Identification of the Neural Correlates Underlying Conflict Resolution Performance Using a Rodent Analogue of the Stroop Tests.

The Stroop test is a widely used neuropsychological test measuring attention and conflict resolution, which shows sensitivity across a range of diseases, including Alzheimer's, Parkinson's and Huntington's diseases. A rodent analogue of the Stroop test, the Response-Conflict task (rRCT), allows for systematic investigation of the neural systems underpinning performance in this test. Little is known about the involvement of the basal ganglia in this neural process. The aim of this study was to use the rRCT to determine whether striatal subregions are recruited during conflict resolution processing. To achieve this, rats were exposed to Congruent or Incongruent stimuli in the rRCT and the expression patterns of the immediate early gene Zif268 were analysed throughout cortical, hippocampal and basal ganglia subregions. The results confirmed the previously reported involvement of prefrontal cortical and hippocampal regions, as well as identifying a specific role for the dysgranular (but not granular) retrosplenial cortex in conflict resolution. Finally, performance accuracy correlated significantly with reduced neural activation in the dorsomedial striatum. Involvement of the basal ganglia in this neural process has not previously been reported. These data demonstrate that the cognitive process of conflict resolution requires not only prefrontal cortical regions, but also recruits the dysgranular retrosplenial cortex and the medial region of the neostriatum. These data have implications for understanding the neuroanatomical changes that underpin impaired Stroop performance in people with neurological disorders.

Alterations in the metabolic and cardiorespiratory response to exercise in Huntington's Disease.

BACKGROUND: Limited data suggests that an altered metabolic and cardiorespiratory exercise response may affect exercise performance in individuals with Huntington's disease (HD). There is no clear exploration of the response in individuals at different stages of the disease or in relation to genetic markers. This study aimed to examine the exercise response and recovery of HD participants, and the relationship to genetic and clinical markers. METHOD: HD gene-positive participants (n = 31; 9 pre-manifest; 22 manifest HD) and a healthy control group (n = 29) performed an incremental exercise test until exhaustion. Performance, cardiorespiratory, metabolic and perceptual responses to exercise were determined from a maximal cycle ergometer test throughout the exercise test and during a recovery period. RESULTS: During sub-maximal exercise, metabolic (lactate levels, oxygen uptake) and cardiorespiratory markers (heart rate) were elevated in HD participants compared to controls. Lactate elevation was specific to pre-manifest HD participants. Work capacity was reduced in both pre-manifest and manifest HD participants with tests terminated with no difference in metabolic, perceptual or cardiorespiratory markers. Submaximal oxygen uptake was correlated with motor score, whilst peak measures were unrelated to genetic or clinical markers. Heart rate recovery was attenuated in pre-manifest and manifest HD participants. CONCLUSIONS: Our findings confirm metabolic and cardiorespiratory deficits reduce exercise performance and affect recovery from an early stage in HD, with submaximal deficits related to phenotypic expression. Exercise capacity appears to be limited by an altered movement economy, thus clinicians should consider an altered exercise response and recovery may affect prescription in HD.

Clinical characterization of dystonia in adult patients with Huntington's disease.

BACKGROUND AND PURPOSE: Huntington's disease (HD) is an autosomal dominant, neurodegenerative movement disorder, typically characterized by chorea. Dystonia is also recognized as part of the HD motor phenotype, although little work detailing its prevalence, distribution, severity and impact on functional capacity has been published to date. METHODS: Patients (>18 years of age) were recruited from the Cardiff (UK) HD clinic, each undergoing a standardized videotaped clinical examination and series of functional assessment questionnaires (Unified Huntington's Disease Rating Scale, Burke-Fahn-Marsden Dystonia Rating Scale and modified version of the Toronto Western Spasmodic Torticollis Rating Scale). The presence and severity of dystonia were scored by four independent neurologists using the Burke-Fahn-Marsden Dystonia Rating Scale and Unified Huntington's Disease Rating Scale. Statistical analysis included Fisher's exact test, Wilcoxon test, anova and calculation of correlation coefficients where appropriate. RESULTS: Forty-eight patients [91% (48/53)] demonstrated evidence of dystonia, with the highest prevalence in the left upper limb (n = 44, 83%), right upper limb most severely affected and eyes least affected. Statistically significant positive correlations (P < 0.05) were observed between dystonia severity and increasing HD disease stage and motor disease duration. Deterioration in functional capacity also correlated with increasing dystonia severity. No significant relationship was observed with age at motor symptom onset or CAG repeat length. CONCLUSIONS: We report a high prevalence of dystonia in adult patients with HD, with worsening dystonia severity with increasing HD disease stage and motor disease duration. The recognition and management of dystonic symptoms in routine clinical practice will aid overall symptomatic treatment and functional improvement.

FoxP1 marks medium spiny neurons from precursors to maturity and is required for their differentiation.

Identifying the steps involved in striatal development is important both for understanding the striatum in health and disease, and for generating protocols to differentiate striatal neurons for regenerative medicine. The most prominent neuronal subtype in the adult striatum is the medium spiny projection neuron (MSN), which constitutes more than 85% of all striatal neurons and classically expresses DARPP-32. Through a microarray study of genes expressed in the whole ganglionic eminence (WGE: the developing striatum) in the mouse, we identified the gene encoding the transcription factor Forkhead box protein P1 (FoxP1) as the most highly up-regulated gene, thus providing unbiased evidence for the association of FoxP1 with MSN development. We also describe the expression of FoxP1 in the human fetal brain over equivalent gestational stages. FoxP1 expression persisted through into adulthood in the mouse brain, where it co-localised with all striatal DARPP-32 positive projection neurons and a small population of DARPP-32 negative cells. There was no co-localisation of FoxP1 with any interneuron markers. FoxP1 was detectable in primary fetal striatal cells following dissection, culture, and transplantation into the adult lesioned striatum, demonstrating its utility as an MSN marker for transplantation studies. Furthermore, DARPP-32 expression was absent from FoxP1 knock-out mouse WGE differentiated in vitro, suggesting that FoxP1 is important for the development of DARPP-32-positive MSNs. In summary, we show that FoxP1 labels MSN precursors prior to the expression of DARPP-32 during normal development, and in addition suggest that FoxP1 labels a sub-population of MSNs that are not co-labelled by DARPP-32. We demonstrate the utility of FoxP1 to label MSNs in vitro and following neural transplantation, and show that FoxP1 is required for DARPP-32 positive MSN differentiation in vitro.

Amelioration of non-motor dysfunctions after transplantation of human dopamine neurons in a model of Parkinson's disease.

BACKGROUND: Patients suffering from Parkinson's disease (PD) display cognitive and neuropsychiatric dysfunctions, especially with disease progression. Although these impairments have been reported to impact more heavily upon a patient's quality of life than any motor dysfunctions, there are currently no interventions capable of adequately targeting these non-motor deficits. OBJECTIVES: Utilizing a rodent model of PD, we investigated whether cell replacement therapy, using intrastriatal transplants of human-derived ventral mesencephalic (hVM) grafts, could alleviate cognitive and neuropsychiatric, as well as motor, dysfunctions. METHODS: Rats with unilateral 6-hydroxydopamine lesions to the medial forebrain bundle were tested on a complex operant task that dissociates motivational, visuospatial and motor impairments sensitive to the loss of dopamine. A subset of lesioned rats received intrastriatal hVM grafts of ~9 weeks gestation. Post-graft, rats underwent repeated drug-induced rotation tests and were tested on two versions of the complex operant task, before post-mortem analysis of the hVM tissue grafts. RESULTS: Post-graft behavioural testing revealed that hVM grafts improved non-motor aspects of task performance, specifically visuospatial function and motivational processing, as well as alleviating motor dysfunctions. CONCLUSIONS: We report the first evidence of human VM cell grafts alleviating both non-motor and motor dysfunctions in an animal model of PD. This intervention, therefore, is the first to improve cognitive and neuropsychiatric symptoms long-term in a model of PD.

Cross-regulation of Connexin43 and ß-catenin influences differentiation of human neural progenitor cells.

Connexin43 (Cx43) is the most widely and abundantly expressed gap junction (GJ) protein and it is strongly associated with the regulation of cell cycle progression. Emerging roles for Cx43 in cell adhesion and migration during neural differentiation have also been recently recognized, and this has emphasized the involvement of Cx43 in different physiological process beyond its role as a GJ protein. In this study, we explore the function of Cx43 in the differentiation of human neural progenitor cells (hNPCs) using viral vectors that mediate the overexpression or knockdown of the protein. Results showed that in the absence of this protein fetal cortex-derived hNPCs differentiated toward a neuronal phenotype at expenses of a glial phenotype. Furthermore, the silencing of Cx43 did not affect hNPC proliferation rate or numbers of apoptotic cells. The increase in the number of neurons was not recapitulated when GJ intercellular communications were pharmacologically blocked, and this suggested that Cx43 was influencing hNPCs differentiation with a GJ-independent effect. In addition, Cx43 knockdown significantly increased ß-catenin signaling, which has been shown to regulate the transcription of pro-neuronal genes during embryonic neural development. Our results add further support to the hypothesis that Cx43 protein itself regulates key signaling pathways during development and neurogenesis beyond its role as GJ protein.

The lateral neostriatum is necessary for compensatory ingestive behaviour after intravascular dehydration in female rats.

Aberrant striatal function results in an array of physiological symptoms, including impaired consummatory and regulatory behaviours, which can lead to weight loss and dehydration. It was hypothesised, therefore, that cell loss in the neostriatum may contribute to altered fluid intake by regulating physiological signals related to dehydration status. To test this theory, rats with lesions of the lateral neostriatum and sham controls underwent a series of physiological challenges, including the experimental induction of intracellular and intravascular dehydration. No baseline differences in prandial or non-prandial drinking were observed, nor were differences in locomotor activity evident between groups. Furthermore, intracellular dehydration increased water intake in lesion rats in a manner comparable to sham rats. Interestingly, a specific impairment was evident in lesion rats after subcutaneous injection of poly-ethylene glycol was used to induce intravascular dehydration, such that lesion rats failed to adapt their water intake to this physiological change. The results suggest that the striatal lesions resulted in regulatory dysfunction by impairing motivational control over compensatory ingestive behaviour after intravascular hydration, while the physiological signals related to dehydration remain intact. Loss of these cells in neurodegenerative disorders, such Huntington's disease, may contribute to regulatory changes evident in the course of the disease.

Molecular regulation of striatal development: a review.

The central nervous system is composed of the brain and the spinal cord. The brain is a complex organ that processes and coordinates activities of the body in bilaterian, higher-order animals. The development of the brain mirrors its complex function as it requires intricate genetic signalling at specific times, and deviations from this can lead to brain malformations such as anencephaly. Research into how the CNS is specified and patterned has been studied extensively in chick, fish, frog, and mice, but findings from the latter will be emphasised here as higher-order mammals show most similarity to the human brain. Specifically, we will focus on the embryonic development of an important forebrain structure, the striatum (also known as the dorsal striatum or neostriatum). Over the past decade, research on striatal development in mice has led to an influx of new information about the genes involved, but the precise orchestration between the genes, signalling molecules, and transcription factors remains unanswered. We aim to summarise what is known to date about the tightly controlled network of interacting genes that control striatal development. This paper will discuss early telencephalon patterning and dorsal ventral patterning with specific reference to the genes involved in striatal development.

Medical terminations of pregnancy: a viable source of tissue for cell replacement therapy for neurodegenerative disorders.

"Proof-of-principle" that cell replacement therapy works for neurodegeneration has been reported, but only using donor cells collected from fetal brain tissue obtained from surgical terminations of pregnancy. Surgical terminations of pregnancy represent an increasingly limited supply of donor cells due to the tendency towards performing medical termination in much of Europe. This imposes a severe constraint on further experimental and clinical cell transplantation research. Therefore, we explore here the feasibility of using medical termination tissue as a donor source. Products of conception were retrieved from surgical terminations over the last 7 years and from medical terminations over the last 2.5 years. The number of collections that yielded fetal tissue, viable brain tissue, and identifiable brain regions (ganglionic eminence, ventral mesencephalon, and neocortex) were recorded. We studied cell viability, cell physiological properties, and differentiation potential both in vitro and following transplantation into the central nervous system of rodent models of neurodegenerative disease. Within equivalent periods, we were able to collect substantially greater numbers of fetal remains from medical than from surgical terminations of pregnancy, and the medical terminations yielded a much higher proportion of identifiable and dissectible brain tissue. Furthermore, we demonstrate that harvested cells retain the capacity to differentiate into neurons with characteristics appropriate to the region from which they are dissected. We show that, contrary to widespread assumption, medical termination of pregnancy-derived fetal brain cells represent a feasible and more readily available source of human fetal tissue for experimental cell transplantation with the potential for use in future clinical trials in human neurodegenerative disease.

Clusterin secreted by astrocytes enhances neuronal differentiation from human neural precursor cells.

Neuronal differentiation from expanded human ventral mesencephalic neural precursor cells (NPCs) is very limited. Astrocytes are known to secrete neurotrophic factors, and so in order to enhance neuronal survival from NPCs, we tested the effect of regional astrocyte-conditioned medium (ACM) from the rat cortex, hippocampus and midbrain on this process. Human NPC's were expanded in FGF-2 before differentiation for 1 or 4 weeks in ACM. The results show that ACM from the hippocampus and midbrain increase the number of neurons from expanded human NPCs, an effect that was not observed with cortical ACM. In addition, both hippocampal and midbrain ACM increased the number and length of phosphorylated neurofilaments. MALDI-TOF analysis used to determine differences in media revealed that although all three regional ACMs had cystatin C, α-2 macroglobulin, extracellular matrix glycoprotein and vimentin, only hippocampal and midbrain ACM also contained clusterin, which when immunodepleted from midbrain ACM eliminated the observed effects on neuronal differentiation. Furthermore, clusterin is a highly glycosylated protein that has no effect on cell proliferation but decreases apoptotic nuclei and causes a sustained increase in phosphorylated extracellular signal-regulated kinase, implicating its role in cell survival and differentiation. These findings further reveal differential effects of regional astrocytes on NPC behavior and identify clusterin as an important mediator of NPC-derived neuronal survival and differentiation.

Challenges facing quantification of rat locomotion along beams of varying widths.

Optoelectronic motion capture systems have been widely used to investigate temporal gait parameters in humans and animals in order to understand function and behavioural attributes of different pathologies, e.g. Parkinson's disease (PD). The aim of the present paper was to investigate the practicality of utilising this system to investigate the effects of a unilateral 6-hydroxydopamine (6-OHDA) lesion on rat locomotion while walking on beams of varying widths (graduated, narrow, and wide). Temporal gait parameters of ten male Lister Hooded rats (five controls and five hemiparkinsonian) were observed using passive markers placed in locations that were representative of their four limbs and their body axis. The results demonstrate that marker-based motion capture can provide an effective and simple approach to quantifying temporal gait parameters for rat models of PD. They also reveal how the width of the path affects the locomotion in both experimental cohorts. Such measurements can be compared with human motion analysis to explore correlations between the animal model and human behaviour, which is an important step for translational medicine.

Human trials for neurodegenerative disease.

The lack of disease-modifying treatments currently available for not just some but most neurodegenerative diseases, including Parkinson's disease, Huntington's disease, and even stroke, helps explain increasing interest in cell-based therapies. One key aim of such treatment is to replace neurons or glia lost as a result of the disease, with a view to the cells integrating functionally within the host tissue in order to reconstruct neural circuitry. Clinical trials using primary human fetal tissue as a cell source commenced in Parkinson's disease (PD) in the 1980s; currently, comparable neural transplantation trials in Huntington's disease are underway. Disappointing results of later controlled trials in PD illustrated not least the vital importance of methodological issues relating to the structure and implementation of clinical trials, and these issues will be considered here in more depth.

Membrane permeability coefficients of murine primary neural brain cells in the presence of cryoprotectant.

Neural cells isolated from the brain have a number of research and clinical applications, including transplantation to patients with neurodegenerative conditions. Tissue supply is one of the major limiting factors to clinical transplantation. Cryopreservation of primary neural cells would improve supply, aid in organisation of transplantation surgery and facilitate research. To date, cryopreservation using standard methods has resulted in reduced yield and/or viability of primary neural tissue. In order to optimise freezing protocols specifically for such cells, the non-osmotic volume (V(b)), water permeability (L(p)) and permeability to cryoprotectant (P(cpa)) were determined. Murine foetal brain tissue from the ganglionic eminence (GE), ventral mesencephalon (VM), or neocortical mantle (Ctx) was trypsinised to a single cell suspension. To determine V(b,) cell volume was measured after exposure to anisotonic solutions of sucrose (150-1500 mOsmol/kg). L(p) (mum/min.atm) and P(cpa) (mum/s) were determined for GE cells by measuring cell volume during exposure to 1.5 mol/l cryoprotectant. Cell volume was determined using an electronic particle counting method. V(b) was 27% for Ctx and GE, and 30% for VM. The osmotic response of GE cells was similar in the presence of propane-1,2-diol and dimethyl sulphoxide. In the presence of ethylene glycol, cell volume decrease was greater on initial exposure to cryoprotectant and recovery slower. Differences in L(p,) but not P(cpa), were found between cryoprotectants. The present results provide key parameters for optimisation of freezing protocols for cryopreservation of primary foetal brain tissues for application in neural cell transplantation.

Mobility and falls in people with Huntington's disease.

OBJECTIVE: The aim of this study was to estimate the frequency of falls in people with Huntington's disease (HD) and make a preliminary assessment of tools appropriate for assessing the risk of falling. DESIGN: Observational study. SETTING: Hospital clinic. SUBJECTS: 24 people with HD. MAIN MEASURES: Balance was assessed using the Berg Balance Scale (BBS) and Timed "Up & Go" (TUG) test. Walking speed over 10 m was recorded. Long-term monitoring of walking activity was undertaken. Unified Huntington Disease Rating Scale (UHDRS) motor, Functional Assessment Scale (FAS), Independence Scale (IS) and Total Functional Capacity (TFC) scores were obtained as well as data about falls and stumbles. Differences between "recurrent fallers" (>or=2 falls/year) and "non-fallers" (or=2 falls in the previous 12 months. Recurrent fallers walked less (p<0.01) and slower than non-fallers. Their balance (BBS) (p<0.01) was worse and TUG scores were higher (p<0.01). People with HD had increased risk of falls if TUG scores were >or=14 s or BBS scores

Dentatorubral pallidoluysian atrophy in South Wales.

BACKGROUND: Dentatorubral pallidoluysian atrophy (DRPLA) is a rare, autosomal dominant, clinically heterogeneous neurodegenerative disorder characterised clinically by progressive dementia, ataxia, chorea, myoclonic epilepsy and psychiatric disturbance and pathologically by combined degeneration of the dentatorubral and pallidoluysian systems. DRPLA has a marked ethnic predilection, most commonly reported in Japan and thought to be rare in Caucasian populations. METHODS: We describe the clinical and genetic characteristics of 17 patients with DRPLA segregating in four families in South Wales. RESULTS: There was marked clinical heterogeneity with considerable overlap of symptoms and signs between and within families. The age of onset ranged from 34 to 60 years with an earlier onset associated with myoclonic epilepsy and a later onset associated with a Huntington disease-like presentation. We identified a distinct haplotype within one family not present within the other three families, suggesting that the expansion in at least one family did not arise from an immediate common ancestor. Analysis of repeat length polymorphisms in 306 Welsh control patients identified 14 (4.6%) with repeat lengths in the high-normal range, compared with 0% and 7.4% in previously reported north American Caucasian and Japanese control populations, respectively. CONCLUSIONS: DRPLA may not be as geographically or ethnically restricted as previously thought and the diagnosis should be considered in non-Asian patients presenting with a wide spectrum of neurological disease, especially if there is a dominant family history of dementia or movement disorder. The prevalence of high-normal length alleles may account for the relatively high prevalence of DRPLA in Wales.

Can directed activity improve mobility in Huntington's disease?

Huntington's disease is an inherited disorder of the CNS that results in progressive deterioration of mobility and cognition and also affects behaviour. There are no disease-modifying interventions available to date, although there has been considerable progress in research directed at understanding the pathological basis of the disease with a view to identifying potential treatments. It is however important not to overlook currently available treatment strategies, including rehabilitation approaches. There has been little work to date to explore the potential of such approaches and here we highlight the need for more systematic studies in this area as well as the need for good objective assessment tools and the potential role that rehabilitation and training may have in the application of novel treatment options.

Neural transplantation in Huntington's disease: the NEST-UK donor tissue microbiological screening program and review of the literature.

Neural transplantation of human fetal tissue for Huntington's disease (HD) is now entering the clinical arena. The safety of the procedure has now been demonstrated in a number of studies, although the efficacy of such an approach is still being investigated. Stringent but practicable screening of the donor tissue for potential pathogens is an essential prerequisite for successful implementation of any novel transplant program that uses human fetal tissue. In this article we summarize the UK-NEST protocol for the screening of human fetal tissue being grafted to patients with mild to moderate HD. We describe the results of microbiological screening of 87 potential tissue donors in a pilot study, and of the first four donor-recipient patients included in the UK-NEST series. The rationale for the adoption and interpretation of the various tests is described and our methodology is compared with those previously used by other centers. This article therefore presents a comprehensive, logical yet pragmatic screening program that could be employed in any clinical studies that use human fetal tissue for neurotransplantation.

Staging and preparation of human fetal striatal tissue for neural transplantation in Huntington's disease.

Transplantation of human fetal central nervous system tissue has been shown to be of benefit in Parkinson's disease, and is currently being explored as a therapeutic option in Huntington's disease. The success of a neural transplant is dependent on a number of factors, including the requirement that donor cells are harvested within a given developmental window and that the cell preparation protocols take account of the biological parameters identified in animal models. Although many of the criteria necessary for a successful neural transplant have been defined in animal models, ultimately they must be validated in human studies, and some issues can only ever be addressed in human studies. Furthermore, because neural transplantation of human fetal tissue is limited to small numbers of patients in any one surgical center, largely due to practical constraints, it is crucial that tissue preparation protocols are clearly defined and reproducible, so that (i) multicenter trials are possible and are based on consistent tissue preparation parameters, and (ii) results between centers can be meaningfully analyzed. Here we describe the preparation of human fetal striatum for neural transplantation in Huntington's disease, and report on the validation of a method for estimating the developmental stage of the fetus based on direct morphometric measurements of the embryonic tissue.

Unilateral transplantation of human primary fetal tissue in four patients with Huntington's disease: NEST-UK safety report ISRCTN no 36485475.

OBJECTIVES: Huntington's disease (HD) is an inherited autosomal dominant condition in which there is a CAG repeat expansion in the huntingtin gene of 36 or more. Patients display progressive motor, cognitive, and behavioural deterioration associated with progressive cell loss and atrophy in the striatum. Currently there are no disease modifying treatments and current symptomatic treatments are only partially effective in the early to moderate stages. Neural transplantation is effective in animal models of HD and offers a potential strategy for brain repair in patients. The authors report a safety study of unilateral transplantation of human fetal striatal tissue into the striatum of four patients with HD. SUBJECTS AND METHODS: Stereotaxic placements of cell suspensions of human fetal ganglionic eminence were made unilaterally into the striatum of four patients with early to moderate HD. All patients received immunotherapy with cyclosporin A, azathioprine, and prednisolone for at least six months postoperatively. Patients were assessed for safety of the procedure using magnetic resonance imaging (MRI), regular recording of serum biochemistry and haematology to monitor immunotherapy, and clinical assessment according to the Core Assessment Protocol For Intrastriatal Transplantation in HD (CAPIT-HD). RESULTS: During the six month post-transplantation period, the only adverse events related to the procedure were associated with the immunotherapy. MRI demonstrated tissue at the site of implantation, but there was no sign of tissue overgrowth. Furthermore, there was no evidence that the procedure accelerated the course of the disease. CONCLUSIONS: Unilateral transplantation of human fetal striatal tissue in patients with HD is safe and feasible. Assessment of efficacy will require longer follow up in a larger number of patients.

Cellular and molecular aspects of striatal development.

The striatum is a key component of the basal ganglia and there is considerable evidence that it has an important role in motor, cognitive and limbic functions. However, very little is known about how this forebrain structure develops. This review considers the role of cellular and molecular mechanisms involved in the development of the striatum, and the potential application of this knowledge to the understanding of the pathology and treatment of primary disease of this structure.