay using [3H]quinuclidinyl benzylate ([3H]QNB), the NO radical (NO*) donor 3-(2-Hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-propanamin e (NOC7) and a gerbil brain cortical membrane preparation. NOC7 (at 10 microM, 100 microM or 1 mM concentrations) significantly reduced the [3H]QNB binding Kd values (from 0.196 +/- 0.009 nM in the control, to 0.151 +/- 0.013, 0.144 +/- 0.012 and 0.153 +/- 0.007 nM respectively). NOC7 did not alter the displacement curves of atropine or carbachol. Reduction of SH groups with dithiothreitol, in the presence of the NO donor, significantly increased [3H]QNB binding affinity whereas alkylation by N-ethylmaleimide markedly decreased it. The observed enhancing effect on mACh-R binding affinity for [3H]QNB, may reflect conformational changes in the receptors mediated by the NO generated, and these changes might be explained by NO reactions with such groups through conditions supporting redox reactions intrinsic to the NO molecule, similar to those occurring in redox regulatory sites reported for other neurotransmitter pathways in the CNS.

Recent evidence has suggested that cyclosporin A (CsA), an immunosuppressive agent, has neuroprotective properties. However, its mechanisms associated with this activity remain unclear. We have previously shown that post-ischemic administration of CsA daily for 14 days prevented the decrease of muscarinic acetylcholine receptor binding in the hippocampus in the gerbil model of 5-min transient forebrain ischemia. In the present study, CsA (5 mg/kg, subcutaneously) was administered to each animal just after, 2 and 6 h after ischemia so as not to exert its immunosuppressive effect. Initial CsA treatment significantly restored the declined muscarinic acetylcholine receptor binding of the hippocampus 14 days after ischemia similar to the previous report. However, CsA did not alter reactive changes of astrocytes and microglia in the CA1 area of the hippocampus, which had been suppressed by daily administration. These results indicate that CsA could positively modulate the hippocampal acetylcholine neurotransmission system broken down through the ischemia-induced pyramidal cell death and its action mechanism may have no relation to the immunosuppressive properties.

To analyze the effect of lecithinized superoxide dismutase (SOD) on superoxide accumulation after traumatic brain injury (TBI) in rats, we studied the SOD activity by NBT-reducing method and the expression of Cu,Zn-SOD mRNA by Northern blot analysis. As determined by the specific gravity method, the administration of lecithinized SOD decreased brain edema in the periphery of the lesion at 6 hr after contusion. SOD activity, without lecithinized SOD administration, increased at the peripheral portion at 30 min after contusion, but decreased to normal level at 6 hr after TBI. By administration of lecithinized SOD, the increase of SOD activity was preserved until 6 hr after TBI. The expression of Cu,Zn-SOD mRNA increased in the core lesion, peripheral portion, and contralateral hemisphere until 6 hr after TBI, then was suppressed in all three areas by lecithinized SOD. These results support the hypothesis that superoxide anions may play an important role in the development of brain edema after TBI, and that leciyhinized SOD appears to prevent brain edema through a protective effect against superoxide anions.

The level of lipid peroxidation reflects the degree of free radical-induced oxidative damage in brain tissue of the elderly. We examined the effects of Manda, a product prepared by yeast fermentation of several fruits and black sugar, on lipid peroxidation in the senescent rat brain as model of aging. Senescent rats were provided with a diet containing 50 g/100 g Manda for 8 days, supplemented on day 8 with an intragastric administration of Manda (6.0 g/kg body wt.) twice daily. The hydroxyl radical scavenging activity was generated by the FeSO4-H2O2 system and analyzed by electron spin resonance spectrometry. Using this method, the addition of Manda (2.88 mg/ml) to brain homogenates of adult rats (0.06 mg/ml) had an additive inhibitory effect on lipid peroxidation compared with control adult rats not treated with Manda. Incubation of brain homogenates with Manda for 2 h and 3 h, significantly inhibited the increase in lipid peroxides (malondialdehydes and 4-hydroxyalkenals) levels in aged rats due to auto-oxidation. In addition, oral administration of Manda significantly suppressed the age-related increase in lipid peroxidation in the hippocampus and striatum, although such change was not observed in the cerebral cortex. Although Manda contains trace level of alpha-tocopherol, the level of alpha-tocopherol in Manda did no correlate with its antioxidant effect. Our results suggest that Manda protects against age-dependent oxidative neuronal damage caused by oxidative stress and that this protective effect may be due, in part, to its scavenging activity against free radicals.

We examined the effect of Oren-gedoku-to (TJ15), which is a traditional herbal Kampo prescription used as an anti-cerebral apoplexy agent on these changes. Chronic pre- and post-ischemia TJ15 oral administration almost completely abolished the ischemia-induced muscarinic receptor reduction and 5-hydroxyindoleacetic acid level increase. These results suggest that TJ15 prevents cholinergic synaptic dysfunction and serotonergic presynaptic hyperactivity induced by transient ischemia.

To analyze the effect of lecithinized superoxide dismutase (SOD) on superoxide accumulation after traumatic injury, the expression of Cu,Zn-SOD mRNA was examined after contusion in rat using Northern blotting. As determined by specific gravity, lecithinized SOD decreased brain edema. The expression of Cu,Zn-SOD mRNA increased at the core, peripheral and contralateral hemisphere of injury. These increases were then suppressed by lecithinized SOD. Our results support the hypothesis that superoxide may play an important role in edema formation after contusion, and that lecithinized SOD appears to prevent brain edema through a protective effect against superoxide injury.

In a cultured hybrid neuronal cell line (BIM) which was produced between human neuroblastoma cells (IMR32) and thymidine auxotrophs (B3T) of rat nerve-like cells (B103), the mRNAs encoding ciliary neurotrophic factor (CNTF) and neurotrophins were detected by the polymerase chain reaction method. The conditioned medium of BIM cells enhanced choline acetyltransferase (ChAT) activity in septal neurons and survival of ciliary ganglion neurons. The mRNA expression of CNTF and neurotrophins in BIM cells was differently regulated by the stimulation with cAMP, FGF and retinoic acid. These data suggest multiple regulation and collaboration of neurotrophic factors.

To clarify the effects of levodopa administration on MPTP-induced alterations in neuropeptides, we examined the effects of repeated levodopa injections (200 mg/kg i.p.) for 2 weeks starting 4 weeks after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment (30 mg/kg i.p. twice/day for 5 days) on cholecystokinin-octapeptide (CCK-8), substance P (SP) and thyrotropin-releasing hormone (TRH) concentrations at 6 weeks after the MPTP treatment. In the striatum, CCK-8 significantly but slightly decreased in the MPTP-treated mice, coinciding with the MPTP-induced marked reduction of dopamine (DA). This considerable reduction of striatal CCK-8 may result from the selectivity of MPTP since the mesolimbic DA neurons coexisting with CCK-8 are intact with the MPTP treatment. Furthermore, this MPTP-induced decrease in CCK-8 persisted with repeated levodopa administration; therefore, the ineffectiveness of the levodopa treatment may have been be due to the degeneration of the nigrostriatal DA neurons. SP and TRH contents showed little or no change with levodopa treatment in the MPTP-treated mouse brain. The CCK-8 level decreased in the thalamus+midbrain, hippocampus and hindbrain of the MPTP+levodopa-treated group, although there were no changes in the MPTP-treated controls. These results suggest that DAergic neurons, except those in the nigrostriatum, strongly interact with the CCK neurons in these brain regions.

Pergolide, along with bromocriptine and lisuride, is one of the most active dopamine receptor agonists. To determine whether or not pergolide protects against dopaminergic neuronal damage, via its activity on monoamine metabolism, we studied the effects of pergolide pretreatment on changes in monomaines and their metabolites in the mouse striatum after intracerebroventricular injection of 6-hydroxydopamine with pretreatment of desipramine. After intracerebroventricular administration of 6-hydroxydopamine (40 micrograms) in mice, the levels of dopamine and its metabolites (DOPAC, HVA) in the striatum rapidly decreased to 49%, 29% and 68%, respectively, of the naive controls at week 1 but then gradually recovered to control levels at weeks 2 and 4. Repeated pretreatment with pergolide (0.5 mg/kg, i.p.) for 7 days before administration of 6-hydroxydopamine, almost completely protected against reduction in striatal dopamine and its metabolites 1 week after injection of 6-hydroxydopamine. Therefore, pergolide could normalize the decreased dopamine synthesis or storage, and has a neuroprotective effect against dopaminergic dysfunction induced by the neurotoxin, 6-hydroxydopamine. Although we found that pergolide did not show radical scavenging activity in an in vitro system that generated hydroxyl radicals, it has been reported in vivo that pergolide treatment may induce Cu/Zn superoxide dismutase in the rat striatum. Considering these findings, pergolide may well be protective to dopaminergic neurons, largely because of its effects on presynaptic autoreceptors and on its induction of Cu/Zn superoxide dismutase. Further research on the neuroprotective effects of pergolide in Parkinson disease models, by injection of 6-hydroxydopamine, is needed to clarify its mechanism of action on dopaminergic indices.

In order to verify whether angiotensin II (Ang II) induced in vivo protooncogene, c-fos, expression in the rat renal cortex and medulla, we administered various concentrations of Ang II to Wistar rats and measured the c-fos expression from the renal cortex and medulla using the method of Northern hybridization. c-fos expression induced by 1 microgram (1.6 x 10(-6) M) of atrial natriuretic peptide (ANP) was also examined. The result was that the peak expression of c-fos mRNA was observed at approximately 10 min after Ang II administration in both rat renal cortex and medulla. This expression was reduced to the control level at 30 min. The measurement of the concentration of injected-Ang II and c-fos mRNA expression revealed that the peak expression of c-fos mRNA in the renal cortex and medulla was detected at the concentration of 1.0 x 10(-8) M and 1.0 x 10(-9) M Ang II, respectively. Nevertheless, ANP had no significant effect on the increase in c-fos mRNA expression. These data revealed that Ang II transiently increases the in vivo c-fos expression in both rat renal cortex and medulla but ANP does not. This protooncogene expression may induce vascular and mesangial proliferation in the kidney.

Regional changes in the mRNA accumulations for cytoskeletal proteins alpha-tubulin and beta-actin were examined by in situ hybridization and Northern blot analysis in spontaneously hypertensive rat brains at chronic stages after 3 hours of transient ischemia. alpha-Tubulin mRNA accumulations showed no significant change at 2 weeks after transient ischemia except for a significant decrease in the frontal cortex (9.7%, p0.01) coinciding with ischemia induced histological changes. beta-Actin mRNA level was significantly increased in the parietal cortex (8.5%), septum (10.0%), amygdala (11.0%), CA4 area (5.8%) and the dentate gyrus (7.5%) of the hippocampus at 2 weeks after recirculation compared with a sham-operated control group (p0.01). The ischemic areas of hippocampal and frontocortical lesions receive afferent neurons from those regions where beta-actin mRNA was increased, suggesting that ischemia-induced increases in beta-actin mRNA may reflect actin synthesis in these neurons to compensate for lost synaptic connections. Two cytoskeletal mRNA concentrations reacted differently to cerebral ischemia, and did not parallel histological signs of ischemia either temporally or spatially.

No definitive evidence for the participation of the immune system in progressive brain damage has been previously reported. However, glial cells continue to accumulate after degeneration of neurons appears to be completed, and a recent study showed that microglia and leukocytes also accumulate after brain damage. Thus, it seemed possible that immune responses might play a role in the delayed effects. Cyclosporin A (CsA) is a cyclic undecapeptide of fungal origin with a strong immunosuppressive action but low myelotoxicity. We examined the effect of CsA administration on three different kinds of animal models for neurological deficits. Late onset reduction of muscarinic receptors after transient forebrain ischemia in gerbils was prevented by daily post-ischemic administration of CsA. This indicates that an immune mechanism may be involved in the progressive brain damage occurring after transient ischemia. On the other hand, CsA exacerbated iminodipropionitrile-induced dyskinesia both behaviorally and biochemically. CsA also mimicked pentylenetetrazol-induced seizures. These findings suggest that immune mechanisms may play important roles in the progression of brain damage and possibly that immunosuppressants might open a new chapter in the pathophysiology and treatment of chronic progressive neurodegenerative diseases. Further investigations on the immune response in the progressive brain damage are needed.

To determine ischemia-induced changes in phosphodiesterase (PDE), changes in the membranous binding sites of rolipram, a cAMP-selective PDE inhibitor, were examined in the gerbil brain following transient 5 min forebrain ischemia. Coinciding with the delayed neuronal death (DND) in the hippocampal CA1 region, affinities for cerebral rolipram bindings decreased on Day 4, when intrinsic cAMP, substrate for PDE, might increase. The number of rolipram binding sites was significantly reduced in the hippocampus Day 14, despite the lack of change on Day 4. This reduction in rolipram binding was in agreement with the previously reported late onset reduction of muscarinic receptors, progressing more slowly than DND. Slowly progressive mechanisms may be involved in the ischemia-induced reduction of the hippocampal rolipram binding sites which may be PDEs.

The effects of a new acetylcholinesterase inhibitor, ENA-713, on ischemia-induced changes in acetylcholine, monoamines, and their metabolites, were studied in the gerbil. ENA-713 (0.2 mg/kg) or saline was administered intraperitoneally to gerbils 30 min before induction of cerebral ischemia by bilateral carotid occlusion. Pretreatment with ENA-713 mitigated the ischemia-induced abnormalities of the cholinergic, dopaminergic and serotoninergic systems in the gerbil brain, although it had virtually no effect on acetylcholine, monoamines, or their metabolites in any region of the normal gerbil brain. These findings suggest that ENA-713 has beneficial effects against ischemia-induced cerebral disorders. Thus, ENA-713 seems to be promising as a preventive or therapeutic agent for cerebrovascular dementia due to cerebral ischemia and might be useful for the treatment of Alzheimer-type dementia which is associated with multiple neurotransmitter abnormalities in the brain.

A patient with progressive supranuclear palsy (PSP) showing severe akinesia, postural instability and gait disturbance like frozen gait was treated with amitriptyline (30 mg/day). Four weeks after drug administration, we recognized tendencies for improvement on akinesia, postural instability, frozen gait and vertical gaze palsy. Thereafter, gait disturbance almost disappeared and slow motor response and depression also improved with the amitriptyline for 8 weeks. The changes of cerebral blood flow (CBF) with amitriptyline treatment were evaluated in this case by 123I-IMP single photon emission computed tomography. Before the treatment, clear reduction of CBF was shown in the frontal lobes, especially in the frontal cortices, while this frontal hypoperfusion markedly improved 6 weeks after the treatment. These results, showing marked improvements of frontal hypoperfusion, as well as parkinsonism with amitriptyline treatment, may imply possibilities that these symptoms of PSP result from the frontal hypoperfusion and the improvement of the hypoperfusion with amitriptyline is involved in the improvement of these symptoms.

Super-delayed loss of muscarinic-1 (M1) receptors and the protective effect of propentofylline on these changes were examined in the gerbil hippocampus after transient ischemia with radioactive pirenzepine. M1 receptors were markedly decreased in the gerbil hippocampus 14 days after transient ischemia. Single administration of propentofylline (20 mg/kg, ip) just after transient ischemia almost completely prevented the ischemia-induced decrease in the number of M1 receptors, and had no effect in sham-operated controls. These findings suggest that one of the therapeutic efficacies of propentofylline is the result of the normalization of the dysfunction of the acetylcholine neuronal system in cerebrovascular disease.

The effects of bifemelane hydrochloride on changes of N-methyl-D-aspartate receptors and muscarinic cholinergic receptors were examined in the gerbil hippocampus after transient ischemia with radioactive-specific ligands. There were marked reductions in both these receptors in the gerbil hippocampus 14 days after transient ischemia, without changes in the respective affinities. Post-ischemia bifemelane treatment almost completely prevented the ischemia-induced decreases in the numbers of these receptors, and had no effect in sham-operated controls. The results of the present study suggest that this drug prevents postsynaptic dysfunction induced by transient ischemia, and may be useful in the therapy of both the chronic and the acute stage of cerebrovascular disease.

Cerebral monoaminergic neurotransmitters and their metabolites show various concentration changes in gerbils following bilateral carotid ligation. The present study evaluated the effect of chronic administrations of lisuride hydrogen maleate (lisuride) on these changes. Lisuride (0.01 mg/kg or 0.05 mg/kg) or vehicle was intraperitoneally administered to gerbils for 14 consecutive days before the induction of a 30 min ischemia by bilateral carotid ligation. Animals were sacrified immediately and the levels of dopamine, DOPAC, homovanillic acid, noradrenaline, serotonin and 5-hydroxyindoleacetic acid determined by HPLC in the striatum, cortex, hippocampus and diencephalon/midbrain. Lisuride itself had no effect on any compound determined in any region. In the carotid-ligated gerbil brain, however, lisuride corrected the reduction of dopamine in the striatum, normalized or reduced increases in the (DOPAC+homovanillic acid)/dopamine ratio in the striatum, hippocampus and diencephalon/midbrain, and increased the levels of serotonin in all four regions. The present study, together with previous reports, indicate that lisuride may interfere with ischemia-induced cerebrovascular disturbances and, in such a way, improve some pathological sequelae of cerebrovascular disease.

The dopamine system in weaver mutant mice, a model of cerebellar atrophy, was studied. Dopamine levels of 6-week-old weaver mice were 37%, 44%, 34%, and 41% of levels in age-matched controls in the cerebral cortex, hippocampus, septal region and striatum, respectively. Noradrenaline levels did not differ from controls. This study shows that weaver mice have specific deficiencies in the dopamine system.

In senescent rats, the binding ability of muscarinic cholinergic receptors (MCR) was markedly decreased in the cerebral cortex, hippocampus, thalamus, and the striatum compared with young adult rats. This decrease was markedly improved by chronic administration of bifemelane hydrochloride (bifemelane). In view of the findings that bifemelane markedly inhibits the decrease in acetylcholine (ACh) associated with cerebral ischemia, and that it improves amnesia induced by scopalamine, it may be inferred that bifemelane acts on the pre- and post-synaptic ACh neuronal systems. It may improve memory disturbance due to aging or cerebral ischemia, decreased conscious level, and decreased motor function.

Regional responses of rat brain opioid receptors (Op-Rs) upon castration and testosterone replacement were studied using [3H]naloxone (NAL) binding for u-type and [3H]D-Ala2-methionine-enkephalin (ENK) binding for delta-type Op-R. Castration itself produced an increase in specific NAL and ENK binding in the olfactory bulb and thalamus + midbrain. In the hypothalamus, specific ENK binding was increased without any change in NAL binding. After the testosterone replacement in castrated rats, specific NAL binding was decreased and returned to the control level in the thalamus + midbrain, but remained high in the olfactory bulb. Specific ENK binding was decreased and returned to the control level in the olfactory bulb, hypothalamus and thalamus + midbrain. These results indicated that there are regional differences between u- and delta-type Op-R responses to castration and testosterone replacement. Op-R subtypes were regulated differentially by endogenous hormonal changes, such as testosterone.

Changes in various neurotransmitter systems of the gerbil brain during ischemia and the effects of administration of mergocryptine, a new ergot derivative, were studied. Gerbils intraperitoneally administered 0.5 mg/kg of mergocryptine or the vehicle once a day for 14 days were used. The administration of mergocryptine to control animals had no effect on the contents of neurotransmitters or their metabolites in various regions of the brain, but it corrected abnormalities in the neurotransmitter systems caused by ischemia. These results suggest the usefulness of mergocryptine against the deleterious effects of brain ischemia.

Flunarizine hydrochloride (FZ) is used to improve cerebral circulation and possesses Ca antagonistic effects. In recent years, this drug has been reported to induce parkinsonism and depressive symptoms as side effects, particularly in the elderly. Effects of FZ on dopamine receptors of the rat striatum were studied by radiolabeled receptor assay to clarify the mechanism of onset of parkinsonism in response to FZ. FZ was found to directly and competitively affect D-2 receptors without affecting D-1 receptors. Furthermore, the effect of FZ on D-2 receptors was found to be antagonistic based on the finding that the displacement curve for FZ in the binding of [3H]spiperone to D-2 receptors remained unchanged even after the addition of GppNHp. The effect of FZ on the D-2 receptors in aged rats was more marked than that in young-adult rats. In addition, the tertiary structures of FZ and the anti-schizophrenic agents, pimozide and haloperidol, were examined using computer graphics. FZ was found to have a tertiary structure highly analogous to pimozide and haloperidol, and FZ also had an alkyl structure linking a fluorophenyl group and a nitrogen atom, believed to be particularly necessary for the binding of anti-schizophrenic agents to D-2 receptors. These results may contribute to clarifying the mechanism of onset of parkinsonism in response to FZ, especially in the elderly.