Normal aging in rats and pathological aging in human Alzheimer's disease decrease FAAH activity: modulation by cannabinoid agonists.

Anandamide is an endocannabinoid involved in several physiological functions including neuroprotection. Anandamide is synthesized on demand and its endogenous level is regulated through its degradation, where fatty acid amide hydrolase plays a major role. The aim of this study was to characterize anandamide breakdown in physiological and pathological aging and its regulation by CB1 and CB2 receptor agonists. Fatty acid amide hydrolase activity was analyzed in an independent cohort of human cortical membrane samples from control and Alzheimer's disease patients, and in membrane and synaptosomes from adult and aged rat cerebral cortex. Our results demonstrate that fatty acid amide hydrolase activity decreases in the frontal cortex from human patients with Alzheimer's disease and this effect is mimicked by Aß(1-40) peptide. This activity increases and decreases in aged rat cerebrocortical membranes and synaptosomes, respectively. Also, while the presence of JWH-133, a CB2 selective agonist, slightly increases anandamide hydrolysis in human controls, it decreases this activity in adults and aged rat cerebrocortical membranes and synaptosomes. In the presence of WIN55,212-2, a mixed CB1/CB2 agonist, anandamide hydrolysis increases in Alzheimer's disease patients but decreases in human controls as well as in adult and aged rat cerebrocortical membranes and synaptosomes. Although a similar profile is observed in fatty acid amide hydrolase activity between aged rat synaptic endings and human Alzheimer's disease brains, it is differently modulated by CB1/CB2 agonists. This modulation leads to a reduced availability of anandamide in Alzheimer's disease and to an increased availability of this endocannabinoid in aging.

Leptin gene therapy attenuates neuronal damages evoked by amyloid-ß and rescues memory deficits in APP/PS1 mice.

There is growing evidence that leptin is able to ameliorate Alzheimer's disease (AD)-like pathologies, including brain amyloid-ß (Aß) burden. In order to improve the therapeutic potential for AD, we generated a lentivirus vector expressing leptin protein in a self-inactivating HIV-1 vector (HIV-leptin), and delivered this by intra-cerebroventricular administration to APP/PS1 transgenic model of AD. Three months after intra-cerebroventricular administration of HIV-leptin, brain Aß accumulation was reduced. By electron microscopy, we found that APP/PS1 mice exhibited deficits in synaptic density, which were partially rescued by HIV-leptin treatment. Synaptic deficits in APP/PS1 mice correlated with an enhancement of caspase-3 expression, and a reduction in synaptophysin levels in synaptosome preparations. Notably, HIV-leptin therapy reverted these dysfunctions. Moreover, leptin modulated neurite outgrowth in primary neuronal cultures, and rescued them from Aß42-induced toxicity. All the above changes suggest that leptin may affect multiple aspects of the synaptic status, and correlate with behavioral improvements. Our data suggest that leptin gene delivery has a therapeutic potential for Aß-targeted treatment of mouse model of AD.

Cortical expression of brain derived neurotrophic factor and type-1 cannabinoid receptor after striatal excitotoxic lesions.

An involvement of one particular neurotrophin, namely, the brain-derived neurotrophic factor (BDNF), has been demonstrated in the pathophysiology Huntington's disease. Type-1 cannabinoid (CB1) receptor has been postulated to upregulate BDNF gene transcription. To better understand the relationship between CB1 and BDNF levels in a situation where the striatum is degenerating, we studied, by dual label immunofluorescence, the distribution of CB1 and BDNF in cortical neurons projecting to the striatum in our rat quinolinic acid model of striatal excitotoxicity. We completed our study with quantitative analyses of BDNF protein levels and CB1 binding activity in the cortex. We show that, 2 weeks post lesion, cortical neurons contain more BDNF compared with controls and to earlier time points. Such BDNF up-regulation coincides with a higher binding activity and an increased protein expression of CB1. We suggest that after excitotoxic lesions, CB1 might, at least transiently, upregulate BDNF in the attempt to rescue striatal neurons from degeneration.

Inhibition of glioma growth in vivo by selective activation of the CB(2) cannabinoid receptor.

The development of new therapeutic strategies is essential for the management of gliomas, one of the most malignant forms of cancer. We have shown previously that the growth of the rat glioma C6 cell line is inhibited by psychoactive cannabinoids (I. Galve-Roperh et al., Nat. Med., 6: 313-319, 2000). These compounds act on the brain and some other organs through the widely expressed CB(1) receptor. By contrast, the other cannabinoid receptor subtype, the CB(2) receptor, shows a much more restricted distribution and is absent from normal brain. Here we show that local administration of the selective CB(2) agonist JWH-133 at 50 microg/day to Rag-2(-/-) mice induced a considerable regression of malignant tumors generated by inoculation of C6 glioma cells. The selective involvement of the CB(2) receptor in this action was evidenced by: (a) the prevention by the CB(2) antagonist SR144528 but not the CB(1) antagonist SR141716; (b) the down-regulation of the CB(2) receptor but not the CB(1) receptor in the tumors; and (c) the absence of typical CB(1)-mediated psychotropic side effects. Cannabinoid receptor expression was subsequently examined in biopsies from human astrocytomas. A full 70% (26 of 37) of the human astrocytomas analyzed expressed significant levels of cannabinoid receptors. Of interest, the extent of CB(2) receptor expression was directly related with tumor malignancy. In addition, the growth of grade IV human astrocytoma cells in Rag-2(-/-) mice was completely blocked by JWH-133 administration at 50 microg/day. Experiments carried out with C6 glioma cells in culture evidenced the internalization of the CB(2) but not the CB(1) receptor upon JWH-133 challenge and showed that selective activation of the CB(2) receptor signaled apoptosis via enhanced ceramide synthesis de novo. These results support a therapeutic approach for the treatment of malignant gliomas devoid of psychotropic side effects.

beta-Amyloid-induced cytotoxicity, peroxide generation and blockade of glutamate uptake in cultured astrocytes.

beta-Amyloid (betaA) is cytotoxic to neurons in culture by increasing hydrogen peroxide and altering calcium homeostasis. We have evaluated betaA-induced cytotoxicity, peroxide generation and glutamate (Glu) uptake in cultured astrocytes. Twenty-four hours after a single addition of either betaA25-35 or betaA1-40there was a concentration-dependent decrease in viability. Catalase or vitamin E showed no protective effect against betaA25-35 Dithiothreitol (DTT), N-acetylcysteine (NAC) and cyclosporine A significantly prevented the toxic effects of both betaA25-35 and peroxide, while inhibition of peroxide detoxifying enzymes enhanced toxicity. Exposure to betaA25-35 or betaA1-40 increased peroxides at 2 h and 24 h, which was prevented by DTT and NAC, but not vitamin E. betaA25-35 inhibited Glu uptake in astrocytes and neurons in culture. Following exposure of neurons to betaA for 24 h there was decreased uptake and increased Glu levels in the culture medium, that resulted in gradual excitotoxicity.

Increased cannabinoid CB1 receptor binding and activation of GTP-binding proteins in the basal ganglia of patients with Parkinson's syndrome and of MPTP-treated marmosets.

Recent evidence obtained in rat models of Parkinson's disease showed that the density of cannabinoid CB1 receptors and their endogenous ligands increase in basal ganglia. However, no data exists from post-mortem brain of humans affected by Parkinson's disease or from primate models of the disorder. In the present study, we examined CB1 receptor binding and the magnitude of the stimulation by WIN55,212-2, a specific CB1 receptor agonist, of [35S]GTPgammaS binding to membrane fractions from the basal ganglia of patients affected by Parkinson's disease. In Parkinson's disease, WIN55,212-2-stimulated [35S]GTPgammaS binding in the caudate nucleus, putamen, lateral globus pallidus and substantia nigra was increased, thus indicating a more effective activation of GTP-binding protein-coupled signalling mechanisms via CB1 receptors. This was accompanied by an increase in CB1 receptor binding in the caudate nucleus and the putamen, although no changes were observed in the lateral globus pallidus and the substantia nigra. Because Parkinson's disease patients had been chronically treated with l-DOPA, brains were studied from normal common marmosets and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated animals with and without chronic L-DOPA treatment. MPTP-lesioned marmosets had increased CB1 receptor binding in the caudate nucleus and the putamen compared to control marmosets, as well as increased stimulation of [35S]GTPgammaS binding by WIN55,212-2. However, following l-DOPA treatment these parameters returned towards control values. The results indicate that a nigro-striatal lesion is associated with an increase in CB1 receptors in the basal ganglia in humans and nonhuman primates and that this increase could be reversed by chronic l-DOPA therapy. The data suggest that CB1 receptor blockade might be useful as an adjuvant for the treatment of parkinsonian motor symptoms.

beta-amyloid peptides are cytotoxic to astrocytes in culture: a role for oxidative stress.

beta-Amyloid is cytotoxic to neurons in culture by increasing hydrogen peroxide and altering calcium homeostasis. We have evaluated the cytotoxicty of beta-amyloid peptides (betaA(25-35) and betaA(1-40)) and generation of hydrogen peroxide on cortical cultured astrocytes. Twenty-four hours after a single addition of either betaA(25-35) or betaA(1-40) there was a concentration-dependent decrease in viability. This toxicity never exceeded 50% of the population independently of exposure time and concentrations. The subpopulation of astrocytes resistant to betaA(25-35) effects were also insensitive to peroxide. Catalase or vitamin E showed no protective effect against betaA(25-35) toxicity. Dithiothreitol (DTT), N-acetylcysteine (NAC), and cyclosporine A significantly prevented the toxic effects of both betaA(25-35) and peroxide. Inhibition of peroxide detoxifying enzymes increased betaA(25-35) and peroxide toxicity. Exposure to betaA(25-35) or betaA(1-40) increased peroxide production at 2 and 24 h, which was prevented by DTT and NAC, but not vitamin E. Despite the inability of added catalase to reduce betaA toxicity, these results suggest that betaA-induced cytotoxicity to astrocytes in culture is, as in neurons, mediated by generation of hydrogen peroxide.

Repeated intracerebroventricular administration of beta-amyloid(25-35) to rats decreases muscarinic receptors in cerebral cortex.

The effects of repeated in vivo administration to rats of beta-amyloid(25-35) (betaA(25-35)) on several cholinergic markers have been studied and compared with those of a peptide with a scrambled sequence. Rats received intracerebroventricular injections of betaA(25-35) (5 or 20 microg/day) for 7 days and they were sacrificed at 2 or 3 weeks survival. The density of total muscarinic receptors labeled with [3H]N-methyl-scopolamine was dose-dependently decreased by betaA(25-35) in the cerebral cortex at 3 weeks survival. No changes were observed at 2 weeks survival in cerebral cortex or in the hippocampus, at any time. BetaA(25-35) administration did not modify choline acetyltranferase activity in cerebral cortex. However, in betaA(25-35)-treated rats hypertrophic/hyperactive positive acetylcholinesterase nucleus basalis cholinergic neurons were observed at 2 weeks survival, while the density of acetylcholinesterase-positive fibers of cerebral cortex was increased along with the number of cortical positive neurons at 3 weeks survival. These results suggest that increased cholinergic function may be responsible of muscarinic receptor down-regulation. Given the involvement of cholinergic systems in memory and learning, repeated administration of betaA(25-35) may represent a good approach to explore the role of betaA in Alzheimer's disease and to develop therapeutic strategies relevant to it.

The AMP-activated protein kinase is involved in the regulation of ketone body production by astrocytes.

The possible role of the AMP-activated protein kinase (AMPK), a highly conserved stress-activated kinase, in the regulation of ketone body production by astrocytes was studied. AMPK activity in rat cortical astrocytes was three times higher than in rat cortical neurons. AMPK in astrocytes was shown to be functionally active. Thus, incubation of astrocytes with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), a cell-permeable activator of AMPK, stimulated both ketogenesis from palmitate and carnitine palmitoyltransferase I. This was concomitant to a decrease of intracellular malonyl-CoA levels and an inhibition of acetyl-CoA carboxylase/fatty acid synthesis and 3-hydroxy-3-methylglutaryl-CoA reductase/cholesterol synthesis. Moreover, in microdialysis experiments AICAR was shown to stimulate brain ketogenesis markedly. The effect of chemical hypoxia on AMPK and the ketogenic pathway was studied subsequently. Incubation of astrocytes with azide led to a remarkable drop of fatty acid beta-oxidation. However, activation of AMPK during hypoxia compensated the depression of beta-oxidation, thereby sustaining ketone body production. This effect seemed to rely on the cascade hypoxia --> increase of the AMP/ATP ratio --> AMPK stimulation --> acetyl-CoA carboxylase inhibition --> decrease of malonyl-CoA concentration --> carnitine palmitoyltransferase I deinhibition --> enhanced ketogenesis. Furthermore, incubation of neurons with azide blunted lactate oxidation, but not 3-hydroxybutyrate oxidation. Results show that (a) AMPK plays an active role in the regulation of ketone body production by astrocytes, and (b) ketone bodies produced by astrocytes during hypoxia might be a substrate for neuronal oxidative metabolism.

Subgroups of parkinsonian patients differentiated by peptidergic immunostaining of caudate nucleus biopsies.

In this study, Met-enkephalin (Met-enk), substance P (SP) and tyrosine hydroxylase (TH) immunostaining was assessed in caudate nucleus biopsies from 15 Parkinson's disease patients who were treated surgically. According to the combination of changes in Met-enk, SP and TH immunostaining, several subgroups of parkinsonian patients were disclosed. Group I: Patients showing low SP and normal Met-enk immunostaining, and variably reduced TH immunoreactivity. Group II: both SP and Met-enk immunostaining were apparently of normal intensity in these PD patients, but they showed the greatest decrease in TH labeling. Group III: PD patients that showed normal SP, very low Met-enk and variably reduced TH immunostaining. Low Met-enk immunostaining tended to correlate with the severity of the disease as judged by higher Unified Parkinson's disease Rating Scale and gait scores. These results suggest that different neurochemical phenotypes may exist among Parkinson's disease patients. Peptidergic deficits should be taken into account for therapeutic intervention.

In vivo effects of new inhibitors of catechol-O-methyl transferase.

1. The effects of two new synthetic compounds showing in vitro catechol-O-methyl transferase (COMT) inhibitor properties were studied in vivo and compared with the effects of nitecapone and Ro-41-0960. 2. QO IA (3-(3-hydroxy-4-methoxy-5-nitrobenzylidene)-2,4-pentanedione), QO IIR ([2-(3,4-dihydroxy-2-nitrophenyl)vinyl]phenyl ketone), nitecapone and Ro-41-0960 (30 mg kg(-1), i.p.) were given to reserpinized rats 1 h before the administration of L-DOPA/carbidopa (LD/CD, 50:50 mg kg(-1), i.p.). Locomotor activity was assessed 1 h later. All the COMT inhibitors (COMTI), with the exception of QO IA, markedly potentiated LD/CD reversal of reserpine-induced akinesia. Similar results were obtained when the COMTI were coadministered with LD/CD. The effect of compound QO IIR was dose-dependent (7.5-30 mg kg(-1), i.p.). 3. The COMTI (30 mg kg(-1), i.p.) potentiated LD/CD reversal of both catalepsy and hypothermia of reserpinized mice. 4. QO IIR, nitecapone and Ro-41-0960 (30 mg kg(-1), i.p.) reduced striatal 3-methyl-DOPA (3-OMD) levels and increased dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) levels. Compound QO IA was devoid of any effect on striatal amine levels. In contrast to the other inhibitors, Ro-41-0961 reduced HVA levels as well. The effect of QO IIR on striatal amine levels was dose-dependent (7.5-60 mg kg(-1), i.p.) 5. These results suggest that the new compound QO IIR is an effective peripherally acting COMT inhibitor in vivo.

Alzheimer's disease: relationship between muscarinic cholinergic receptors, beta-amyloid and tau proteins.

Senile dementia is one of the most important health problems in developed countries. The main disease causing dementia is Alzheimer's disease that is characterized by the progressive deterioration of the cholinergic system, beta-amyloid production and deposition, and neurofibrillary tangle formation. Most of the reviewed data, along with data from experiments performed in our laboratory, suggest that there are no changes in the number of muscarinic receptors between Alzheimer and control brains, although the receptors expressed in Alzheimer's disease brains can be anomalous in their function. The muscarinic receptor-G-protein interaction also seems to be impaired in Alzheimer's disease compared with control brains, as well as the G-protein system, with an important decrease in the function of the Gq/11, the most important G-protein stimulating phosphoinositide hydrolysis in human brain; in addition, the second messenger system is also impaired, with a decrease in the synthesis of phosphoinositides and in the number of IP3 receptors. Muscarinic cholinergic receptors are also linked to beta-amyloid production, stimulation of the M1 subtype with agonists results in the processing of the beta-amyloid precursor protein to non-amyloidogenic products and administration of a fraction of the beta-amyloid (beta-amyloid 25-35) to rats, results in a decrease in the number of muscarinic receptors in brain. M1 agonists also decrease the phosphorylation of tau proteins, playing again a modulatory role in the pathogenesis of Alzheimer's disease. The existence of a link between beta-amyloid and tau proteins also has been reported; treatment of hippocampal neurones with beta-amyloid, or the 25-35 residue fragment, resulted in an increase in tau protein phosphorylation. The particular contribution of muscarinic receptors, beta-amyloid and tau proteins in the pathogenesis of Alzheimer's disease remains still unclear. Probably Alzheimer's disease could be due to a progressive degeneration in the relationship between the three components covered in this review.

Alterations in peptide levels in Parkinson's disease and incidental Lewy body disease.

The levels of the neuropeptides Met- and Leu-enkephalin (MET-ENK, LEU-ENK), substance P and neurotensin were measured by a combined high performance liquid chromatography/radioimmunoassay (HPLC/RIA) method in postmortem samples of basal ganglia from Parkinson's disease patients, incidental Lewy body disease patients (pre-symptomatic Parkinson's disease) and matched controls. Dopamine (DA) levels were reduced in the caudate nucleus and putamen in Parkinson's disease, but unaltered in incidental Lewy body disease. The levels of MET-ENK were reduced in the caudate nucleus, putamen and substantia nigra in Parkinson's disease. Met-enkephalin levels were reduced in the caudate nucleus and in the putamen in incidental Lewy body disease. Leu-enkephalin levels were decreased in the putamen and were undetectable in the substantia nigra in Parkinson's disease. Leu-enkephalin levels were unchanged in incidental Lewy body disease, although there was a tendency to a reduction in putamen. Substance P levels were reduced in the putamen in Parkinson's disease. No significant changes in substance P content were observed in incidental Lewy body disease. Neurotensin levels were increased in the substantia nigra in Parkinson's disease. Neurotensin levels in incidental Lewy body disease were not altered significantly, but tended to parallel the changes in Parkinson's disease. The changes in basal ganglia peptide levels in incidental Lewy body disease generally followed a trend similar to those seen in Parkinson's disease, but were less marked. This suggests that they are an integral part of the pathology of the illness and not secondary to DA neuronal loss or a consequence of prolonged drug therapy.

Synthesis and affinities for dopamine (D2) and 5-hydroxytryptamine (5-HT2A) receptors of 1-(benzoylpropyl)-4-(1-oxocycloalkyl-2-ethyl)-piperazines as cyclic butyrophenone derivatives.

Starting from benzo- or thienocycloalkaneacetic acids, we have prepared a series of 1-(3-p-fluorobenzoylpropyl)-4-(1-oxo-benzo- or thienocycloalkyl-2-ethyl)piperazines 8a-e containing both semirigid and linear butyrophenones pharmacophores. The affinities of these compounds for dopamine (D2) and 5-hydroxytryptamine (5-HT2A) receptors were evaluated in vitro in receptor-binding assays and in functional experiments. The ratios of pKi's for 5-HT2A/D2 receptors may be useful for rapid screening of new compounds and assessing potential induction of extrapyramidal symptoms; ratio values > or = 1.12 (Meltzer's ratio) are predictive of an atypical antipsychotic profile. The new molecules had a ratio in the range of 0.96-1.11 while haloperidol showed a ratio of 0.93. The 2-piperazinoethyl thiotetralone derivative 8d (QF 0506B) with a ratio of 1.11 was the most active compound.

Characterization of neurotensin-like immunoreactivity in human basal ganglia: increased neurotensin levels in substantia nigra in Parkinson's disease.

A method that combines high performance liquid chromatography with radioimmunoassay (HPLC/RIA) has been used to characterize neurotensin-like immunoreactivity (NT-IR) in the basal ganglia from control subjects and Parkinson's disease (PD) patients. In samples from the caudate nucleus and putamen, NT-IR eluted as two HPLC peaks. One was indistinguishable from the synthetic tridecapeptide, while the other peak corresponded to oxidized NT, as judged by its chromatographic behaviour and its reaction with the antiserum employed. There were marked discrepancies between the IR detected in crude extracts and that in HPLC purified samples. NT levels (HPLC/RIA) were unaltered in the caudate nucleus, putamen and both segments of the globus pallidus in the parkinsonian brain. In contrast, there was a two-fold increase in NT content in both zona compacta and zona reticulata of the substantia nigra in PD patients compared to controls. Degeneration of the nigrostriatal pathway and/or prolonged antiparkinsonian treatment in PD appears to alter neurotensin levels in an attempt to activate the dopaminergic nigrostriatal pathway.

Neurotensin, substance P, delta and mu opioid receptors are decreased in basal ganglia of Parkinson's disease patients.

The specific binding of [3H]neurotensin, [3H]substance P, [3H]D-Ala2-D-Leu5-enkephalin (delta receptors) and [3H]-Tyr-D-Ala-Gly-(NMe)Phe-Gly-ol (mu receptors) were studied in membrane preparations of caudate nucleus, putamen, globus pallidus and substantia nigra from patients with Parkinson's disease and from age-matched controls. The density of neurotensin receptors was decreased in globus pallidus (lateral and medial segments) in parkinsonian brain. Substance P receptors were reduced in the putamen (anterior and posterior) and in lateral globus pallidus in Parkinson's disease. There was a reduction in the density of opioid receptors in posterior putamen and in mu receptors in caudate nucleus and putamen (anterior and posterior). No differences in neuropeptide receptor binding were observed in substantia nigra from parkinsonian brains compared with control subjects. The reductions in neuropeptide receptor density were less marked than the decrease in caudate and putamen content of dopamine and its metabolites. This suggests that neuropeptide receptors are only partially localized to striatal dopamine terminals.

Parallel alterations in Met-enkephalin and substance P levels in medial globus pallidus in Parkinson's disease patients.

Met-enkephalin (Met-enk) and substance P (SP) were measured by a combined high-performance liquid chromatography/radioimmunoanalysis method in medial (GPM) and lateral globus pallidus (GPL) from controls and from Parkinson's disease (PD) patients. All patients showed a similar marked (> 90%) reduction in dopamine (DA) levels in putamen compared with controls. However, based on DA levels in the caudate nucleus, two subgroups of PD patients were differentiated. In patients with > 80% decrease in caudate nucleus DA content, there was a three-fold increase in both Met-enk and SP levels in GPM. In contrast, in patients showing an approximately 50% reduction in DA content in caudate, levels of both peptides were markedly reduced (approximately 80%). Met-enk and SP levels in GPL were unchanged in PD. These results suggest that neurons containing Met-enk and SP projecting to GPM adapt according to the extent of degeneration in the substantia nigra in PD.

Synthesis and inhibitory activities against enkephalin degrading aminopeptidase of H-Trp(Nps)-Lys-OMe analogues bearing chelating groups.

With the aim of increasing the inhibitory potency of the analgesic dipeptide H-Trp(Nps)-Lys-OMe against enkephalin-degrading aminopeptidases, the following derivatives bearing chelating groups at the N-terminus have been synthesized: Ac-Trp(Nps)-Lys-OMe (3), HS(CH2)nCO-Trp(Nps)-Lys-OMe [n = 1 (4), n = 2 (5)], MeOCO(CH2)n-Trp(Nps)-Lys-OMe [n = 1 (6), n = 2 (7)] and analogues in which the N alpha-amino group has been replaced by a methoxycarbonyl group (8) and a bidentate hydroxamate function (9), respectively. The inhibitory activities of all these compounds and the S-protected derivatives EtNHCOS(CH2)nCO-Trp(Nps)-Lys-OMe [n = 1 (16), n = (17)] against the mentioned enzyme, isolated from rat striatum, are compared with those of the parent dipeptide 2 and bestatin. All the new derivatives showed, in general, inhibitory potencies of the same order of magnitude as compound 2.

Striatal neuropeptide levels in Parkinson's disease patients.

Substance P (SP), Met-enkephalin (Met-enk) and cholecystokinin-8-S (CCK-8-S) were measured by a combined HPLC/RIA method in the caudate nucleus and anterior putamen from controls and from Parkinson's disease (PD) patients. SP levels were reduced in caudate in PD, but unchanged in putamen. No differences in Met-enk content were found in parkinsonians compared to controls. However, a significant correlation between DA and Met-enk levels in caudate nucleus from PD was observed. The concentration of CCK-8-S was unaltered in caudate nucleus or putamen in PD. The decrease in caudate nucleus SP levels might be related to the decrease in nigral SP levels in PD, while the reduction in Met-enk levels appears to be a feature of a subgroup of parkinsonian patients.