Longitudinal Behavioral and 6-[F]Fluoro-l-DOPA-PET Assessment in MPTP-Hemiparkinsonian Monkeys

Biochemical and behavioral criteria were established to determine the long-term stability of a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced unilateral striatal dopamine deficiency in the vervet monkey. At time points over a 12-month period, post-MPTP striatal dopamine synthesis capacity was indexed with 6-[¹⁸F]fluoro-l-DOPA (FDOPA)-positron emission tomography. For the MPTP-treated subjects (n = 4), an intrasubject FDOPA influx rate constant (K_i) ratio method of right (lesioned) striatum/left (unlesioned) striatum values was used to assess changes in striatal activity. Striatal FDOPAK_iratios differed less than 5% between studies conducted at 1–2, 5–7, and 9–11 months post-MPTP; these results indicated a stable MPTP-induced striatal lesion over this time period. At the 5–7 and 9–11 month time points, behavioral indices of the MPTP-induced deficits were obtained within a species-typical group setting. For three of the four subjects, persistent decrements in motoric, affiliative, and vigilance behavior were observed while the frequency of aggression toward group members was increased. At the 9–11 month time point, one subject showed a 30% improvement in the social measures, indicative of a partial recovery from the MPTP-induced behavioral decrements although its striatal FDOPAK_iratio remained unchanged. Thus, behavioral and noninvasive biochemical methods can provide complementary indices to assess individual differences in sensitivity to MPTP-induced deficits. Both types of data are required to determine lesion stability and, subsequently, the efficacy of interventions designed to restore normal function in this primate Parkinsonian model.     

Cerebral 6-[F]fluoro-l-DOPA uptake in rhesus monkey: pharmacological influence of aromatic amino acid decarboxylase (AAAD) and catechol-O-methyltransferase (COMT) inhibition

FDOPA/PET scans were performed in one rhesus monkey to study the influence of three catechol-O-methyltransferase (COMT) inhibitors (CGP 28014, OR-611 and Ro 40-7592) on FDOPA pharmacokinetics. COMT inhibitors were administered in combination with carbidopa, a peripherally acting inhibitor of the aromatic amino acid decarboxylase (AAAD). FDOPA was administered intravenously and its metabolic fate in plasma was determined using an HPLC system with an on-line γ-γ coincidence detector. Cerebral tracer uptake was assessed in the striatum and in a non-dopaminergic brain region (occipital cortex). In the periphery, the pharmacokinetic efficiency of FDOPA was increased due to the combined inhibition of COMT and AAAD activity. All three COMT inhibitors reduced the FDOPA methylation rate constant in plasma, with complete suppression obtained in the case of Ro 40-7592. In the brain, specific ¹⁸F radioactivity (striatal minus brain reference radioactivity) increased as a result of the increase in FDOPA plasma availability following the administration of COMT and AAAD inhibitors. We established a significant linear correlation between striatal radioactivity and FDOPA plasma levels (r=0.924±0.048, P<0.0001 for total striatal and r=0.948±0.054, P<0.0001 for specific striatal radioactivity). Using plasma FDOPA radioactivity as input, we found that the striatal FDOPA uptake rate constant K_i^FD was not changed by any of the inhibitors. Thus, the enhancement of striatal radioactivity after application of enzyme inhibitors is a consequence of the increase in plasma FDOPA that becomes available for conversion to fluorodopamine in the striatal dopaminergic nerve terminals. By contrast, using the radioactivity in a non-dopaminergic region (cortex) as input, we found that the striatal FDOPA uptake rate constant K_i^ref was significantly (P<0.0001) increased following pretreatment with COMT inhibitors. Our analysis demonstrated that K_i^ref and the 3-OMFD contribution to the cerebral radioactivity were inversely correlated.     

6-[18F]fluoro-L-DOPA-PETstudies show partial reversibility of long-term effects of chronic amphetamine in monkeys

The acute and long-term effects of chronic amphetamine administration on the striatal dopamine system in monkeys were assessed with 6-[¹⁸F]fluoro-L-DOPA (FDOPA) and positron emission tomography (PET). Vervet monkeys (Cerecopithecus aethiops) were administered amphetamine doses, i.m., that increased from 4 mg/kg/d to 18 mg/kg/d over a 10 day period. Post-amphetamine FDOPA-PET scans at 1–2, 3–4, and 6 week time points in individual subjects showed persistent decrements in dopamine synthesis capacity as reflected by FDOPA influx rate constant (K_i) values being ∼30% that of pre-drug assessment. In other animals that were administered the same drug regimen, biochemical analysis of striatal regions at 1–2 weeks post-drug indicated that dopamine concentrations were decreased by ∼95% throughout caudate and putamen regions, while the homovanillic acid/dopamine level ratio was increased 3–10-fold. Post-drug FDOPA-PET K_i values remained consistently low up to 6 weeks; however, at the 5–6 month time point, relative increases in FDOPA-K_i values (∼53% of pre-drug values) were observed for all subjects, indicative of partial recovery of striatal dopamine synthesis capacity. These results demonstrate that FDOPA-PET can reveal temporal activity changes within the striatal dopamine system of individual subjects. The apparent, partial reversibility of amphetamine's neurotoxic effects suggests a plasticity of dopaminergic function that may include regeneration of dopaminergic terminals and compensatory increases in residual dopamine synthesis rates. The persistence of the partial decrement in dopamine synthesis capacity, however, may indicate a long term component of amphetamine's toxic effects. © 1996 Wiley-Liss, Inc.     

Ventral striatal prediction error signaling is associated with dopamine synthesis capacity and fluid intelligence

Fluid intelligence represents the capacity for flexible problem solving and rapid behavioral adaptation. Rewards drive flexible behavioral adaptation, in part via a teaching signal expressed as reward prediction errors in the ventral striatum, which has been associated with phasic dopamine release in animal studies. We examined a sample of 28 healthy male adults using multimodal imaging and biological parametric mapping with (1) functional magnetic resonance imaging during a reversal learning task and (2) in a subsample of 17 subjects also with positron emission tomography using 6‐[¹⁸F]fluoro‐L‐DOPA to assess dopamine synthesis capacity. Fluid intelligence was measured using a battery of nine standard neuropsychological tests. Ventral striatal BOLD correlates of reward prediction errors were positively correlated with fluid intelligence and, in the right ventral striatum, also inversely correlated with dopamine synthesis capacity (FDOPA K). When exploring aspects of fluid intelligence, we observed that prediction error signaling correlates with complex attention and reasoning. These findings indicate that individual differences in the capacity for flexible problem solving relate to ventral striatal activation during reward‐related learning, which in turn proved to be inversely associated with ventral striatal dopamine synthesis capacity. Hum Brain Mapp, 2013. © 2012 Wiley Periodicals, Inc.     

Striatal [18F]fluorodopa utilization after COMT inhibition with entacapone studied with PET in advanced Parkinson's disease

Summary The effect of peripheral catechol-O-methyltransferase (COMT) inhibition with entacapone on striatal uptake of 6-[¹⁸F]fluoro-L-dopa (FDOPA) was studied with PET both without and with entacapone in fifteen advanced parkinsonian patients and six healthy controls. Entacapone significantly enhanced the fraction of unmetabolized FDOPA in plasma from 16% to about 50% at 80 minutes after FDOPA injection in all subjects. The striatal to occipital ratios and the striatal FDOPA uptake, expressed as a modified decarboxylation coefficient (k₃R₀), was significantly increased in healthy controls, whereas in parkinsonian patients the increase was significant only in the caudate. On the other hand, the influx constant (K_i) decreased significantly in the caudate and putamen in parkinsonian patients; in healthy controls the K_i remained virtually unchanged.Effective peripheral COMT inhibition markedly increased the fraction of FDOPA in plasma and thus its availability in the brain for decarboxylation both in patients and control subjects. However, the change in striatal FDOPA uptake was modest in the advanced parkinsonian patients as compared to that in control subjects, because of the advanced disease, decreased storage capacity, or both.     

6-[18F]fluoro-L-DOPA PET studies of the turnover of dopamine in MPTP-induced parkinsonism in monkeys

This report describes a method to assess, in vivo, the turnover of dopamine (DA) and describes its application to the evaluation of DA function in normal monkeys and monkeys with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced lesions of the DA nigro–striatal pathway. Using positron emission tomography with the tracer of presynaptic DA function, 6-[¹⁸F]fluoro-L -DOPA (FDOPA), and an extension of the graphical method of analysis, we measured the striatal FDOPA uptake rate constant, K_i, and the rate of reversibility of FDOPA trapping k_loss in normal and MPTP-treated monkeys, either neurologically normal or displaying a parkinsonian symptomatology. An index of effective DA turnover was defined as the ratio of k_loss/K_i. Compared to normal controls, K_i was decreased and k_loss was increased in the MPTP-lesioned monkeys. The index of DA turnover was significantly increased in the monkeys displaying a parkinsonian symptomatology as compared to the controls and the neurologically normal MPTP-treated monkeys. The DA turnover index was also significantly increased in the neurologically normal MPTP-lesioned animals compared to normals. This suggests that an increase in DA turnover develops early in the disease process and may be one of the compensatory mechanisms partly responsible for the delay in the development of the clinical manifestations in Parkinson's disease. Synapse 29:225–232, 1998. © 1998 Wiley-Liss, Inc.     

Influence of O-methylated metabolite penetrating the blood–brain barrier to estimation of dopamine synthesis capacity in human L-[β-11C]DOPA PET

O-methyl metabolite (L-[β-¹¹C]OMD) of ¹¹C-labeled L-3,4-dihydroxyphenylalanine (L-[β-¹¹C]DOPA) can penetrate into brain tissue through the blood–brain barrier, and can complicate the estimation of dopamine synthesis capacity by positron emission tomography (PET) study with L-[β-¹¹C]DOPA. We evaluated the impact of L-[β-¹¹C]OMD on the estimation of the dopamine synthesis capacity in a human L-[β-¹¹C]DOPA PET study. The metabolite correction with mathematical modeling of L-[β-¹¹C]OMD kinetics in a reference region without decarboxylation and further metabolism, proposed by a previous [¹⁸F]FDOPA PET study, were implemented to estimate radioactivity of tissue L-[β-¹¹C]OMD in 10 normal volunteers. The component of L-[β-¹¹C]OMD in tissue time-activity curves (TACs) in 10 regions were subtracted by the estimated radioactivity of L-[β-¹¹C]OMD. To evaluate the influence of omitting blood sampling and metabolite correction, relative dopamine synthesis rate (k_ref) was estimated by Gjedde–Patlak analysis with reference tissue input function, as well as the net dopamine synthesis rate (K_i) by Gjedde–Patlak analysis with the arterial input function and TAC without and with metabolite correction. Overestimation of K_i was observed without metabolite correction. However, the k_ref and K_i with metabolite correction were significantly correlated. These data suggest that the influence of L-[β-¹¹C]OMD is minimal for the estimation of k_ref as dopamine synthesis capacity.     

In-vivo measurement of LDOPA uptake,dopamine reserve and turnover in the rat brain using [18F]FDOPA PET

Longitudinal measurements of dopamine (DA) uptake and turnover in transgenic rodents may be critical when developing disease-modifying therapies for Parkinson''s disease (PD). We demonstrate methodology for such measurements using [¹⁸F]fluoro-3,4-dihydroxyphenyl-L-alanine ([¹⁸F]FDOPA) positron emission tomography (PET). The method was applied to 6-hydroxydopamine lesioned rats, providing the first PET-derived estimates of DA turnover for this species. Control (n=4) and unilaterally lesioned (n=11) rats were imaged multiple times. Kinetic modeling was performed using extended Patlak, incorporating a k_loss term for metabolite washout, and modified Logan methods. Dopaminergic terminal loss was measured via [¹¹C]-(+)-dihydrotetrabenazine (DTBZ) PET. Clear striatal [¹⁸F]FDOPA uptake was observed. In the lesioned striatum the effective DA turnover increased, shown by a reduced effective distribution volume ratio (EDVR) for [¹⁸F]FDOPA. Effective distribution volume ratio correlated (r>0.9) with the [¹¹C]DTBZ binding potential (BP_ND). The uptake and trapping rate (k_ref) decreased after lesioning, but relatively less so than [¹¹C]DTBZ BP_ND. For normal controls, striatal estimates were k_ref=0.037±0.005 per minute, EDVR=1.07±0.22 and k_loss=0.024±0.003 per minute (30 minutes turnover half-time), with repeatability (coefficient of variation) ≤11%. [¹⁸F]fluoro-3,4-dihydroxyphenyl-L-alanine PET enables measurements of DA turnover in the rat, which is useful for developing novel therapies for PD.     

Neurotoxic methamphetamine regimens produce long‐lasting changes in striatal G‐proteins

Animals repeatedly dosed with methamphetamine during a single day suffer damage to brain dopamine and serotonin terminals and show behavioral deficits. These methamphetamine regimens also produce long‐term reductions in dopamine agonist‐stimulated immediate‐early gene responses both in striatum and several cortical areas, but the mechanism(s) underlying these long‐lasting effects of methamphetamine remain uncertain. Six weeks after a neurotoxic regimen of methamphetamine (4 × 4 mg/kg) or saline, α subunit levels of striatal G‐proteins that couple dopamine receptors to second messenger systems were measured. Because the damage to striatal monoamine terminals produced by methamphetamine is regionally heterogeneous, we used radioimmunocytochemistry, which combines quantification with regional resolution. We found significant increases in G_iα and G_olfα expression in the ventral striatum (but not in the dorsolateral striatum or nucleus accumbens) of methamphetamine‐pretreated rats, a regional pattern similar to that reported for methamphetamine effects on dopamine terminal markers. By contrast, G_qα expression was unaffected in all striatal subregions. The central roles of G_i and G_olf in modulating the activity of a series of interlinked intracellular signaling pathways suggest that methamphetamine‐induced changes in G_i and G_olf can have lasting effects on striatal neuronal function. Synapse 64:839–844, 2010. © 2010 Wiley‐Liss, Inc.     

Longitudinal Positron Emission Tomography of Dopamine Synthesis in Subjects with GBA1 Mutations

Mutations in GBA1, the gene mutated in Gaucher disease, are a common genetic risk factor for Parkinson disease, although the penetrance is low. We performed [¹⁸F]-fluorodopa positron emission tomography studies of 57 homozygous and heterozygous GBA1 mutation carriers (15 with parkinsonism) and 98 controls looking for early indications of dopamine loss using voxelwise analyses to identify group differences in striatal [¹⁸F]-fluorodopa uptake (K_i). Forty-eight subjects were followed longitudinally. Cross-sectional and longitudinal comparisons of K_i and K_i change found significant effects of Parkinson disease. However, at baseline and over time, striatal [¹⁸F]-fluorodopa uptake in mutation carriers without parkinsonism did not significantly differ from controls. ANN NEUROL 2020;87:652–657     

Differential effects of dopamine agonists on evoked dopamine release from slices of striatum and nucleus accumbens in rats

The effects of dopamine receptor agonists on electrically evoked dopamine release from slices of nucleus accumbens were compared with the effects on release from striatal slices in rats. Apomorphine, which has equal potency at the dopamine D₂ and D₃ receptors, reduced the evoked dopamine release from both regions to the same extent (ED₅₀, 0.42 μM for nucleus accumbens; ED₅₀, 0.46 μM for striatum). Quinpirole of 7-[³H]hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OHDPAT), which are much more potent at the D₃ receptor than at the D₂ receptor, reduced the evoked dopamine release from the nucleus accumbens (ED₅₀, 0.12 μM for quinpirole; 0.02 μM for 7-OHDPAT) much more than the release from the striatum (ED₅₀, 1.6 μM for quinpirole; 0.55 μM for 7-OHDPAT). These results suggest that the contribution of D₃ receptors in nucleus accumbens to regulate dopamine release from dopamine nerve terminals is much greater than that in striatum.     

Increased striatal neuropeptide Y immunoreactivity and its modulation by deprenyl,clonidine and L-dopa in MPTP-treated mice

Summary. The aim of this study was to evaluate the effect of MPTP (2 × 45mg/kg s.c., 20h apart) on striatal neuropeptide Y-like immunoreactivity (NPY-LI) in C57BL/6 mice. NPY-LI markedly increased 2 weeks after MPTP but it remained unchanged after 24h, 1 or 6 weeks. The increase in NPY-LI was accompanied by depletion of dopamine (–80%), DOPAC (–70%), 3-MT (–44%) and HVA (–52%). L-Deprenyl completely prevented the MPTP-induced NPY-LI increase, neurodegeneration of the striatal dopamine system and motor dysfunction. Clonidine attenuated the neurotoxin effect on NPY-LI and dopaminergic neurons. L-dopa/carbidopa protected NPY neurons against MPTP but slightly enhanced MPTP-induced decrease in the levels of dopamine and its metabolites. The relationship between changes in NPY-LI and dopamine and serotonin metabolism determined by HPLC was discussed. The results further extend the range of MPTP-elicited modifications in striatum and demonstrate that drugs with antiparkinsonian activity can protect NPY neurons against MPTP toxicity.     

Effects of Radiochemical Impurities on Measurements of Transfer Constants for [C]Sucrose Permeation of Normal and Injured Blood–Brain Barrier of Rats

Radiolabeled sucrose is often used to assess blood–brain barrier (BBB) injury in the rat, but published transfer constants (K_is) for sucrose permeation of the intact BBB (control K_is) are highly discrepant. A potential problem with the commonly used tracer, [¹⁴C(U)]sucrose, is radiolytic generation, preuse, of radiocontaminants that might readily penetrate the BBB. How such contaminants might affect measurements of sucrose K_is was examined for both the intact and the ischemically injured BBB. Three stocks of [¹⁴C(U)]sucrose were studied: newly purchased (“new”), 4-year-old, and 7-year-old. A high purity (99.9%) “new” and a 2-year-old stock of [³H(fructose-1)]sucrose were also tested. Pentobarbital-anesthetized male Sprague–Dawley rats were injected IV with each tracer separately (six to eight rats) and K_is in five brain regions were measured by the multiple-time graphical method. The “new” ¹⁴C-, “new” ³H-, and 2-year-old ³H-sucrose yielded comparable K_is, ranging from 1.2 ± 0.1 to 2.4 ± 0.3 nl · g⁻¹ · s⁻¹ (mean ± SE) across the regions. The two old stocks of ¹⁴C-sucrose yielded significantly higher regional K_is: 5.1–6.3 (4-year-old) and 8.4–9.7 (7-year-old). Thin-layer chromatography of the three ¹⁴C-tracers revealed that each contained radioimpurities (ca. 2% in both the “new” and 4-year-old, and 9% in the 7-year-old), but that the old stocks contained larger amounts of relatively mobile (more lipophilic) impurities, which can be suspected as the main cause of the elevated K_is obtained. Additional rats were subjected to 10 min of cerebral ischemia, which effects a delayed BBB injury, and 6 h later the “new” ³H- and the 4-year-old ¹⁴C-sucrose were injected together. The K_is for both tracers were elevated by like, absolute amounts (ΔK_is), but by very different percentages, over their disparate baseline values in uninjured rats (for striatum and hippocampus, the most injured regions, ΔK_is were 3.9 to 4.4 nl · g⁻¹ · s⁻¹). It is concluded that radiolysis of [¹⁴C(U)]sucrose yields certain labeled products that readily cross the BBB and that can seriously distort baseline K_is, even if present only in very small amounts. While this appears not to compromise assessment of BBB injury, definition of the authentic range of baseline, sucrose K_is for the rat BBB would appear to remain a challenge.     

Striatal 6-[18F]fluorodopa accumulation after combined inhibition of peripheral catechol-O-methyltransferase and monoamine oxidase type B: Differing response in relation to presynaptic dopaminergic dysfunction

The aim was to investigate the effects of inhibition of monoamine oxidase type B (MAO-B) with selegiline alone and the combined inhibition of peripheral catechol-O-methyltransferase (COMT) with entacapone and MAO-B with selegiline on striatal 6-[¹⁸F]fluorodopa (FDOPA) accumulation, and whether the effect of entacapone + selegiline on FDOPA uptake differed depending on the severity of the presynaptic dopaminergic dysfunction. Thus, eight healthy controls, eight de novo patients with Parkinson's disease (PD), and 18 levodopa-treated PD patients were investigated with positron emission tomography (PET). Half of the subjects in each population belonged to the selegiline group and half to the entacapone + selegiline group. Both groups were studied twice with PET using FDOPA. After the first (baseline) FDOPA PET investigation, both groups were on 2 weeks of selegiline treatment, 10 mg daily. Thereafter, the second FDOPA PET was performed for all subjects with a premedication administered 60 min before the PET imaging; one group received 10 mg of selegiline, and the other group received a single 400 mg dose of entacapone coadministered with 10 mg of selegiline. Selegiline treatment alone had no significant influence on striatal FDOPA metabolism. The FDOPA accumulation, expressed as striatal-to-occipital ratios and modified decarboxylation coefficients (k₃R₀), increased significantly after entacapone + selegiline administration in all subject populations. The FDOPA uptake rate constant (K_i) remained virtually unchanged in controls and in de novo patients but decreased significantly in levodopa-treated PD patients after entacapone + selegiline intake. Entacapone + selegiline administration did not influence significantly the unidirectional blood-to-brain clearance for FDOPA (K₁^D) or the relative dopadecarboxylase activity (k₃^D). The changes in the studied parameters after entacapone + selegiline administration probably reflect the effects of entacapone, since entacapone alone has caused similar changes in previous PET studies. Response in FDOPA accumulation to entacapone + selegiline was higher in controls and de novo patients compared with levodopa-treated PD patients. The milder response in levodopa-treated patients might reflect the reduced ability of the degenerated dopaminergic neurons to utilize the prolonged FDOPA availability, produced by entacapone. Synapse 27:336–346, 1997. © 1997 Wiley-Liss, Inc.     

Post-ischemic leakiness of the blood–brain barrier: A quantitative and systematic assessment by Patlak plots

The Patlak plot analysis of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) allows estimation of blood–brain barrier (BBB) leakage following temporary focal cerebral ischemia. Thus far, a systematic and quantitative in vivo evaluation of post-ischemic BBB leakage is lacking. Here, using DCE-MRI and the Patlak plot method, we quantitatively assessed BBB leakage in rats at the following time-points after reperfusion: 25 min, 2, 4, 6, 12, 18, 24, 36, 48, and 72 h, and 1, 2, 3, 4, and 5 weeks. Sham-operated animals served as controls. Data collected for each time-point were: the blood-to-brain transfer rate constant (K_i) of the contrast agent gadolinium, distribution volume (V_p), ischemic lesion volume, and apparent diffusion coefficient (ADC) values. Compared to controls, K_i, measured at all time-points, except for 5 weeks, appeared significantly different (p < 0.001). At several time-points (25 min, 48 and 72 h, 4 and 5 weeks), V_p was similar compared to that of controls, but for the remaining groups the difference was significant (p < 0.001). Analyzing the relationship of K_i values to time-points, we observed a trend towards a decrease over time (r = − 0.61, p = 0.014). Both ADC values (r = − 0.58, p = 0.02) and ischemic lesion volumes (r = 0.75, p = 0.0015) correlated with K_i values. These results suggest that after ischemia–reperfusion in rats, BBB leakage is continuous during a 4-week period. Its magnitude diminishes over time and correlates with severity and extent of ischemic injury.     

Striatal dopaminergic D1 and D2 receptors after intracerebroventricular application of alloxan and streptozocin in rat

Summary Intracerebroventricular application of low, nondiabetogenic doses (500 g kg^–1) of alloxan and streptozocin is followed by alterations of the dopaminergic system in rat striatum. In this brain region the dopamine content significantly increased, while the density of dopaminergic D₁ receptors significantly decreased seven days after the intracerebroventricular application of betacytotoxics, as compared with the control group. The density of dopaminergic D₂ receptors in striatum remained unchanged. Dopaminergic D₁ and D₂ receptors operate through signalling mechanism of G proteins, but no changes of G_s and G_i proteins content have been found in rat striatum after the intracerebroventricular application of betacytotoxics. As intracerebroventricular, nondiabetogenic administration of betacytotoxics produces changes of the striatal dopamine content and D₁ receptor density similar to that produced by peripheral, diabetogenic administration of these drugs, the effect might be related not solely to pancreatic beta cells damage, but to alterations of the brain insulin system, as well.     

Neuropharmacological characterization of local ibogaine effects on dopamine release

Summary Local perfusion with ibogaine (10^–6M—10^–3M) via microdialysis probes in the nucleus accumbens or striatum of rats produced a biphasic dose-response effect on extracellular dopamine levels. Lower doses (10^–6M—10^–4M) produced a decrease while higher doses (5 × 10^–4M—10^–3M) produced an increase in dopamine levels. Dihydroxyphenylacetic acid (DOPAC) levels were not effected. Naloxone (10^–6M) and norbinaltorphimine (10^–6M—10^–5M) did not affect dopamine levels, but when co-administered with ibogaine (10^–4M) blocked the decrease in dopamine levels produced by ibogaine. Ibogaine (10^–3M) stimulation of dopamine levels in the striatum was calcium independent and not blocked by tetrodotoxin (10^–5M). Pretreatment with cocaine (15mg/kg), reserpine (5mg/kg) or alpha-methyl-paratyrosine (250mg/kg) given intraperitoneally significantly reduced ibogaine (10^–3M) stimulation of striatal dopamine levels. In striatal synaptosomes, both ibogaine and harmaline (10^–7—10^–4M) produced dose-dependent inhibition of [³H]-dopamine uptake. These findings suggest that ibogaine has both inhibitory and stimulatory effects on dopamine release at the level of the nerve terminal. It is suggested that the inhibitory effect is mediated by kappa opiate receptors while the stimulatory effect is mediated by interaction with the dopamine uptake transporter.     

Poly(ADP-ribose) polymerase inhibitors protect against MPTP-induced depletions of striatal dopamine and cortical noradrenaline in C57B1/6 mice

Treatment of C57B1/6 mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) reduced striatal dopamine and cortical noradrenaline levels by 77–83% and 43–46%, respectively, at 7 days post-treatment. Co-treatments with five different inhibitors of poly(ADP-ribose) polymerase (PARP), including benzamide, significantly prevented the MPTP-induced catecholamine depletions. Benzamide was present in the striatum, 30 min after single i.p. injection, at low millimolar concentrations known to selectively inhibit PARP in vitro. The protective activities of benzamide and its derivatives paralleled their in vitro efficacies and potencies both as neuroprotective agents and as inhibitors of PARP, while the activity of 1,5-dihydroxyisoquinoline, a structurally-unrelated compound, did not. In naive animals, the PARP inhibitors by themselves did not alter striatal dopamine levels at 7 days post-treatment. However, in acute studies, 1,5-dihydroxyisoquinoline and nicotinamide caused marked alterations in striatal dopamine metabolite levels; on the contrary, benzamide and its amino-derivatives showed little or no effect on dopamine metabolism. These results indicate that, although these compounds might act at other sites in addition to PARP, PARP inhibitors possess neuroprotective potential in vivo and suggest a role for PARP in MPTP neurotoxicity.     

Noninvasive assessment of aromatic L-amino acid decarboxylase activity in aging rhesus monkey brain in vivo

The effect of aging on aromatic L-amino acid decarboxylase (AAAD) activity in rhesus monkey striatum was assessed in vivo using PET imaging. Two analogs of L-DOPA, 6-fluoro-m-tyrosine (FMT) and 6-fluoro-L-DOPA (FDOPA), were used to image rhesus monkeys of various ages. Results show that when the animals were grouped between young (3-11 years) and aged (25-37 years), FDOPA uptake in the older animals showed a 21% decline (P < 0.0005), while FMT uptake in young and older animals were not different. On the other hand, when individual uptake values were plotted vs. age, linear regression analysis showed FDOPA uptake similarly declined with age (r = -0.84, P < 0.001) while FMT uptake increased with age (r = 0.66, P < 0.05). Since FMT pharmacokinetics has been shown to be unaffected by metabolic steps occurring after the AAAD step, while FDOPA traces all the steps involved in L-DOPA metabolism, FMT is a suitable tracer to assess AAAD activity while FDOPA traces dopamine turnover. Based on these tracer characteristics, this study found that AAAD activity is maintained or increased in the aging rhesus monkey striatum while the FDOPA uptake decreases with age consistent with age-related declines in neuronal mechanisms whose overall effect is increased striatal dopamine turnover and clearance. Furthermore, comparison of results of this study with previous studies support the notion that the effect of aging in the dopamine system is different from that of MPTP-induced parkinsonism.     

Decreases in mouse brain NAD and ATP induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP): prevention by the poly(ADP-ribose) polymerase inhibitor, benzamide

Inhibitors of poly(ADP-ribose) polymerase (PARP), including benzamide, protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopamine neurotoxicity in vivo [Cosi et al., Brain Res. 729 (1996) 264–269]. In vitro, the activation of PARP by free radical damaged DNA has been shown to be correlated with rapid decreases in the cellular levels of its substrate nicotinamide adenine dinucleotide (NAD⁺), and ATP. Here, we investigated in vivo whether MPTP acutely caused region- and time-dependent changes in brain levels of NAD⁺, ATP, ADP and AMP in C57BL/6N mice killed by head-focused microwave irradiation, and whether such effects were modified by treatments with neuroprotective doses of benzamide. At 1 h after MPTP injections (4×20 mg/kg i.p.), NAD⁺ was reduced by 11–13% in the striatum and ventral midbrain, but not in the frontal cortex. The ATP/ADP ratio was reduced by 10% and 32% in the striatum and cortex, respectively, but was unchanged in the midbrain. All of these regional changes were prevented by co-treatment with benzamide (2×160 mg/kg i.p.), which by itself did not alter regional levels of NAD⁺, ATP, ADP or AMP in control mice. In a time-course study, a single dose of MPTP (30 mg/kg i.p.) resulted in maximal and transient increases in striatal levels of MPP⁺ and 3-methoxytyramine (+540%) at 0.5–2 h, followed by maximal and coincidental decreases in NAD⁺ (−10%), ATP (−11%) and dopamine content (−39%) at 3 h. Benzamide (1×640 mg/kg i.p., 30 min before MPTP) partially reduced MPP⁺ levels by 30% with little or no effect on MPTP or MPDP⁺ levels, did not affect or even slightly potentiated the increase in 3-methoxytyramine, and completely prevented the losses in striatal NAD⁺, ATP and dopamine content, without by itself causing any changes in these latter parameters in control mice. These results (1) confirm that MPTP reduces striatal ATP levels [Chan et al., J. Neurochem. 57 (1991) 348–351.]; (2) show that MPTP causes a regionally-dependent (striatal and midbrain) loss of NAD⁺; (3) indicate that the PARP inhibitor benzamide can prevent these losses without interfering with MPTP-induced striatal dopamine release; and (4) provide further evidence to suggest an involvement of PARP in MPTP-induced neurotoxicity in vivo.