Effects of tyrosine administration on serum biopterin in normal controls and patients with Parkinson's disease.

After administration of tyrosine, total concentration of biopterin, the cofactor for tyrosine hydroxylase, was increased in the striatum, adrenal glands, and serum of rats, and in the serum of humans. Serum biopterin is lower in patients with Parkinson's disease than in normal controls. After oral administration of tyrosine, the increase in serum biopterin concentration was smaller in patients with Parkinson's disease (less than twofold) than in healthy controls (three-to sevenfold). These results suggest that tyrosine may have a regulatory role in biopterin biosynthesis and that patients with Parkinson's disease may have some abnormality in the regulation of biopterin biosynthesis.

Pergolide in the treatment of Parkinson's disease.

Three trials evaluated the efficacy and safety of pergolide. Eighty-six de novo patients and 314 patients already receiving levodopa were enrolled in an open-label study. Of the de novo patients, 47.5% showed a marked or moderate improvement and 32% showed a mild improvement. In the levodopa add-on group, 53.8% showed marked or moderate improvement and 36.3% showed mild improvement. In a short-term, double-blind study, the efficacy of pergolide was compared with that of bromocriptine. One hundred seventy-two patients were randomized to receive pergolide, and 173 were randomized to receive bromocriptine. In de novo patients, bromocriptine (n = 49) and pergolide (n = 49) demonstrated similar efficacy. However, significantly more levodopa-treated patients in the pergolide group, compared with the bromocriptine group, demonstrated marked or moderate improvements in several items of the rating scale score. In a long-term study, 151 of 314 patients receiving pergolide in combination with levodopa remained in the study for 3 years, and 127 for 4 years, and in these patients the initial improvement was maintained. In 18 of 62 de novo patients, the initial improvement was maintained for up to 3 years. These trials indicate that pergolide has efficacy in patients with Parkinson's disease, either as monotherapy or in combination with levodopa.

New pathologic observations in juvenile onset parkinsonism with dystonia.

A patient with hereditary juvenile onset parkinsonism with dystonia died at age 39. There were Lewy bodies and regionally selective neuronal damage in the substantia nigra pars compacta. These changes closely resemble those seen in Parkinson's disease, and emphasize the selective vulnerability of the ventral tier of the pars compacta in these degenerations.

Treatment of parkinsonism with L-threo-3,4-dihydroxyphenylserine: a pharmacokinetic study.

We have examined the kinetics of oral L-threo-3,4-dihydroxyphenylserine (DOPS) alone and combined with peripheral decarboxylase inhibitors in patients with Parkinson's disease and other degenerative diseases of the brain. Combined administration of L-threo-DOPS and carbidopa or benserazide produced higher plasma concentrations of L-threo-DOPS and suppressed the increase in plasma norepinephrine. This finding indicates some advantages of combined therapy with L-threo-DOPS and decarboxylase inhibitors. Measurable quantities of DL-threo-DOPS were found in the CSF during repeated administration, but there was no consistent change in norepinephrine concentration.

Kinetics of superoxide formation by respiratory chain NADH- dehydrogenase of bovine heart mitochondria.

Formation of superoxide anions (O2-) by bovine heart NADH-dehydrogenase preparation (Complex I) supported by an NADH- or NADPH-generating system was studied kinetically. Both NADH- and NADPH-dependent superoxide-forming activities of Complex I were biphasic in double reciprocal plots. The NADH-dependent reaction had two sets of kinetic parameters: One Km value for NADH was 10 times higher than the other one, and the Vmax of the reaction with high Km was about 4 times higher than that of the reaction with low Km. Similar Vmax values were obtained for the NADPH-dependent reactions but the Km values were a thousand times higher than those of the NADH-dependent reactions. The plots of the NADH-dependent activity of rotenone-treated submitochondrial particles were also biphasic. The double reciprocal plots of NADH- and NADPH-dependent 2,6-dichlorophenolindophenol (DCIP) reductase activities of Complex I were linear and the plots of the superoxide-forming activities against the DCIP reductase activities were biphasic. These results indicate that the biphasic double reciprocal plots of the superoxide-forming activities against reduced coenzymes are not caused by interaction between reduced coenzymes and the NADH-dehydrogenase but are due to the presence of at least two superoxide-forming sites in the respiratory chain NADH-dehydrogenase.

Sandwich enzyme immunoassay of dopamine-?-hydroxylase in cerebrospinal fluid from control and parkinsonian patients.

A sandwich enzyme immunoassay (EIA) was established by using purified human serum dopamine-?-hydroxylase (DBH) as a standard protein and a monospecific polyclonal antibody raised against human DBH purified from human pheochromocytoma. The EIA was applied to measuring DBH levels in human CSF from Parkinsonian patients and control patients devoid of neurological diseases. The control group had DBH content of 21.1 +/- 3.1 ng/ml CSF and DBH activity of 24.0 +/- 3.7 ? U/ml CSF, and Parkinsonian group 3.3 ? 0.7 ng/ml CSF (16% of control) and 4.6 +/- 0.7 ?U/ml CSF (19% of control) (mean +/- SEM). Thus, both DBH content and DBH activity in CSF were reduced in Parkinsonian patients to less than 20% of the control values (P $ ? 0.005 ). However, the specific activity (units of enzyme activity/mg of DBH protein) in CSF of Parkinsonian patients was similar to that of control patients. These results suggest that the reduced DBH activity in CSF from Parkinsonian patients is caused by a reduction in DBH protein content, and is not due to production of an inactive form of DBH, for example, by combining with endogenous inhibitor(s). These data support our previous findings that DBH activities in the Parkinsonian brain and CSF are decreased.

Difference in recovery patterns of striatal dopamine content, tyrosine hydroxylase activity and total biopterin content after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration: a comparison of young and older mice.

Striatal dopamine (DA) content, tyrosine hydroxylase (TH) activity, and total biopterin content were measured as parameters of recovery, after administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in 7-week-old (young) and 28-week-old (older) C57 BL/6 mice. After 10 consecutive days of injection of MPTP (30 mg/kg/day) young mice were sacrificed at one day and 2, 4, 8, 12 and 20 weeks; older mice (20 mg/kg/day) at one day and 4, 8, and 12 weeks. All 3 parameters were markedly reduced one day after the last injection of MPTP. During the observation period, the parameters showed a gradual and partial recovery. The recovery rates of the 3 parameters differed significantly, especially during the early phases of 2-8 weeks. Total biopterin content showed a greater rate of recovery than TH activity and TH activity, a greater rate of recovery than DA content. In the older mice group, the recovery of all 3 parameters was retarded significantly, and dissociation of the recovery rates between the 3 parameters was more prominent. The results of our present study suggest that, following neurotoxic injury, the recovery of biopterin may play a significant role in dopaminergic terminal regeneration.

Hemiparkinsonism in monkeys after unilateral caudate nucleus infusion of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP): behavior and histology.

Systematically administered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is biotransformed into 1-methyl-4-phenylpyridinium ion (MPP+), which enters dopaminergic neurons via the dopamine uptake system to destroy nigral cells. Either MPP+ is retrogradely transported to the cell body after being taken up at the nerve terminals, or the dopamine uptake sites on the cell body and its dendritic processes are responsible for the toxin directly entering the neuron. Using a 200 microliter osmotic minipump, we administered 4 mg of MPTP HCl directly into the unilateral caudate nucleus, i.e., the dopamine nerve terminal area, of monkeys for 14 days. Persistent hemiparkinsonism began to appear in a week. Each monkey exhibited a flexed posture and hypokinesia of the contralateral limbs and circling toward the MPTP-treated side. These disturbances developed within 3 months and maintained a plateau for 3 months until the day of sacrifice. After treatment with apomorphine, there appeared a striking circling away from the MPTP-treated side. Selective cell loss in the MPTP-treated side of the substantia nigra pars compacta was found along the entire rostrocaudal extent relative to the untreated side. In conclusion, MPP+ uptake only at the dopamine nerve terminals and retrograde axonal transport to the cell body seemed sufficient to destroy nigral dopamine cells in the monkey.

Histochemical study of the muscle spindles in parkinsonism, motor neuron disease and myasthenia. An examination of the pathological fusimotor endings by the acetylcholinesterase technic.

Pathological changes of the fusimotor endings in parkinsonism, motor neuron disease and myasthenia were examined by the acetylcholinesterase technic on serial sections. In parkinsonism, the diffuse endings, which are thought to be supplied by the static gamma nerve fibers, showed remarkable enlargement, while en plaque and en grappe endings were atrophic. In motor neuron disease, en plaque and en grappe endings, which are thought to be innervated by the beta nerve fibers and dynamic gamma nerve fibers respectively, revealed marked atrophy. However the diffuse endings were normal. In myasthenia gravis and myasthenic syndrome (Eaton-Lambert syndrome), en plaque and en grappe endings were atrophic, though only the diffuse endings were spared. The significance of these changes in the fusimotor endings is discussed.

Histochemical study of normal human muscle spindle. Histochemical classification of intrafusal muscle fibers and intrafusal nerve endings.

Normal muscle spindles of human skeletal muscle were studied histochemically. 1) Four histochemical types of intrafusal muscle fibers were classified by ATPase stain: Bag I fiber, Bag II fiber, Chain I fiber and Chain II fiber. Moreover, two types of nuclear bag fibers were classified by NADH Tetrazolium Reductase stain and PAS stain: Bag I fiber and Bag II fiber. 2) Three kinds of fusimotor endings were verified by the cholinesterase technic: en plaque, en grappe and diffuse endings. 3) Two kinds of fusisensory endings were verified by NADH TR stain and also electron-microscopically: primary and secondary sensory endings.

Destruction of norepinephrine terminals in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice reduces locomotor activity induced by L-dopa.

There is little information concerning the effects of norepinephrine (NE) depletion on clinical features of patients with Parkinson's disease. By inducing two types of experimental parkinsonism, one with a dopamine (DA) deficiency alone and the other with both a DA and NE deficiency, we attempted to evaluate the differences in locomotor activity and behavioral responses between the two groups after L-DOPA administration. The results of the study revealed that increases in locomotor activity were markedly suppressed in the DA and NE deficient group. This may suggest that with striatal DA deficiency the central NE terminals play a significant role in the increase in locomotor activity after L-DOPA administration.

Transforming growth factor-beta 1 levels are elevated in the striatum and in ventricular cerebrospinal fluid in Parkinson's disease.

Transforming growth factor (TGF)-beta 1 content was measured for the first time in the brain (caudate nucleus, putamen, and cerebral cortex) and in ventricular cerebrospinal fluid (VCSF) from control and parkinsonian patients by a sandwich enzyme immunoassay. The concentrations of TGF-beta 1 were significantly higher in the dopaminergic striatal regions in parkinsonian patients than those in controls, but were not significantly different in the cerebral cortex between parkinsonian and control patients. Furthermore, the concentrations of TGF-beta 1 in VCSF were significantly higher in parkinsonian patients than those in non-parkinsonian control patients. Since TGF-beta 1 has potent regulatory activity on cell growth, these results suggest that TGF-beta 1 may have some significant modulatory role in the process of neurodegeneration in Parkinson's disease.

Tumor necrosis factor-alpha (TNF-alpha) increases both in the brain and in the cerebrospinal fluid from parkinsonian patients.

Tumor necrosis factor-alpha (TNF-alpha), a glial-cell-related factor, was measured for the first time in the brain (striatum) and cerebrospinal fluid (CSF) from control and parkinsonian patients by a sensitive sandwich enzyme immunoassay. The concentrations of TNF-alpha in the brain and CSF were significantly higher in parkinsonian patients than those in controls. Since TNF-alpha is an important signal transducer of the immune system with cytotoxic and stimulator properties, these results suggest that an immune response may occur in the nigrostriatal dopaminergic regions in Parkinson's disease and that TNF-alpha may be related, at least in part, to the neuronal degeneration.

Interleukin (IL)-1 beta, IL-2, IL-4, IL-6 and transforming growth factor-alpha levels are elevated in ventricular cerebrospinal fluid in juvenile parkinsonism and Parkinson's disease.

Interleukin (IL)-1 beta , IL-2, IL-4, IL-6, epidermal growth factor (EGF), and transforming growth factor (TGF)-alpha were measured for the first time in ventricular cerebrospinal fluid (VCSF) from control non-parkinsonian patients, patients with juvenile parkinsonism (JP) and patients with Parkinson's disease (PD) by highly sensitive sandwich enzyme immunoassays. All cytokines were detectable in VCSF from control and parkinsonian patients, and the concentrations were much higher than those in lumbar CFS. The concentrations of IL-1 beta, IL-2, IL-4 and TGF-alpha in VCSF were higher in JP than those in controls (P < 0.05). In contrast, the concentrations of IL-2 and IL-6 in VCSF from patients with PD were higher than those from control patients (P < 0.05). These results agree with our previous reports, in which the cytokine levels were elevated in the striatal dopaminergic region of the brain from patients with PD. Since VCSF is produced in the ventricles, the alteration of cytokines in VCSF may reflect the changes of cytokines in the brain. Because cytokines play an important role as mitogens and neurotrophic factors in the brain, the increases in cytokines as a compensatory response may occur in the brain of patients of JP or PD during the progress of neurodegeneration. Increase in cytokines may contribute not only as a compensatory response but as a primary initiating trigger for the neurodegeneration.

bcl-2 protein is increased in the brain from parkinsonian patients.

The proto-oncogene bcl-2 is involved in the regulation of cell death and may be able to block apoptosis in neurons through reduced generation of reactive oxygen species (ROS). The bcl-2 product was measured for the first time in brain (caudate nucleus, putamen and cerebral cortex), ventricular cerebrospinal fluid (VCSF), and lumber CSF (LCSF) from control and parkinsonian patients by highly sensitive two-site sandwich enzyme-linked immunosorbent assay (ELISA). The concentrations of bcl-2 in the nigrostriatal dopaminergic regions were significantly higher in parkinsonian patients than those in controls, whereas this product in cerebral cortex showed no significant difference between parkinsonian and control subjects. Neither VCSF nor LCSF from control or parkinsonian subjects contained the bcl-2 product in the detectable amount (< 5 U/ml). Since oxidative stress may be involved in neurogenerative disorders, accumulation of bcl-2 may reflect a mechanism for counterbalancing ROS-mediated damage, or it might represent the impairment of bcl-2-dependent protection from ROS in parkinsonian brain.

The soluble form of Fas molecule is elevated in parkinsonian brain tissues.

Fas is an apoptosis-signaling receptor molecule on the surface of a number of cell types. The soluble form of Fas (sFas) was measured for the first time in brain (caudate nucleus, putamen and cerebral cortex), ventricular cerebrospinal fluid (VCSF), and lumbar CSF (LCSF) from control and parkinsonian patients by a highly sensitive two-site sandwich enzyme-linked immunosorbent assay (ELISA). The concentrations of sFas in nigro-striatal dopaminergic regions were significantly higher in parkinsonian patients than those in controls, whereas this product in cerebral cortex showed no significant difference between parkinsonian and control subjects. Neither VCSF nor LCSF contained the sFas molecule in the detectable amounts (< 16 pg/ml). These results suggest that the presence of sFas possibly leads to cell death/neurodegeneration in parkinsonian brain.

Beta 2-microglobulin decrease in cerebrospinal fluid from parkinsonian patients.

A sandwich enzyme immunoassay (EIA) was established by using purified beta 2-microglobulin (beta 2-MG) as a standard protein and a polyclonal antibody raised against human beta 2-MG. The EIA was applied for the measurement of beta 2-MG levels in human cerebrospinal fluid (CSF) from parkinsonian patients and control patients devoid of neurological diseases. beta 2-MG contents in CSF of the control group and the parkinsonian group were 1.81 +/- 0.11 micrograms/ml CSF and 0.63 +/- 0.09 microgram/ml CSF, respectively. Thus, beta 2-MG content in CSF was reduced in parkinsonian patients to less than 35% of the control value (P less than 0.005). We had previously reported that the activity and content of dopamine beta-hydroxylase (DBH) were decreased in CSF from parkinsonian patients. A significant positive correlation (r = 0.87) was observed between the beta 2-MG content and DBH activity for CSF from 45 patients. These results suggest a probable link between an immunological change and the changes in catecholaminergic neurons in Parkinson's 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.

Total biopterin levels in the ventricular CSF of patients with Parkinson's disease: a comparison between akineto-rigid and tremor types.

Total L-erythro-biopterin (T-BP) levels in the ventricular cerebrospinal fluid (CSF) were measured in 43 patients with Parkinson's disease (PD) and 12 age-matched neurological controls. In 5 of the PD patients and 1 control, lumbar CSF T-BP values were also measured. The mean ventricular CSF T-BP level in the PD patients, 15.6 +/- 0.5 pmol/ml (mean +/- SE), was significantly lower than that in the controls (21.3 +/- 1.4 pmol/ml, P less than 0.0001). The mean T-BP concentration in the ventricular CSF was 1.9 times higher than that in the lumbar CSF (P less than 0.0005), indicating a rostrocaudal gradient for the T-BP value in the CSF. When the PD patients were classified according to their predominant clinical features into 24 akineto-rigid (A-R) type and 19 tremor (T) type, there was a significant negative correlation between the T-BP levels and duration of illness only for the A-R type patients (rho = -0.605, P less than 0.005). No such significant correlation was found in the T type patients. These results may indicate a difference of pathophysiological changes in the brain between the 2 types of PD.

Juvenile parkinsonism: ventricular CSF biopterin levels and clinical features.

Total biopterin (T-BP) levels in the ventricular cerebrospinal fluid (CSF) and clinical features of 19 patients with juvenile parkinsonism (JP: Parkinson's disease manifesting below the age of 40) were evaluated and compared with 61 patients with classical Parkinson's disease (classical PD: symptoms developing at the age of 40 or above). The JP patients were divided into two subgroups: JP-I; those with good response to levodopa followed by marked motor fluctuations and dopa-induced dyskinesias (DID), JP-II; those with milder response than JP-I with less fluctuations and DID being more similar to classical PD. Both of the mean ventricular CSF T-BP concentrations in the JP and classical PD patients were significantly lower than that in neurological controls. Moreover, the mean T-BP level in the JP-I was markedly lower than that in the JP-II or classical PD. Total biopterin levels revealed a gaussian distribution in the classical PD. However, a bimodal distribution was noted in the JP, with the lower peak consisting of only JP-I patients. These results seem to indicate that JP-II represents early-onset classical PD, while JP-I represents a distinct subgroup having a different physiopathology from classical PD.