Wakako Maruyama

N-methylation of dopamine-derived 6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, (R)-salsolinol, in rat brains : in vivo microdialysis study

Abstract: N‐Methylation of (R)‐1‐methyl‐6,7‐dihydroxy‐1,2,3,4‐tetrahydroisoquinoline [(R)‐salsolinol] derived from dopamine was proved by in vivo microdialysis study in the rat brain. The striatum was perfused with (R)‐salsolinol and N‐methylated compound was identified in the dialysate using HPLC and electrochemical detection with multichanneled electrodes. N‐Methylation of (R)‐salsolinol was confirmed in three other regions of the brain, the substantia nigra, hypothalamus, and hippocampus. In the substantia nigra, the amount of N‐methylated (R)‐salsolinol was significantly larger than in the other three regions. These results indicate that around dopaminergic neurons, particularly in the substantia nigra, (R)‐salsolinol was methylated into N‐methyl‐(R)‐salsolinol, which has a chemical structure similar to that of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine, the selective dopaminergic neurotoxin. N‐Methylation of tetrahydroisoquinolines and β‐carbolines have already been proven to increase their toxicity to dopaminergic neurons and N‐methylation might be an essential step for these alkaloids to increase their toxicity. On the other hand, after perfusion of (R)‐salsolinol, release of dopamine and 5‐hydroxytryptamine was observed and inhibition of monoamine oxidase was indicated. (R)‐Salsolinol and its derivatives may be candidates for being dopaminergic neurotoxins.

Dopamine-derived endogenous 1(R),2(N)-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, N-methyl-(R)-salsolinol, induced parkinsonism in rat: biochemical, pathological and behavioral studies

Dopamine-derived 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (salsolinol, Sal) and related compounds were examined for their selective neurotoxicity to dopamine neurons by injection into the rat striatum. Among salsolinol analogs examined, only N-methyl-(R)- salsolinol (NM(R)Sal) induced behavioral changes very similar to those in Parkinsons disease: hypokinesia, stiff tail, limb twitching at rest and postural abnormality. Biochemical analysis showed that after NM(R)Sal injection, NM(R)Sal itself and its oxidation product, 1-2-dimethyl-6,7-dihydroxyisoquinolinium ion (DMDHIQ+) accumulated in the striatum, and also in the substantia nigra definite amount of DMDHIQ+ was detected. Dopamine and noradrenaline were reduced in the striatum and more markedly in the substantia nigra, whereas serotonin and its metabolite were not affected. Morphological analysis revealed selective reduction of tyrosine hydroxylase (TH)-containing neurons in the substantia nigra after continuous NM(R)Sal administration in the striatum. These results demonstrate the selective cytotoxicity of NM(R)Sal to the dopamine neurons in the substantia nigra, and the possible involvement of this 6,7-dihydroxy-isoquinoline in the pathogenesis of Parkinsons disease is discussed.

N-methyl(r)salsolinol produces hydroxyl radicals: Involvement to neurotoxicity

Recently, (R)-1,2-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline [N-methyl-(R)salsolinol, NM(R)Sal] and 1,2-dimethyl-6,7-dihydroxyisoquinolinium ion [DiMeDHIQ+] were found to cause a syndrome similar to parkinsonism in rodents. NM(R)Sal is produced in the brain by N-methylation of a naturally occurring catechol isoquinoline, 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline [(R)salsolinol, (R)Sal], which is formed from dopamine. The mechanism of NM(R)Sal cytotoxicity to dopamine neurons was examined using in vitro experiments. NM(R)Sal was found to be nonenzymatically oxidized into DiMeDHIQ+, with concomitant formation of hydroxyl radicals. The oxidation and the radical production were completely inhibited by the antioxidants, ascorbic acid and reduced glutathione, and the radical formation was enhanced by Fe(II) and, to a less extent, by Fe(III). The oxidation of NM(R)Sal into DiMeDHIQ+ and the production of hydroxyl radicals may be essential for neurotoxicity to develop in dopamine neurons. The possible involvement of this catechol isoquinoline in the pathogenesis of Parkinsons disease is discussed.

Uptake of a neurotoxin-candidate, (R)-1,2-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline into human dopaminergic neuroblastoma SH-SY5Y cells by dopamine transport system

Uptake of catechol isoquinolines to dopamine cells was studied using human dopaminergic neuroblastoma SH-SY5Y cells. Only (R)-1,2-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline [(R)-1,2-DiMeDHTIQ] was transported by dopamine uptake system, while (S)-1,2-DiMeDHTIQ, (R)- and (S)-1-methyl-6,7-dihydroxy-tetrahydroisoquinoline, and 1,2-dimethyl-6,7-dihydroxyisoquinolinum ion were not. Kinetical study showed that the uptake of (R)-1,2-DiMeDHTIQ followed the Michaelis-Menten equation, and the values of the Michaelis constant and the maximal velocity were obtained to be 102.6 ± 36.9 μM and 66.0 ± 2.8 pmol/min/mg protein. Dopamine was found to inhibit (R-1-DiMeDHTIQ uptake competitively. These results suggest that the selective uptake by dopamine transporter may account for the specific neurotoxicity of (R)-1,2-DiMeDHTIQ to dopamine neurons.

Neurotoxins induce apoptosis in dopamine neurons: protection by N-propargylamine-1(R)- and (S)-aminoindan, rasagiline and TV1022

In Parkinsons disease, apoptosis was proposed to cause cell death in nigral dopamine neurons. An endogenous dopaminergic neurotoxin, N-methyl(R)salsolinol, stereo-selectively induced apoptosis in human neuroblastoma SH-SY5Y cells. In this paper the intracellular mechanism of apoptosis was studied using N-methyl(R)salsolinol, 6-hydroxydopamine and peroxynitrite as inducers of apoptosis. Apoptotic cascade was initiated by opening of mitochondrial permeability transition pore, as shown by collapse of mitochondrial membrane potential, deltapsim. Apoptosis was executed by caspase 3 activation, followed by DNA fragmentation, which was antagonized by overexpressed Bcl-2. Propargylamines were found to protect the cells from apoptosis, and rasagiline, a selective irreversible inhibitor of type B monoamine oxidase was the most potent to prevent the cell death. Rasagiline preserved deltapsim, which was proved also in isolated mitochondria, and rasagiline completely suppressed the activation of caspases and DNA fragmentation. These results suggest that mitochondria regulate apoptotic process, which may be a target of neuroprotection by rasagiline.

A dopaminergic neurotoxin, 1(R), 2(N)-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, N-methyl(R)salsolinol, and its oxidation product, 1,2(N)-dimethyl-6,7-dihydroxyisoquinolinium ion, accumulate in the nigro-striatal system of the human brain

N-Methyl(R)salsolinol was found to be an endogenous dopaminergic neurotoxin inducing parkinsonism in rodents and to increase in the cerebrospinal fluid of parkinsonian patients. The amounts of N-methyl(R)salsolinol and related compounds in the human brain regions were quantitatively analyzed. Only the (R)-enantiomer of salsolinol derivatives were detected, which suggests their enzymatic synthesis in situ. In the nigro-striatal system, the concentration of N-methyl(+)salsolinol was higher than in the frontal cortex, and its oxidized catechol isoquinolinium ion was detected only in the substantia nigra significantly. The accumulation of these neurotoxins in the nigro-striatal region might account for selective cell death of dopamine neurons in the substantia nigra of Parkinsons disease.

The metabolism of l-DOPA and l-threo-3,4-dihydroxyphenylserine and their effects on monoamines in the human brain: analysis of the intraventricular fluid from parkinsonian patients

The monoamines and their metabolites were analyzed in the intraventricular fluid of parkinsonian patients treated with L-DOPA alone or together with L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS), the precursor amino acids of dopamine and noradrenaline, respectively. In the intraventricular fluid of the patients administered with L-DOPA, the level of dopamine metabolites were higher than control, suggesting enhanced turnover of dopamine in the brain. However, L-DOPA administration increased free noradrenaline only slightly, and did not affect serotonin and its metabolite. On the other hand, by administration of L-DOPA combined with L-threo-DOPS, the levels of monoamines increased in general, whereas the monoamine metabolites by catechol-O-methyltransferase were reduced compared with those in the patients treated with L-DOPA alone. Only a minor part of L-threo-DOPS was metabolized into noradrenaline by aromatic L-amino acid decarboxylase, and it was metabolized mainly by two other enzymes, catechol-O-methyltransferase and DOPS-aldolase in the brain. An overview of the metabolism of neurotransmitters in the brain proved to be useful to evaluate the therapeutic effects of these precursor amino acids.

Dopamine‐Derived 1‐Methyl‐6,7‐Dihydroxyisoquinolines as Hydroxyl Radical Promoters and Scavengers in the Rat Brain: In Vivo and In Vitro Studies

Abstract: The effects of derivatives of dopamine‐derived isoquinoline, (R)‐1‐methyl‐6,7‐dihydroxy‐1,2,3,4‐tetrahydroisoquinoline [or (R)‐salsolinol] on hydroxyl radical production were studied in vivo and in vitro. As reported previously, (R)‐salsolinol is N‐methylated in the brain into N‐methyl‐(R)‐salsolinol, which is further oxidized into the 1,2‐dimethyl‐6,7‐dihydroxyisoquinolinium ion. Using in vivo microdialysis, we measured hydroxyl radical levels in the rat striatum by HPLC after derivatization to 2,3‐dihydroxybenzoic acid with salicylic acid. (R)‐Salsolinol and the isoquinolinium ion (40 and 200 µM) and N‐methyl‐(R)‐salsolinol (200 µM) reduced in vivo radical formation, with reduction of dopamine catabolism. (R)‐Salsolinol and the isoquinolinium ion reduced in vitro hydroxyl radical production from dopamine autoxidation. On the other hand, 40 µMN‐methyl‐(R)‐salsolinol increased the hydroxyl radical level in the striatum, and the radical production by its autoxidation was confirmed in vitro. N‐Methyl‐(R)‐salsolinol affected neither in vivo dopamine catabolism nor in vitro production of hydroxyl radicals from dopamine. These results show that (R)‐salsolinol and N‐methyl‐(R)‐salsolinol may be neuroprotective and neurotoxic, respectively, and thus might be involved in the pathogenesis of Parkinsons disease.

N-methyl-(R)salsolinol as a dopaminergic neurotoxin: from an animal model to an early marker of Parkinson's disease.

A dopamine-derived 1(R), 2(N)-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydrosioquinoline [N-methyl-(R)salsolinol] was found to occur enantioselectively in human brain. This isoquinoline induced parkinsonism in rat after injection in the striatum, and the behavioral, biochemical and pathological changes were very similar to those in Parkinsons disease. N-Methyl-(R)salsolinol depleted dopamine neurons in the rat substantia nigra without necrotic tissue reaction, which may be due to the apoptotic death process, as proved by its induction of DNA damage in dopaminergic neuroblastoma SH-SY5Y cells. N-Methyl-(R)salsolinol was found to increase significantly in the cerebrospinal fluid of parkinsonian patients. All these results suggest that N-methyl-(R)salsolinol may be an endogenous neurotoxin to cause Parkinsons disease and the enzymes involved in its biosynthesis and catabolism may be endogenous factors in the pathogenesis of this disease.

Inhibition of Tyrosine Hydroxylase by R and S Enantiomers of Salsolinol, 1-Methyl-6,7-Dihydroxy-1,2,3,4- Tetrahydroisoquinoline

Abstract: Salsolinol is one of the dopamine‐derived tetrahydroisoquinolines and is synthesized from pyruvate or acetaldehyde and dopamine. As it cannot cross the blood‐brain barrier, salsolinol as the R enantiomer in the brain is considered to be synthesized in situ in dopaminergic neurons. Effects of R and S enantiomers of salsolinol on kinetic properties of tyrosine hydroxylase [tyrosine, tetrahydrobiopterin:oxygen oxidoreductase (3‐hydroxylating); EC 1.14.16.2], the rate‐limiting enzyme of catecholamine biosynthesis, were examined. The naturally occurring co‐factor of tyrosine hydroxylase, l‐erythro‐5,6,7,8‐tetra‐hydrobioptein, was found to induce allostery to the enzyme polymers and to change the affinity to the biopterin itself. Using l‐erythro‐5,6,7,8‐tetrahydrobiopterin, tyrosine hydroxylase recognized the stereochemical structures of the salsolinols differently. The asymmetric center of salsolinol at C‐1 played an important role in changing the affinity to l‐tyrosine. The allostery of tyrosine hydroxylase toward biopterin cofactors disappeared, and at low concentrations of biopterin such as in brain tissue, the affinity to the cofactor changed markedly. A new type of inhibition of tyrosine hydroxylase, by depleting the allosteric effect of the endogenous biopterin, was found. It is suggested that under physiological conditions, such a conformational change may alter the regulation of DOPA biosynthesis in the brain.