Combined effects of cabergoline and L-dopa on parkinsonism in MPTP-treated cynomolgus monkeys

Summary The behavioral effects of L-dopa or cabergoline alone were compared with those of the joint administration of the two drugs in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned parkinsonian cynomolgus monkeys with attention to the induction of hyperactivity and dyskinesia. Cabergoline alone at 0.2mg/kg or less improved in a dose-dependent fashion the parkinsonism without inducing hyperactivity and dyskinesia following a single subcutaneous injection. L-dopa alone improved the parkinsonism, but induced hyperactivity and dyskinesia, depending on the dose applied. Doses required for 50% amelioration by L-dopa and cabergoline were 10 and 0.038mg/kg, s.c., respectively. With low doses (50%-amelioration doses), cabergoline or L-dopa alone improved the parkinsonism without induction of hyperactivity and dyskinesia, but the duration of action was brief. Cabergoline in combination with L-dopa was highly effective in improving motor disability without induction of hyperactivity and dyskinesia. Moreover, the duration of action was more prolonged with the coadministration than with the single administration of each drug. These findings suggest that the combined therapy with low doses of L-dopa and cabergoline is beneficial for treating patients with advanced Parkinson's disease.     

Progressive changes of pre- and post-synaptic dopaminergic biomarkers in conscious MPTP-treated cynomolgus monkeys measured by positron emission tomography

Positron emission tomography (PET) is a useful technique for the consecutive investigation of the relationship between changes in neurotransmission biomarkers and behavioral signs in animal models of Parkinson's disease (PD). In this study, we aimed to investigate the threshold of dopamine (DA) neuron damage for the appearance of tremor by observing the longitudinal changes of pre- and post-synaptic DA biomarkers in awake monkeys using PET with multiple tracers. Three cynomolgus monkeys were treated with MPTP every 3-6 weeks until tremor was observed. Brain uptake of [11C]PE2I, [beta-11C]DOPA, and [11C]raclopride for DA transporter (DAT), DOPA utilization, and DA D2 receptor were measured using PET as a single set in awake condition. Sets of PET scans were repeated in parallel with continuous behavioral estimation. The pre-synaptic biomarkers of DA neuron in the striatum decreased [11C]PE2I binding and [beta-11C]DOPA uptake in an MPTP dose-dependent manner. Tremor was not observed until striatal [11C]PE2I binding was reduced to about 15% of the pretreatment level and [beta-11C]DOPA uptake was reduced to about 34%. DA D2 receptor measured by [11C]raclopride was not significantly changed throughout the experiment. Our results revealed that it is possible to quantitatively define the threshold of the onset of behavioral PD signs by monitoring spontaneous motor activity, and in vivo PET with DAT marker can be a biomarker for early diagnosis at the presymptomatic stage of PD and for high-risk groups.     

Locus ceruleus lesions and eosinophilic inclusions in MPTP-treated monkeys

Systemic administration of the recently discovered neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces severe clinical parkinsonism and degeneration of the substantia nigra in humans and monkeys. In previous studies, no convincing structural damage to nerve cells outside the substantia nigra could be demonstrated in subhuman primates. Using a protracted MPTP regimen and older animals, we now report locus ceruleus lesions and eosinophilic inclusion bodies in squirrel monkeys. The inclusions were seen only in areas where Lewy bodies are found in human Parkinson's disease. No such abnormalities were seen in control animals. These findings suggest that similarities between the neuropathology of MPTP-induced parkinsonism in the monkey and human Parkinson's disease are greater than first thought and increase the usefulness of the MPTP monkey model for research in Parkinson's disease.     

Alterations in event-related potentials (ERPs) of MPTP-treated monkeys

Using an auditory 'oddball' paradigm and classical conditioning, we have studied auditory evoked potentials (AEPs) and P300-like potentials in monkeys pre- and post-MPTP treatment. Free-field acoustic stimuli were 500 Hz and 4000 Hz tones, which were designated as the 'frequent' and 'rare' conditions, respectively. The 4000 Hz stimuli were reinforced with mild somatosensory electrical stimulation. During the first few weeks following 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP) administration, all monkeys gradually developed a parkinsonian syndrome, which partially, but not completely improved within 30-40 days in 2 animals. The amplitudes of the AEP were initially significantly decreased, but progressively returned to pretreatment magnitudes in the 2 monkeys which partially recovered. P300-like potentials were initially abolished in all animals; however, 30-40 days later P300 spontaneously re-emerged in the same 2 monkeys. Latencies of both of these signals were unaffected by MPTP. Acute administration of dopamine precursor during the first phase of neurotoxicity partially and temporarily improved depressed AEP amplitudes, but did not restore absent P300-like potentials. The relevance of these results for Parkinson's disease is discussed.     

Differential effects of three dopamine receptor agonists in MPTP-treated monkeys

Summary The behavioral effects of cabergoline, pergolide and bromocriptine were investigated in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned parkinsonian cynomolgus monkeys with attention to the induction of hyperactivity, as evidenced by irritability, excitability and aggressiveness. All three drugs improved the parkinsonism in a dose-dependent fashion following a single injection. Among the three dopamine (DA) receptor agonists used, the antiparkinsonian effect of pergolide was the strongest and had an immediate effect, while cabergoline showed the longest duration of the antiparkinsonian effect and was least potent in inducing hyperactivity.     

Haloperidol induces Fos expression in the globus pallidus and substantia nigra of cynomolgus monkeys

Systemic injections of the dopamine antagonist haloperidol (0.1-2.5 mg/kg) induced a dose dependent increase in Fos-like immunoreactivity (FLI) in the internal segment of the globus pallidus (GPi) and in the substantia nigra (SN) of cynomolgus monkeys. These findings are consistent with models of basal ganglia organization which predict that blockade of dopamine receptors should result in a disinhibition of cells in these structures. In the GPi, labeling was most pronounced along the ventral, lateral and medial borders of the nucleus and none of the pallidal cells expressing FLI were immunopositive for choline acetyltransferase. In the SN, immunoreactive nuclei were concentrated in the pars reticulata and the majority of labeled nigral neurons did not display tyrosine hydroxylase-like immunoreactivity. A small number of cells displaying FLI were also observed in the external pallidal segment, but no labeling was seen in the subthalamic nucleus. These findings indicate that blockade of dopamine receptors induces a characteristic pattern of Fos expression in the primate brain which strongly resembles that previously reported in rodents.     

Neuronal localisation of neuropeptide Y gene expression in rat brain

The cellular location of Neuropeptide Y (NPY) synthesis in rat brain is identified by using in-situ hybridisation histochemistry. The results show that NPY mRNA is widely distributed through the rat brain, although the levels of NPY mRNA are surprisingly low. There is a large degree of variation in the content of NPY mRNA in different regions. The highest cellular levels of NPY mRNA are found in the arcuate nucleus of the hypothalamus, in the hypothalamus, in the cerebral cortex, and in the hilar region of the hippocampus. In general, the distribution of perikarya containing NPY mRNA corresponds to the reported distribution of perikarya containing NPY-immunoreactivity. However, NPY mRNA was detected in the majority of the perikarya in the reticular nucleus of the thalamus, an area not previously known to contain NPY neurones. In many areas of the forebrain the distribution of NPY mRNA parallels that of somatostatin mRNA, supporting suggestions of their coexistence. The ability to detect NPY mRNA at the cellular level should be of considerable use in dynamic studies of the activity of NPY neurones.     

Striatal enkephalin gene expression does not reflect parkinsonian signs

Loss of striatal dopamine has been associated with an increase in striatal enkephalin expression. However, the relationship between increased striatal enkephalin expression and the manifestation of parkinsonian motor deficits is not clear. Administration of MPTP to cats produces a severe parkinsonian condition from which the animals spontaneously recover. Using in situ hybridization histochemistry, preproenkephalin (PPE) mRNA expression was examined in the striatum of cats when normal, symptomatic for or spontaneously recovered from MPTP-induced parkinsonism. In all areas of the striatum, PPE mRNA levels were significantly elevated in animals exhibiting severe parkinsonian motor deficits and remained elevated even after recovery of gross motor functioning. These results show that striatal PPE gene expression and parkinsonian motor deficits are not directly correlated.     

Dopaminergic denervation and spine loss in the striatum of MPTP-treated monkeys

Striatal spine loss is a key pathological feature of human Parkinson's disease (PD) that can be induced after complete degeneration of the nigrostriatal dopaminergic system in rodent models of parkinsonism. In line with these observations, our findings reveal a significant (30–50%) reduction in spine density in both the caudate nucleus and putamen of severely DA-depleted striata of MPTP-treated monkeys; the sensorimotor post-commissural putamen being the most severely affected region for both dopamine depletion and spine loss. Using MPTP-treated monkeys with complete or partial striatal dopamine (DA) denervation, we also demonstrate that striatal spine loss is an early pathological feature of parkinsonism, which progresses along a positive rostrocaudal and mediolateral gradient in parallel with the extent of striatal dopamine denervation. Quantitative electron microscopy immunocytochemistry for D1 dopamine receptor (D1) in the striatum of control and severely DA-depleted animals revealed that both D1-immunoreactive and immunonegative spines are lost in the putamen of MPTP-treated monkeys.These data demonstrate that striatal spine loss in MPTP-treated monkeys is an early pathological event of parkinsonism, tightly correlated with the degree of nigrostriatal dopamine denervation that likely affects both direct and indirect striatofugal pathways.     

Systemic administration of the immunophilin ligand GPI 1046 in MPTP-treated monkeys

Systemic administration of immunophilin ligands provides trophic influences to dopaminergic neurons in rodent models of Parkinson's disease (PD) resulting in the initiation of clinical trials in patients with Parkinson's disease. We believe that prior to clinical trials, novel therapeutic strategies should show safety and efficacy in nonhuman models of PD. The present study assessed whether oral administration of the immunophilin 3-(3-pyridyl)-1-propyl (2S)-1-(3,3-dimethyl-1,2-dioxopentyl)-2-pyrrollidinecarboxylate (GPI 1046) could prevent the structural and functional consequences of n-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration in nonhuman primates. Twenty-five rhesus monkeys received daily oral administration of vehicle (n = 5) or one of four doses of GPI 1046 (0.3 mg/kg, n = 5; 1.0 mg/kg, n = 5; 3.0 mg/kg, n = 5; 10.0 mg/kg, n = 5). Two weeks after starting the drug treatment, all monkeys received a unilateral intracarotid injection of MPTP-HCl (3 mg). Daily drug administration continue for 6 weeks postlesion after which time the monkeys were sacrificed. Monkeys were assessed for performance on a hand reach task, general activity, and clinical dysfunction based on a clinical rating scale. All groups of monkeys displayed similar deficits on each behavioral measure as well as similar losses of tyrosine hydroxylase (TH)-immunoreactive (ir) nigral neurons, TH-mRNA, and TH-ir striatal optical density indicating that in general treatment failed to have neuroprotective effects.     

食蟹猴脊髓小脑性共济失调模型的制作研究

目的 探讨食蟹猴脊髓小脑性共济失调(SCAs)模型的制作. 方法选取2只成年雄性食蟹猴,在不同的时间段内分别采用腹腔注射3-乙酰吡啶(3-AP)、静脉注射3-AP及立体定向注射3-AP损毁小脑中央核群(顶核、齿状核、栓状核)的方法制作食蟹猴SCAs模型,并建立行为学评价体系(行为学评定、取食实验、水平旋转实验等)用于食蟹猴共济运动的评估. 结果首先采用腹腔注射3-AP 8倍有效剂量并观察12周,行为学评价体系结果未出现有关共济失调的症状;再次采用静脉注射3-AP6倍的有效剂量并观察12周,行为学评价体系结果未出现有关共济失调的症状;最后立体定向注射3-AP损毁小脑中央核群,行为学评价体系结果统计分析显示造模后8周内均较造模前差异有统计学意义(P<0.05),而造模后12周起又逐步恢复至造模前状态. 结论立体定向注射3-AP损毁小脑中央核群方法可以制作食蟹猴SCAs的急性模型,行为学评定、取食实验、水平旋转实验适用于该模型的评价.     

食蟹猴脊髓小脑性共济失调模型的制作研究

目的 探讨食蟹猴脊髓小脑性共济失调(SCAs)模型的制作. 方法选取2只成年雄性食蟹猴,在不同的时间段内分别采用腹腔注射3-乙酰吡啶(3-AP)、静脉注射3-AP及立体定向注射3-AP损毁小脑中央核群(顶核、齿状核、栓状核)的方法制作食蟹猴SCAs模型,并建立行为学评价体系(行为学评定、取食实验、水平旋转实验等)用于食蟹猴共济运动的评估. 结果首先采用腹腔注射3-AP 8倍有效剂量并观察12周,行为学评价体系结果未出现有关共济失调的症状;再次采用静脉注射3-AP6倍的有效剂量并观察12周,行为学评价体系结果未出现有关共济失调的症状;最后立体定向注射3-AP损毁小脑中央核群,行为学评价体系结果统计分析显示造模后8周内均较造模前差异有统计学意义(P<0.05),而造模后12周起又逐步恢复至造模前状态. 结论立体定向注射3-AP损毁小脑中央核群方法可以制作食蟹猴SCAs的急性模型,行为学评定、取食实验、水平旋转实验适用于该模型的评价.     

慢性实验性变应性猴脑脊髓炎的超微结构改变

目的：探讨自身免疫性中枢神经系统脱髓鞘疾病慢性型的病理特点及其可能机制。方法：成功建立猴实验性态反应性脑脊髓炎模型数年后，根据MR摄片结果位病灶并作分类，然后进行病理取材和电镜观察超微结构。结果：（1）活动性病灶内成片的髓鞘松解、断裂或融合，轴突空泡样变性，皱缩或消失，少突胶质细胞变性，未见淋巴细胞浸润，仅见散在巨噬细胞，伴明显的间质水肿；（2）可疑活动性病灶内见部分髓鞘内板松解，其内轴突有较度空泡样变，亦见少突胶质细胞变性，散在巨噬细胞，未见淋巴瘤浸润，结论：慢性EAE的病理改变不仅有髓鞘的变性，同时轴突的病变也十分明显。     

Quantitative analysis of dopaminergic loss in relation to functional territories in MPTP-treated monkeys

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication in primates results in a heterogeneous loss of dopamine in the striatum, predominating in the dorsal and caudal parts of the structure, causing functional impairment that appears to be essentially motor and cognitive. The aim of the present study was to quantify the loss of dopamine in relation to the anatomo-functional subdivisions of the striatum, and also of the pallidum and cortex of MPTP-treated monkeys. A severe loss of dopaminergic innervation was observed in both the sensorimotor and associative territories of all these structures in MPTP-treated monkeys. Comparatively, the limbic territories of all these structures were little affected. The preservation of dopaminergic innervation of the limbic part of cerebral structures may explain the preservation of motivational processes mediated by these limbic regions in MPTP-treated monkeys.     

Social instability and coronary artery atherosclerosis in cynomolgus monkeys

Epidemiologic research has increasingly implicated psychosocial factors in the emergence of atherosclerotic coronary heart disease in human beings. The study of behavioral influences on atherogenesis in man, however, is impeded by the difficulty of assessing coronary artery atherosclerosis in asymptomatic individuals and by the fact that significant arterial lesions typically develop only over relatively protracted intervals. Consequently, we have recently attempted development of an appropriate animal model for examining the atherogenic effects of psychosocial variables. In the first of two investigations, an experimental stressor—involving repeated reorganization of socially housed groups of adult, male cynomolgus monkeys (Macaca fasicularis) fed a moderately atherogenic diet—resulted in increased coronary artery atherosclerosis relative to control animals, though only among monkeys which retained dominant social status over the 22 months of the study. In the second investigation, which employed the same experimental procedures among monkeys fed a low cholesterol/low saturated fat diet, periodic social group reorganization similarly led to development of greater atherosclerosis in the coronary arteries. In neither experiment were psychosocial influences on coronary atherogenesis attributable to the concomitant effects of other physiologic variables commonly associated with atherosclerosis (e.g., serum lipids, blood pressure).     

Social instability and coronary artery atherosclerosis in cynomolgus monkeys

Epidemiologic research has increasingly implicated psychosocial factors in the emergence of atherosclerotic coronary heart disease in human beings. The study of behavioral influences on atherogenesis in man, however, is impeded by the difficulty of assessing coronary artery atherosclerosis in asymptomatic individuals and by the fact that significant arterial lesions typically develop only over relatively protracted intervals. Consequently, we have recently attempted development of an appropriate animal model for examining the atherogenic effects of psychosocial variables. In the first of two investigations, an experimental stressor—involving repeated reorganization of socially housed groups of adult, male cynomolgus monkeys (Macaca fasicularis) fed a moderately atherogenic diet—resulted in increased coronary artery atherosclerosis relative to control animals, though only among monkeys which retained dominant social status over the 22 months of the study. In the second investigation, which employed the same experimental procedures among monkeys fed a low cholesterol/low saturated fat diet, periodic social group reorganization similarly led to development of greater atherosclerosis in the coronary arteries. In neither experiment were psychosocial influences on coronary atherogenesis attributable to the concomitant effects of other physiologic variables commonly associated with atherosclerosis (e.g., serum lipids, blood pressure).     

Differential structural plasticity of corticostriatal and thalamostriatal axo-spinous synapses in MPTP-treated Parkinsonian monkeys

Striatal spine loss is a key pathological feature of Parkinson's disease (PD). Knowing that striatal glutamatergic afferents target dendritic spines, these data appear difficult to reconcile with evidence for an increased expression of the vesicular glutamate transporter 1 (vGluT1) in the striatum of PD patients and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys, as well as in some electrophysiological studies showing overactivity of the corticostriatal glutamatergic system in models of parkinsonism. To address the possibility that structural changes in glutamatergic afferents may underlie these discrepancies, we undertook an ultrastructural analysis of vGluT1-positive (i.e., corticostriatal) and vGluT2-positive (i.e., mostly thalamostriatal) axo-spinous glutamatergic synapses using a 3D electron microscopic approach in normal and MPTP-treated monkeys. Three main conclusions can be drawn: 1) spines contacted by vGluT1-containing terminals have larger volume and harbor significantly larger postsynaptic densities (PSDs) than those contacted by vGluT2-immunoreactive boutons; 2) a subset of vGluT2-, but not vGluT1-immunoreactive, terminals display a pattern of multisynaptic connectivity in normal and MPTP-treated monkeys; and 3) VGluT1- and vGluT2-positive axo-spinous synapses undergo ultrastructural changes (larger spine volume, larger PSDs, increased PSD perforations, larger presynaptic terminal) indicative of increased synaptic activity in parkinsonian animals. Furthermore, spines contacted by cortical terminals display an increased volume of their spine apparatus in MPTP-treated monkeys, suggesting an increased protein synthesis at corticostriatal synapses. These findings demonstrate that corticostriatal and thalamostriatal glutamatergic axo-spinous synapses display significantly different ultrastructural features, and that both systems undergo complex morphological changes that could underlie the pathophysiology of corticostriatal and thalamostriatal systems in PD.     

Cortical connections of the lateral mediodorsal thalamus in cynomolgus monkeys

The prefrontal cortex has been defined as that cortical territory that has "essential or sustaining" connections with the mediodorsal (MD) nucleus of the thalamus. However, recent studies in the monkey have documented projections from MD to the more caudal, agranular regions of the frontal cortex, suggesting that the connections of MD may be characterized by a breadth of distribution and diversity of functional roles too great to be useful as a unifying and defining feature for a specific cortical territory. In this study, we placed tracer injections in the lateral divisions of MD in cynomolgus monkeys (Macaca fascicularis) to assess the relative proportions of connections devoted to diverse regions of the frontal cortex (FC). Three different patterns of label were observed in the cortex, associated with different locations within lateral MD. We have designated these as the ventrolateral MD-arcuate FC circuit, having most label in areas 8 and 6; the caudoventral MD-dorsomedial FC circuit, having most label in areas 24 and presupplementary motor area (SMA); and the anterodorsal MD-anterior FC circuit, with the most label in areas 9, 46, 12, and 10. Only two of the nine cases injected in lateral MD were predominantly connected with the anterior FC. Thus, particular locales within lateral MD are connected with multiple, functionally diverse cortical regions, including several not classically recognized as "prefrontal" areas. This divergence may distinguish MD-frontocortical and reciprocal corticothalamic pathways from the largely segregated pathways arising from the other thalamic nuclei that are interconnected with the frontal cortex, such as those from the ventrolateral nuclear group.     

Reduced cholinergic olfactory centrifugal inputs in patients with neurodegenerative disorders and MPTP-treated monkeys

Olfactory impairment is a common feature of neurodegenerative diseases such as Parkinson’s disease (PD), Alzheimer’s disease (AD) and dementia with Lewy bodies (DLB). Olfactory bulb (OB) pathology in these diseases shows an increased number of olfactory dopaminergic cells, protein aggregates and dysfunction of neurotransmitter systems. Since cholinergic denervation might be a common underlying pathophysiological feature, the objective of this study was to determine cholinergic innervation of the OB in 27 patients with histological diagnosis of PD (n = 5), AD (n = 14), DLB (n = 8) and 8 healthy control subjects. Cholinergic centrifugal inputs to the OB were clearly reduced in all patients, the most significant decrease being in the DLB group. We also studied cholinergic innervation of the OB in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys (n = 7) and 7 intact animals. In MPTP-monkeys, we found that cholinergic innervation of the OB was reduced compared to control animals (n = 7). Interestingly, in MPTP-monkeys, we also detected a loss of cholinergic neurons and decreased dopaminergic innervation in the horizontal limb of the diagonal band, which is the origin of the centrifugal cholinergic input to the OB. All these data suggest that cholinergic damage in the OB might contribute, at least in part, to the olfactory dysfunction usually exhibited by these patients. Moreover, decreased cholinergic input to the OB found in MPTP-monkeys suggests that dopamine depletion in itself might reduce the cholinergic tone of basal forebrain cholinergic neurons.     

Task persistence and learning ability in normal and chronic low dose MPTP-treated monkeys

Monkeys exposed to low doses of the dopamine neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) develop cognitive deficits in the absence of gross motor dysfunction. Attentional deficits and task impersistence are now also described in these animals. The task impersistence correlated with no-response errors (i.e. errors of omission) on a delayed response task and improved with dopamine agonist therapy. In parallel studies, it was observed that there were significant differences in the ability of normal monkeys to learn to perform cognitive tasks. We found that monkeys classified as poor learners had similar deficits in task persistence as did MPTP-exposed monkeys, suggesting a relationship between poor cognitive performance and task impersistence in untreated as well as MPTP-treated monkeys. The possible significance of these results for two clinical disorders, early Parkinson's disease and attention deficit hyperactivity disorder is discussed. Cognitive and behavioral similarities between chronic low dose MPTP-treated monkeys, early Parksinson's disease patients and people with attention deficit hyperactivity disorder may suggest the existence of related pathophysiological mechanisms in these disorders.