Clinical and genetic analyses of a Swedish patient series diagnosed with ataxia.

Hereditary ataxia is a heterogeneous group of complex neurological disorders. Next-generation sequencing methods have become a great help in clinical diagnostics, but it may remain challenging to determine if a genetic variant is the cause of the patient's disease. We compiled a consecutive single-center series of 87 patients from 76 families with progressive ataxia of known or unknown etiology. We investigated them clinically and genetically using whole exome or whole genome sequencing. Test methods were selected depending on family history, clinical phenotype, and availability. Genetic results were interpreted based on the American College of Medical Genetics criteria. For high-suspicion variants of uncertain significance, renewed bioinformatical and clinical evaluation was performed to assess the level of pathogenicity. Thirty (39.5%) of the 76 families had received a genetic diagnosis at the end of our study. We present the predominant etiologies of hereditary ataxia in a Swedish patient series. In two families, we established a clinical diagnosis, although the genetic variant was classified as "of uncertain significance" only, and in an additional three families, results are pending. We found a pathogenic variant in one family, but we suspect that it does not explain the complete clinical picture. We conclude that correctly interpreting genetic variants in complex neurogenetic diseases requires genetics and clinical expertise. The neurologist's careful phenotyping remains essential to confirm or reject a diagnosis, also by reassessing clinical findings after a candidate genetic variant is suggested. Collaboration between neurology and clinical genetics and combining clinical and research approaches optimizes diagnostic yield.

Novel insights regarding the measurement properties of the SCOPA-AUT.

BACKGROUND: The Scale for Outcomes in Parkinson's disease for Autonomic symptoms (SCOPA-AUT) is an instrument intended to assess overall and domain-specific autonomic symptom burden. In this study the SCOPA-AUT is translated into Swedish and its measurement properties are assessed. METHODS: Following translation the SCOPA-AUT was field-tested regarding comprehensibility, relevance, and respondent burden (n = 20). It was then tested according to Rasch measurement theory using data from 242 persons with PD, of whom 162 completed SCOPA-AUT at baseline and 1-2 years later, giving a total of 404 data points for analysis. RESULTS: The Swedish SCOPA-AUT took a mean of 6 min to complete and was considered easy to use and relevant by respondents. SCOPA-AUT exhibited acceptable Rasch model fit, represents more severe levels of dysautonomia than that reported by the sample, and response categories were not working as expected for 17 items. Local dependency was identified and followed a pattern resembling the suggested subscales. Accounting for the subscale structure eliminated local dependency and reduced the initially inflated reliability from 0.81 to 0.68. CONCLUSIONS: The SCOPA-AUT is useful as a clinical check-list but requires further developmental work in order to meet more rigorous standards as an outcome measurement instrument.

The Orthostatic Hypotension Questionnaire in Swedish tested in patients with parkinsonism.

BACKGROUND: Orthostatic hypotension (OH) is common among older people and in particular in conditions like Parkinson's disease (PD). The OH Questionnaire (OHQ) has been proposed as a useful patient-reported assessment tool consisting of the OH Symptom Assessment (OHSA), OH Daily Activity Scale (OHDAS), and a composite score. AIMS OF THE STUDY: To translate the OHQ into Swedish and assess its psychometric properties. METHODS: Following forward-backward translation, the Swedish OHQ was field-tested (n = 6) for relevance, comprehensibility, and respondent burden. It was then tested regarding scaling assumptions, targeting, reliability, and construct validity in persons with PD (n = 27) and multiple system atrophy (n = 2). RESULTS: The Swedish OHQ was considered relevant and easy to use, with a mean completion time of 5.3 min. Scaling assumptions were acceptable for OHSA and OHDAS (corrected item-total correlations, .30-.67) but not for the total score (.12-.69). Floor/ceiling effects were ≤3.4% and reliability was >.64. Construct validity was supported by expected correlations with the SCOPA-AUT, RAND-36, and blood pressure measurements. CONCLUSIONS: The Swedish OHQ was well received, and psychometric results suggest that the OHQ (particularly the OHDAS) is a useful tool for OH assessment in parkinsonian disorders. Further testing in larger samples is needed.

A Phase 2a Trial Investigating the Safety and Tolerability of the Novel Cortical Enhancer IRL752 in Parkinson's Disease Dementia.

BACKGROUND: IRL752 is a novel small-molecule compound that acts to regioselectively enhance norepinephrine, dopamine, and acetylcholine neurotransmission in the cerebral cortex. OBJECTIVE: The primary objective of the trial was to investigate the safety and tolerability of IRL752 in patients with Parkinson's disease and dementia. METHODS: Patients with Parkinson's disease and dementia were randomized to IRL752 or placebo treatment (3:1 ratio) for 28 days. The study drug was given as an adjunct treatment to the patients' regular stable antiparkinsonian medication. Dosing was individually titrated for 14 days after which the dose was kept stable for an additional 14 days. RESULTS: A total of 32 patients were randomized to treatment, and 29 patients completed the 4-week treatment. Adverse events were generally mild and transient and were mostly reported during the dose titration phase. There were 2 serious adverse events, and none of them were related to the experimental treatment. The average dose achieved in the stable dose phase was 600 mg daily, yielding a 2-hour postdose plasma concentration of about 4 µM on day 28. Exploratory assessment of secondary outcomes indicated efficacy for symptoms and signs known to be poorly responsive to levodopa. CONCLUSIONS: IRL752 appears to be safe and well tolerated for a 4-week treatment in patients with Parkinson's disease and dementia. © 2020 International Parkinson and Movement Disorder Society.

Myoclonus-dystonia (DYT11, DYT-SGCE) - a channelopathy?

INTRODUCTION: Kaczynska et al. reported a family with myoclonus-dystonia (M-D) caused by a truncating SGCE mutation, in which two members had epilepsy. Further, patients had mild psychiatric and developmental deficits. CLINICAL REFLECTIONS: Characteristic motor features of M-D include myoclonus, dystonia and tremor. A wide range of additional disease manifestations are known. A few patients with M-D have seizures. CLINICAL IMPLICATIONS: Altered neuronal excitability has been found in the pathogenesis of M-D. This may explain the partial effectiveness of antiepileptics and a lower seizure threshold, and could encourage trials of other membrane stabilisers. Careful clinical observations of seemingly well-known diseases remain important.

Eltoprazine counteracts l-DOPA-induced dyskinesias in Parkinson's disease: a dose-finding study.

In advanced stages of Parkinson's disease, serotonergic terminals take up L-DOPA and convert it to dopamine. Abnormally released dopamine may participate in the development of L-DOPA-induced dyskinesias. Simultaneous activation of 5-HT1A and 5-HT1B receptors effectively blocks L-DOPA-induced dyskinesias in animal models of dopamine depletion, justifying a clinical study with eltoprazine, a 5-HT1A/B receptor agonist, against L-DOPA-induced dyskinesias in patients with Parkinson's disease. A double-blind, randomized, placebo-controlled and dose-finding phase I/IIa study was conducted. Single oral treatment with placebo or eltoprazine, at 2.5, 5 and 7.5 mg, was tested in combination with a suprathreshold dose of L-DOPA (Sinemet®) in 22 patients with Parkinson's disease (16 male/six female; 66.6 ± 8.8 years old) with L-DOPA-induced dyskinesias. A Wilcoxon Signed Ranked Test was used to compare each eltoprazine dose level to paired randomized placebo on the prespecified primary efficacy variables; area under the curve scores on Clinical Dyskinesia Rating Scale for 3 h post-dose and maximum change of Unified Parkinson's Disease Rating Scale part III for 3 h post-dose. Secondary objectives included effects on maximum Clinical Dyskinesia Rating Scale score, area under the curve of Rush Dyskinesia Rating Scale score for 3 h post-dose, mood parameters measured by Hospital Anxiety Depression Scale and Montgomery Asberg Depression Rating Scale along with the pharmacokinetics, safety and tolerability profile of eltoprazine. A mixed model repeated measures was used for post hoc analyses of the area under the curve and peak Clinical Dyskinesia Rating Scale scores. It was found that serum concentrations of eltoprazine increased in a dose-proportional manner. Following levodopa challenge, 5 mg eltoprazine caused a significant reduction of L-DOPA-induced dyskinesias on area under the curves of Clinical Dyskinesia Rating Scale [-1.02(1.49); P = 0.004] and Rush Dyskinesia Rating Scale [-0.15(0.23); P = 0.003]; and maximum Clinical Dyskinesia Rating Scale score [-1.14(1.59); P = 0.005]. The post hoc analysis confirmed these results and also showed an antidyskinetic effect of 7.5 mg eltoprazine. Unified Parkinson's Disease Rating Scale part III scores did not differ between the placebo and eltoprazine treatments. The most frequent adverse effects after eltoprazine were nausea and dizziness. It can be concluded that a single dose, oral treatment with eltoprazine has beneficial antidyskinetic effects without altering normal motor responses to L-DOPA. All doses of eltoprazine were well tolerated, with no major adverse effects. Eltoprazine has a favourable risk-benefit and pharmacokinetic profile in patients with Parkinson's disease. The data support further clinical studies with chronic oral eltoprazine to treat l-DOPA-induced-dyskinesias.

Autosomal dominant cerebellar ataxia with slow ocular saccades, neuropathy and orthostatism: a novel entity?

BACKGROUND: We describe the clinical characteristics of a Swedish family with autosomal dominant cerebellar ataxia, sensory and autonomic neuropathy, additional neurological features and unknown genetic cause. METHODS: Fourteen affected family members were identified. Their disorder was characterized by neurological examination, MRI, electroneurography, electromyography, MIBG-scintigraphy, and tilt-testing. RESULTS: The disorder presented as a balance and gait disturbance starting between 16 and 47 years of age. Cerebellar ataxia progressed slowly over the course of decades, and MRI showed mild to moderate cerebellar atrophy. Sensory axonal polyneuropathy was the most prominent additional feature and occurred in all patients examined. Autonomic neuropathy caused pronounced orthostatic dysregulation in at least four patients. Several affected members showed muscle wasting, and mild upper or lower motor neuron signs were documented. Patients had no nystagmus but slow or hypometric horizontal saccades and ocular motor apraxia. Cognition remained unimpaired, and there were no non-neurological disease manifestations. The disorder affected men and women in successive generations in a pattern compatible with autosomal dominant inheritance without evidence of anticipation. A second family where 7 members had very similar symptoms was identified and its origin traced back to the same village in southern Sweden as that of the first family's ancestors. All relevant known genetic causes of cerebellar ataxia were excluded by a novel next-generation sequencing approach. CONCLUSION: We present two probably related Swedish families with a characteristic and novel clinical syndrome of cerebellar ataxia and sensory polyneuropathy. The study serves as a basis for the mapping of the underlying genetic cause.

Development and testing of a self administered version of the Freezing of Gait Questionnaire.

BACKGROUND: The Freezing of Gait Questionnaire (FOGQ) was developed in response to the difficulties of observing and quantifying freezing of gait (FOG) clinically as well as in laboratory settings. However, as the FOGQ is a clinician-administered patient-reported rating scale it cannot be used in postal surveys. Here we report the development and measurement properties of a self-administered version of the FOGQ (FOGQsa). METHODS: A clinical sample and a postal survey sample of non-demented people with Parkinson's disease (PD; total n = 225) completed the FOGQsa and questionnaires concerning physical functioning (PF) and fall-related self efficacy (FES). Additional questions (No/Yes) regarded previous falls and whether they were afraid of falling. The clinical sample was also assessed with the Unified PD Rating Scale (UPDRS). Thirty-five participants completed FOGQsa and were also assessed with the original version (FOGQ) in a clinical interview. RESULTS: There were no differences (P = 0.12) between FOGQ (median, 10; q1-q3, 2-14) and FOGQsa (median, 8; 2-14) scores. The Spearman (rs) and intra-class correlations between the two were 0.92 and 0.91 (95% CI, 0.82-0.95), respectively. For FOGQsa, corrected item-total correlations ranged between 0.68-0.89. Reliability was 0.93 (95% CI, 0.91-0.94). FOGQsa scores correlated strongest with UPDRS Item 14 (Freezing; rs, 0.76) and with FES (rs, -0.74). The weakest correlation was found with age (rs, 0.14). Fallers scored significantly (p < 0.001) higher on FOGQsa compared to non-fallers, median scores 8 (q1-q3, 4-14) versus 2 (0-7). Those expressing a fear of falling scored higher (p < 0.001) than those who did not, median scores 2 (0-7) versus 6 (2-14). CONCLUSIONS: The present findings indicate that the FOGQsa is as reliable and valid as the original interview administered FOGQ version. This has important clinical implications when investigating FOG in large scale studies.

Neuronal differentiation following transplantation of expanded mouse neurosphere cultures derived from different embryonic forebrain regions.

In vitro, expanded neurospheres exhibit multipotent properties and can differentiate into neurons, astrocytes and oligodendrocytes. In vivo, cells from neurospheres derived from mouse fetal forebrain have previously been reported to predominantly differentiate into glial cells, and not into neurons. Here we isolated stem/progenitor cells from E13.5 lateral ganglionic eminence (LGE), medial ganglionic eminence (MGE) and cortical primordium, of a green fluorescent protein (GFP)-actin transgenic mouse. Free-floating neurospheres were expanded in the presence of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) and implanted after five to six passages into the striatum, hippocampus and cortex of neonatal rats. Cell suspensions of primary LGE tissue were prepared and grafted in parallel. Grafted cells derived from the primary tissue displayed widespread incorporation into all regions, as visualized with the mouse-specific antibody M2, or mouse satellite DNA in situ hybridization, and differentiated into both neurons, astrocytes and oligodendrocytes. Grafts of neurosphere cells derived from the LGE, MGE and cortical primordium differentiated primarily into astrocytes, but contained low but significant numbers of GFP-immunoreactive neurons. Neurons derived from LGE neurospheres were of three types: cells with the morphology of medium-sized densely spiny projection neurons in the striatum; cells with interneuron-like morphologies in striatum, cortex and hippocampus; and cells integrating into SVZ and migrating along the RMS to the olfactory bulb. MGE- or cortical primordium-derived neurospheres differentiated into interneuron-like cells in both striatum and hippocampus. The results demonstrate the ability of in vitro expanded neural stem/progenitor cells to generate both neurons and glia after transplantation into neonatal recipients, and differentiate in a region-specific manner into mature neurons with morphological features characteristic for each target site.

Neuronal and glial differentiation within expanded glial cultures derived from the lateral and medial ganglionic eminences.

Attached glial-like cell cultures were established from the lateral and medial ganglionic eminences (LGE and MGE) and from the neocortex (Cx) of E13.5 mouse embryos, and expanded over four to five passages under epidermal growth factor (EGF) stimulation. Following removal of EGF and serum, we analysed the generation of neurons and glial cells within the cultures. Significant numbers of betaIII-tubulin-positive neurons were generated in both the LGE (about 7% of total cell numbers) and the MGE (around 2%). However, only few betaIII-tubulin-positive cells with neuronal morphologies were detected in the differentiated Cx cultures. The newly formed neurons were to a large extent GABAergic, and many of the MGE-derived, but not the LGE-derived, cells expressed the MGE-marker NKX2.1. Most cells in all cultures still appeared astroglial-like, expressing glial fibrillary acidic protein (GFAP), but in addition, CNPase-positive cells with oligodendroglial morphologies were present in the MGE (0.68%), and, to a lesser extent (0.2%), in the LGE cultures. The present results demonstrate that cells of expanded glial cultures from both the LGE and MGE can give rise to significant and, to a certain extent, region-specific neuronal and glial cell types under differentiating conditions.

Grafted neural stem cells develop into functional pyramidal neurons and integrate into host cortical circuitry.

In vitro expanded neural stemprogenitor cells can undergo region-specific differentiation after transplantation to the developing or adult brain, and display morphologies and markers characteristic of mature neurons. Here we have used patch-clamp techniques to explore whether grafted stem cells also can develop physiological properties of mature neurons and become functionally integrated within host neural circuitry. The immortalized neural progenitor cell line, RN33B, prelabeled with GFP by using a lentiviral vector, was transplanted into the cortex or hippocampus of neonatal rats. We found that the grafted GFP-positive cells differentiated into cells with morphological features of cortical or hippocampal pyramidal neurons, and that many of them had established appropriate cortico-thalamic and contralateral hippocampal connections, respectively, as revealed by retrograde tracing. Whole-cell patch-clamp recordings from grafted cells with morphological characteristics of pyramidal neurons showed that they were able to generate action potentials, and received functional excitatory and inhibitory synaptic inputs from neighboring cells. These data provide evidence that grafted neural progenitors can differentiate into morphologically mature pyramidal projection neurons, establish appropriate long-distance axonal projections, exhibit normal electrophysiological properties, and become functionally integrated into host cortical circuitry.

Subretinal transplantation of brain-derived precursor cells to young RCS rats promotes photoreceptor cell survival.

The potential use of in vitro-expanded precursor cells or cell lines in brain repair includes transplantation of such cells for cell replacement purposes and the activation of host cells to provide 'self-repair'. Recently, it has been reported that the immortalized brain-derived cell line RN33B (derived from the embryonic rat medullary raphe) survive, integrate and differentiate after subretinal grafting to normal adult rats. Here, it is demonstrated that grafts of these cells survive for at least 6 weeks after implantation into postnatal days 21 and 35 retinas of normal and Royal College of Surgeons rats, a model of retinal degeneration. Implanted cells integrate into the retinal pigment epithelium and the inner retinal layers, and the anterior part of the optic nerve of both normal and Royal College of Surgeons rats. The RN33B cells migrate within the retina, occupying the whole retina from one eccentricity to the other. A significant number of the grafted cells differentiate into glial cells, as shown by the double labelling of the reporter genes LacZ or green fluorescent protein, with several glial markers, including oligodendrocytic markers. Many implanted cells in the host retina were in a proliferative stage judging from proliferative cell nuclear antigen and SV40 large T-antigen immunohistochemistry. Interestingly, there was a promotion of photoreceptor survival, extending over more than 2/3 of the superior hemisphere, in Royal College of Surgeons rats transplanted at postnatal day 21, but not at postnatal day 35. In addition, grafted cells were found in the surviving photoreceptor layer in these rats.

Differentiation of the RN33B cell line into forebrain projection neurons after transplantation into the neonatal rat brain.

The rat neural cell line RN33B has a remarkable ability to undergo region-specific neuronal differentiation after transplantation into the CNS. To further study its neurogenic properties in vivo, we used a recombinant lentiviral vector to genetically label the cells with the Green Fluorescent Protein (GFP) gene before implantation into the striatum/cortex, hippocampus, or mesencephalon of newborn rats. Three weeks after implantation, about 1-2% of the GFP-expressing cells had developed morphologies typical of neurons, astrocytes, or oligodendrocytes, the rest remained as either immature or undifferentiated nestin-positive cells. At 15-17 weeks postgrafting, the immature cells had disappeared in most graft recipients and only cells with neuronal or glial morphologies remained in similar numbers as at 3 weeks. The GFP distributed throughout the expressing cells, revealing fine morphological details, including dendrites with spines and extensive axonal projections. In all forebrain regions, the grafted cells differentiated into neurons with morphologies characteristic for each site, including large numbers of pyramidal-like cells in the cortex and the hippocampus, giving rise to dense projections to normal cortical target regions and to the contralateral hippocampus, respectively. In lower numbers, it was also possible to identify GFP-positive granulelike cells in the hippocampus, as well as densely spiny neurons in the striatum. In the mesencephalon by contrast, cells with astrocytic features predominated. The ability of the grafted RN33B cells to undergo region-specific differentiation into highly specialized types of forebrain projection neurons and establish connections with appropriate targets suggests that cues present in the microenvironment of the neonatal rat brain can effectively guide the development of immature progenitors, also in the absence of ongoing neurogenesis.

Migration patterns and phenotypic differentiation of long-term expanded human neural progenitor cells after transplantation into the adult rat brain.

We have examined long-term growth-factor expanded human neural progenitors following transplantation into the adult rat brain. Cells, obtained from the forebrain of a 9-week old fetus, propagated in the presence of epidermal growth factor, basic fibroblast growth factor, and leukemia inhibitory factor were transplanted into the striatum, subventricular zone (SVZ), and hippocampus. At 14 weeks, implanted cells were identified using antisera recognizing human nuclei and the reporter gene green fluorescent protein. Different migration patterns of the grafted cells were observed: (i) target-directed migration of doublecortin (DCX, a marker for migrating neuroblasts)-positive cells along the rostral migratory stream to the olfactory bulb and into the granular cell layer following transplantation into the SVZ and hippocampus, respectively; (ii) non-directed migration of DCX-positive cells in the grey matter in striatum and hippocampus, and (iii) extensive migration of above all nestin-positive/DCX-negative cells within white matter tracts. At the striatal implantation site, neuronal differentiation was most pronounced at the graft core with axonal projections extending along the internal capsule bundles. In the hippocampus, cells differentiated primarily into interneurons both in the dentate gyrus and in the CA1-3 regions as well as into granule-like neurons. In the striatum and hippocampus, a significant proportion of the grafted cells differentiated into glial cells, some with long processes extending along white matter tracts. Although the survival time was over 3 months in the present study a large fraction of the grafted cells remained undifferentiated in a stem or progenitor cell stage as revealed by the expression of nestin and/or GFAP.

Transplantation of human neural progenitor cells into the neonatal rat brain: extensive migration and differentiation with long-distance axonal projections.

Here we examined the ability of human neural progenitors from the embryonic forebrain, expanded for up to a year in culture in the presence of growth factors, to respond to environmental signals provided by the developing rat brain. After survival times of up to more than a year after transplantation into the striatum, the hippocampus, and the subventricular zone, the cells were analyzed using human-specific antisera and the reporter gene green fluorescent protein (GFP). From grafts implanted in the striatum, the cells migrated extensively, especially within white matter structures. Neuronal differentiation was most pronounced at the striatal graft core, with axonal projections extending caudally along the internal capsule into mesencephalon. In the hippocampus, cells migrated throughout the entire hippocampal formation and into adjacent white matter tracts, with differentiation into neurons both in the dentate gyrus and in the CA1-3 regions. Directed migration along the rostral migratory stream to the olfactory bulb and differentiation into granule cells were observed after implantation into the subventricular zone. Glial differentiation occurred at all three graft sites, predominantly at the injection sites, but also among the migrating cells. A lentiviral vector was used to transduce the cells with the GFP gene prior to grafting. The reporter gene was expressed for at least 15 weeks and the distribution of the gene product throughout the entire cytoplasmic compartment of the expressing cells allowed for a detailed morphological analysis of a portion of the grafted cells. The extensive integration and differentiation of in vitro-expanded human neural progenitor cells indicate that multipotent progenitors are capable of responding in a regionally specific manner to cues present in the developing rat brain.

Ex vivo and in vitro studies of transgene expression in rat astrocytes transduced with lentiviral vectors.

Implantation of cells genetically modified to express therapeutic genes into the brain has been proposed as a potential treatment for neurodegenerative diseases. In the current study embryonic rat-derived astrocytes were cultured and transduced with a lentiviral vector expressing the reporter gene green fluorescent protein (GFP) and subsequently grafted into the adult rat brain. The proportion of GFP expressing cells was stable, albeit small (1%), at all survival times, up to 6 weeks, the longest time point studied. In parallel in vitro studies, the astrocytes were lentivirally transduced to express either one of the two isoforms of glutamate decarboxylase (GAD(65) or GAD(67)) or glial cell line-derived neurotrophic factor (GDNF). When transducing 293T cells with the two GAD vectors, released GABA could be measured using high-performance liquid chromatography. Further studies of rat astrocytes transduced with the same vectors resulted in a level of GAD activity about 10 times higher than the activity of an intact rat striatum. One hundred thousand astrocytes transduced with LV-GDNF released approximately 27 ng of GDNF per hour. Thus, taken together, our observations provide support for the use of rat astrocytes in ex vivo gene transfer of these proteins in animal models of CNS disorders, e.g., Parkinson's disease or epilepsy.

Striatal c-fos Induction by Cocaine or Apomorphine Occurs Preferentially in Output Neurons Projecting to the Substantia Nigra in the Rat.

Fluorogold or rhodamine-labelled latex beads were injected in the substantia nigra (SN) or the globus pallidus (GP) in order retrogradely to label striatal output neurons that project to the two target structures. Ten days later, striatal c-fos was induced by systemic administration of cocaine (five normal rats; 25 mg/kg cocaine i.p. 2 h before killing) or apomorphine (five unilaterally dopamine-denervated rats; 0.25 mg/kg apomorphine s. c. 2 h before killing), and detection of the Fos protein in the striatum was achieved by immunofluorescence. Sections through the caudate-putamen that displayed good labelling from both SN and GP were selected for a quantitative analysis: the number of retrogradely labelled cells that exhibited Fos immunoreactivity, as well as the total number of retrogradely labelled cells located within a grid (0.16 mm2 in size) were counted manually at 25 x magnification. Cocaine induced a proportionally higher c-fos expression in striato-nigral compared to striato-pallidal neurons, whereas apomorphine activated Fos almost exclusively in striato-nigral neurons. The present findings are consistent with the idea that striatal c-fos induction by dopaminergic agents is primarily mediated by an interaction with D1-receptors, which are thought to be selectively localized on neurons projecting to SN.