Hiroyasu Kinemuchi

Release of dopamine by perfusion with 1-methyl-4-phenylpyridinium ion (MPP(+)) into the striatum is associated with hydroxyl free radical generation.

In Parkinsons disease (PD), the dopamine (DA) neuronal cell death in the nigrostriatal system has been proposed to be mediated by reactive oxygen radicals such as hydroxyl radicals (.OH). This.OH production may cause lipid peroxidation of cell membranes leading to neuronal cell death. This paper report that the DA-selective neurotoxin, 1-methyl-4-phenylpyridinium ion (MPP(+)), (1 nmol/microl per min for 1 h) infusion into the striatum of rats induces elevation of extracellular DA and.OH formation. These elevations seem to induce lipid peroxidation of striatum membranes, as detected by increases in non-enzymatic formation of 2,3-dihydroxybenzoic acid (DHBA) levels. To test the involvement of DA release in the.OH generation and lipid peroxidation, the rats were pretreated with reserpine (5 mg/kg, i.v., 24 h before MPP(+) or without MPP(+)) to deplete presynaptic DA. Reserpine treatment alone did not change the levels of DA or 2,3-DHBA, while the combined treatment with both MPP(+) and reserpine clearly decreased 2,3-DHBA, as well as DA levels, compared to those in the group treated with MPP(+) alone. After injection into reserpinized rats, DA at various doses (2, 5 and 10 microM) small increased 2,3-DHBA levels dose-dependently, as compared to the MPP(+) alone-treated group. These results clearly indicate that MPP(+) perfusion into the striatum increases extracellular DA levels and this increase may concomitantly induce the formation of reactive free oxygen radicals, such as.OH free radicals. These events may contribute, at least in part, to the nigrostriatal neurons cell death after MPP(+).

Selective Inhibitors of Membrane-Bound Semicarbazide-Sensitive Amine Oxidase (SSAO) Activity in Mammalian Tissues

Semicarbazide-sensitive amine oxidase (SSAO, EC 1.4.3.6) is a group of enzymes highly sensitive to inhibition by semicarbazide. This high sensitivity distinguishes these enzymes from monoamine oxidase (MAO). Various mammalian tissues contain membrane-bound SSAO which metabolizes only the primary monoamines. Vascular and non-vascular smooth muscle cells have particularly high SSAO activity, but recently the enzyme activity has also been found in non-vascular smooth muscle cells. The substrate specificity of SSAO shows considerable species-related variations. A variety of compounds inhibiting MAO activity has also been identified as SSAO inhibitors. Among inhibitors, there is no specific SSAO inhibitor so far tested. Many studies reinforce the conclusion that inhibitory properties of some compounds against MAO activities has been markedly differed from their properties as SSAO inhibitors. 2-bromoethylamine has been recently developed with a potent, selective and suicide SSAO inhibitor without any inhibitory effect on MAO activity Using this inhibitor, it is possible to study the role of the enzyme in mammalian tissues. As physiological role the increased concentrations of SSAO, especially in blood plasma, have been found in diabetic patients and experimental animals. This enzyme was found to be associated with translocation of the glucose transporter GLUT 4 into the adipose cell surface and involved in the signaling of glucose uptake. Recent studies showed that vascular SSAO metabolizes endogenous primary amines, allylamine, methylamine and aminoacetone, to the corresponding cytotoxic aldehydes. These aldehydes have been linked to the ability of diabetic complications such as neuropathy, retinopathy and nephropathy. Overproduction of such toxic aldehydes produced by increased SSAO activity was proposed to be potentially hazardous in diabetic complications. Thus, reduction or inhibition of SSAO may be beneficial in these pathological conditions. Clearly species-related differences in properties of SSAO must be taken into account in this respect, particularly when assessing if SSAO inhibition may have great application in human.

A possible mechanism of antidepressant activity of beta-amyrin palmitate isolated from Lobelia inflata leaves in the forced swimming test

A mechanism of antidepressant activity of beta-amyrin palmitate was studied using the forced swimming method in mice. Beta-amyrin palmitate (10 mg/kg) reduced the increase in the duration of immobility induced by tetrabenazine (100 and 200 mg/kg), but showed no effect on that in mice treated with alpha-methyl-para-tyrosine (500 mg/kg). Beta-amyrin palmitate (5 and 10 mg/kg) decreased the duration of immobility in mice treated with desipramine plus 6-hydroxy-dopamine (50 micrograms/mouse), but did not affect that induced by nomifensine plus 6-hydroxydopamine. The decreased immobility produced by desipramine (15 mg/kg) was not affected by beta-amyrin palmitate. A study of norepinephrine release in mouse brain synaptosomes indicated that beta-amyrin palmitate caused a release of [3H]norepinephrine. The results of the present study suggest that beta-amyrin palmitate might release norepinephrine from newly synthesized pools, and thus, it might activate noradrenergic activity.

Nicotine suppresses 1-methyl-4-phenylpyridinium ion-induced hydroxyl radical generation in rat striatum.

The present study examined the ability of antioxidant effects of nicotine on 1-methyl-4-phenylpyridinium ion (MPP(+))-induced hydroxyl radical (*OH) formation of rat striatum. Rats were anesthetized and sodium salicylate in Ringers solution (0.5 nmol/microl per min) was infused through a microdialysis probe to detect the generation of.OH as reflected by non-enzymatic formation trapped as 2,3-dihydroxybenzoic acid (DHBA) in the striatum. MPP(+) enhanced generation of.OH formation trapped as DHBA. However, nicotine (100 microM) significantly suppressed.OH formation induced by MPP(+). Iron (II) (2, 5 and 10 microM) increased the levels of DHBA in a concentration-dependent manner. However, in the presence of nicotine (100 microM), the effect of nicotine was suppressed. These results suggest that nicotine has a preventive effect on.OH generation by MPP(+) in rat striatum.

Inhibition of rat brain monoamine oxidase by some analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and 1-methyl-4-phenylpyridinium ion

To clarify the essential chemical structures of the neurotoxins, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its oxidized product, 1-methyl-4-phenylpyridinium ion (MPP+), that govern nigrostriatal dopamine neuron toxicity, interactions of several structurally related compounds of MPTP or MPP+ with monoamine oxidase (MAO) in rat forebrain homogenates were studied. Of the compounds tested, 4-phenyl-1,2,3,6-tetrahydropyridine (PTP), 4-phenylpyridine and 4-phenylpiperidine strongly and dose-dependently inhibited MAO-A and -B activity. Inhibition of PTP and 4-phenylpiperidine was MAO-A-selective, while that by 4-phenylpyridine was MAO-B-selective. Of these 3 compounds, only PTP time-dependently inhibited MAO-B, but not -A. Without preincubation, the modes of inhibition of MAO-A and -B by PTP were competitive. After 1 h preincubation, the mode of MAO-B inhibition changed to non-competitive, while inhibition of -A remained unchanged. PTP was oxidized by MAO-B, but not by -A, under these conditions. In contrast, 4-phenylpyridine and 4-phenylpiperidine were not substrates for either form of MAO in rat forebrain homogenates. These results, along with the other observations, indicate that PTP may essentially cause a neurotoxic effect on the nigrostriatal dopamine pathway.

Protective effect of diltiazem, a L-type calcium channel antagonist, on bisphenol A-enhanced hydroxyl radical generation by 1-methyl-4-phenylpyridinium ion in rat striatum

The present study examined whether bisphenol A enhanced 1-methyl-4-phenylpyridinium ion (MPP+)-induced hydroxyl radical (*OH) generation in rat striatum using microdialysis technique. These elevations were detected by increases in non-enzymatic formation of 2,3-dihydroxybenzoic acid levels. Bisphenol A significantly enhanced MPP+-induced.OH generation. Further, the effect of diltiazem, a L-type Ca2+ channel antagonist, on bisphenol A (10 microM) and MPP+ (5 mM)-induced.OH generation were studied. Ddiltiazem (100 microM) significantly suppressed bisphenol A and MPP+-induced.OH generation. The results indicate bisphenol A augmented *OH generation, and diltiazem may have preventive effect on bisphenol A and MPP+-induced.OH generation.

Antinociceptive effect following dietary-induced thiamine deficiency in mice: Involvement of substance P and somatostatin

We produced thiamine deficiency by treating mice with a thiamine deficient (TD) diet, but not with pyrithiamine, a thiamine antagonist. Twenty days after TD feeding, a significant antinociceptive effect was observed in the formalin test. A single injection of thiamine HCl (50 mg/kg, s.c.) on the 19th day after TD feeding (on the late TD stage) failed to reverse the antinociceptive effect, the muricide effect, and impairment of avoidance learning induced by TD feeding, as compared to pair-fed controls. These results indicate the possibility that the TD-induced antinociceptive effect may result from irreversible changes in the spinal and/or brain neurons. To clarify the involvement of substance P (SP) and somatostatin (SST) systems in the spinal cord, we examined the effect of intrathecal (i.t.) injections of these agonists on TD feeding-inducd elevation of pain threshold. I.t. injection of SP and SST elicited a behavioral response consisting of reciprocal hindlimb scratching, biting and/or licking of hindpaws. There was no significant difference in the behavioral response to SP between TD mice and PF mice on the 5th day after feeding. However, on the 10th and 20th day after TD feeding the response to SP was significantly increased compared with PF mice. This phenomenon was also observed with SST on the 20th day after TD feeding. These results indicate the possibility that TD feeding may produce an increased behavioral response to SP and SST through an enhanced sensitivity of neurokinin-1 and SST receptors in the spinal cord. Taken together, the antinociceptive effect following TD feeding may result from a decrease in spinal SP and SST contents.

Molecular characteristics of a single and novel form of carp (Cyprinus carpio) monoamine oxidase

Two mammalian monoamine oxidases (MAO), MAO-A and MAO-B, are similar in primary structures but have unique substrate/inhibitor selectivities. Carp (Cyprinus carpio) contains a MAO enzyme (C-MAO) with properties different from MAO-A and MAO-B. To determine the molecular characteristics of C-MAO and its phylogenetic relationship with other fish and mammalian MAOs, the primary structure of C-MAO was estimated. The putative C-MAO cDNA encodes 526 amino acids with 59.001 Da, and the deduced amino acid sequence showed as much as 68.9% homology with some mammalian MAO-A proteins, 69.8% homology with some mammalian MAO-B proteins, and as much as 92.4% homology with some fish MAOs. Comparison of two regions in the polypeptide sequence of C-MAO determining possible substrate/inhibitor preferences of MAO-A and MAO-B showed both 79.5% homologies.

Nucleotide sequences of putative cDNAs for guinea-pig monoamine oxidase.

To study the molecular structure of guinea pig monoamine oxidase (MAO) and its phylogenetic relationship with other mammalian MAOs, we determined nucleotide sequences of putative MAO cDNAs isolated from guinea pig tissues. Both the 5′- and 3′-ends of the cDNAs were amplified using the RACE (rapid amplification of cDNA ends) method. The sequence (1924 bp) of a putative guinea-pig MAO-B cDNA covers a complete coding region that corresponds to 521 amino acids. We also analyzed a partial sequence of a putative guinea-pig MAO-A cDNA, which corresponds to 506 amino acids, but have left the region of 66 bp at the 3′-end undetermined. The nucleotide and deduced amino-acid sequences of the putative guinea-pig MAO cDNAs showed the highest homology with that of human MAO cDNAs among the known mammalian MAO sequences. These results suggest that guinea-pig MAOs are structurally similar to human MAOs. Our molecular phylogenetic data support the idea that guinea pigs and rodents diverged before the separation between rodents and other lineage leading to Primates and Artiodactyla.

In vivo generation of hydroxyl radicals and MPTP-induced dopaminergic neurotoxicity in the striatum

Free radical production might make a major contribution at certain stages in the progression of injury to the brain. Oxygen free radical formation has been implicated in lesions caused by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and iron. Although MPTP produces a parkinsonian syndrome after its conversion to 1-methyl-4-phenylpyridine (MPP+) by type B monoamine oxidase (MAO-B) in the brain, the etiology of this disease remains obscure. MPP+ is a highly potent dopamine (DA)-releasing agent and DA autoxidation catalyzed by iron and oxidative stress may be involved in the pathogenesis of Parkinsons disease. Histidine, a singlet oxygen (1O2) scavenger, protects MPP+-induced hydroxyl radical (•OH) formation. The inhibitory effect on the susceptibility of LDL oxidation can reduce •OH generation. These drugs may be applied as antiparkisonian agents. Further clinical investigation is necessary in future. This finding may be useful in elucidating the actual mechanism of free radical formation in the pathogenesis of neurodegenerative brain disorders, including Parkinsons disease and traumatic brain injuries.