A longitudinal motor characterisation of the HdhQ111 mouse model of Huntington's Disease

Background: Huntington’s disease (HD) is a rare, incurable neurodegenerative disorder caused by a CAG trinucleotide expansion with the first exon of the huntingtin gene. Numerous knock-in mouse models are currently available for modelling HD. However, before their use in scientific research, these models must be characterised to determine their face and predictive validity as models of the disease and their reliability in recapitulating HD symptoms. Objective: Manifest HD is currently diagnosed upon the onset of motor symptoms, thus we sought to longitudinally characterise the progression and severity of motor signs in the HdhQ111 knock-in mouse model of HD, in heterozygous mice. Methods: An extensive battery of motor tests including: rotarod, inverted lid test, balance beam, spontaneous locomotor activity and gait analysis were applied longitudinally to a cohort of HdhQ111 heterozygous mice in order to progressively assess motor function. Results: A progressive failure to gain body weight was demonstrated from 11 months of age and motor problems in all measures of balance beam performance were shown in HdhQ111 heterozygous animals in comparison to wild type control animals from 9 months of age. A decreased latency to fall from the rotarod was demonstrated in HdhQ111 heterozygous animals in comparison to wild type animals, although this was not progressive with time. No genotype specific differences were demonstrated in any of the other motor tests included in the test battery. Conclusions: The HdhQ111 heterozygous mouse demonstrates a subtle and progressive motor phenotype that begins at 9 months of age. This mouse model represents an early disease stage and would be ideal for testing therapeutic strategies that require elongated lead-in times, such as viral gene therapies or striatal transplantation

Transplantation site influences the phenotypic differentiation of dopamine neurons in ventral mesencephalic grafts in Parkinsonian rats

Foetal midbrain progenitors have been shown to survive, give rise to different classes of dopamine neurons and integrate into the host brain alleviating Parkinsonian symptoms following transplantation in patients and animal models of the disease. Dopamine neuron subpopulations in the midbrain, namely A9 and A10, can be identified anatomically based on cell morphology and ascending axonal projections. G protein-gated inwardly rectifying potassium channel Girk2 and the calcium binding protein Calbindin are the two best available histochemical markers currently used to label (with some overlap) A9- and A10-like dopamine neuron subtypes, respectively, in tyrosine hydroxylase expressing neurons both in the midbrain and grafts. Both classes of dopamine neurons survive in grafts in the striatum and extend axonal projections to their normal dorsal and ventral striatal targets depending on phenotype. Nevertheless, grafts transplanted into the dorsal striatum, which is an A9 input nucleus, are enriched for dopamine neurons that express Girk2. It remains to be elucidated whether different transplantation sites favour the differential survival and/or development of concordant dopamine neuron subtypes within the grafts. Here we used rat foetal midbrain progenitors at two developmental stages corresponding to a peak in either A9 or A10 neurogenesis and examined their commitment to respective dopaminergic phenotypes by grafting cells into different forebrain regions that contain targets of either nigral A9 dopamine innervation (dorsal striatum), ventral tegmental area A10 dopamine innervation (nucleus accumbens and prefrontal cortex), or only sparse dopamine but rich noradrenaline innervation (hippocampus). We demonstrate that young (embryonic day, E12), but not older (E14),mesencephalic tissue and the transplant environment influence survival and functional integration of specific subtypes of dopamine neurons into the host brain. We also show that irrespective of donor age A9-like, Girk2-expressing neurons are more responsive to environmental cues in adopting a dopaminergic phenotype during differentiation postgrafting. These novel findings suggest that dopamine progenitors use targets of A9/A10 innervation in the transplantation site to complete maturation and the efficacy of foetal cell replacement therapy in patients may be improved by deriving midbrain tissue at earlier developmental stages than in current practice

Environmental housing and duration of exposure affect striatal graft morphology in a rodent model of Huntington's disease

Clinical trials of cell replacement therapy in Huntington's disease have shown its safety, feasibility, and potentially long-lasting effects. However, more needs to be known regarding the conditions that stimulate plasticity and compensation achieved by neural grafts to maximize posttransplantation recovery of such neurorehabilitative therapies. The effects of enriched environment (EE), behavioral experience, and transplantation can each separately influence neuronal plasticity and recovery of function after brain damage, and the mechanisms by which these factors interact to modify the survival, integration, or function of grafted tissues are at present unknown. To investigate the effects of variable housing conditions and duration on morphological and cellular changes within embryonic striatal transplants, rats received unilateral excitotoxic lesions of the striatum, followed by E15 whole-ganglionic eminence suspension grafts. The rats were divided into three groups according to housing: full-time EE, 1 h/day exposure to EE, or standard laboratory cages. The experimental design included “early” (7 weeks postgrafting) and “late” (13 weeks postgrafting) survival time points to explore the effects of exposure lengths to the three housing conditions. The morphological and cellular effects on the grafts were analyzed using immunohistochemistry, cell morphology, image analysis, and enzyme-linked immunoassay. Both the duration of the exposure and the housing conditions were seen to influence multiple parameters of grafted cell morphology. The factors acted either independently (e.g., on graft size), complementarily (e.g., on spine density), or had no distinctive effect (e.g., on lesion size) on graft development. Features of embryonic striatal grafts and their trophic milieu were influenced both by the complexity of the environmental conditions and by the length of exposure to them. The data suggest that neurorehabilitation should be a feature of clinical trials of cell transplantation in order to exploit the underlying mechanisms that promote anatomical integration of the grafted cells and maximize transplant-mediated functional recovery

Mechanisms and use of neural transplants for brain repair

Under appropriate conditions, neural tissues transplanted into the adult mammalian brain can survive, integrate, and function so as to influence the behavior of the host, opening the prospect of repairing neuronal damage, and alleviating symptoms associated with neuronal injury or neurodegenerative disease. Alternative mechanisms of action have been postulated: nonspecific effects of surgery; neurotrophic and neuroprotective influences on disease progression and host plasticity; diffuse or locally regulated pharmacological delivery of deficient neurochemicals, neurotransmitters, or neurohormones; restitution of the neuronal and glial environment necessary for proper host neuronal support and processing; promoting local and long-distance host and graft axon growth; formation of reciprocal connections and reconstruction of local circuits within the host brain; and up to full integration and reconstruction of fully functional host neuronal networks. Analysis of neural transplants in a broad range of anatomical systems and disease models, on simple and complex classes of behavioral function and information processing, have indicated that all of these alternative mechanisms are likely to contribute in different circumstances. Thus, there is not a single or typical mode of graft function; rather grafts can and do function in multiple ways, specific to each particular context. Consequently, to develop an effective cell-based therapy, multiple dimensions must be considered: the target disease pathogenesis; the neurodegenerative basis of each type of physiological dysfunction or behavioral symptom; the nature of the repair required to alleviate or remediate the functional impairments of particular clinical relevance; and identification of a suitable cell source or delivery system, along with the site and method of implantation, that can achieve the sought for repair and recovery

Cognitive training modifies disease symptoms in a mouse model of Huntington's disease

Huntington's disease (HD) is an incurable neurodegenerative disorder which causes a triad of motor, cognitive and psychiatric disturbances. Cognitive disruptions are a core feature of the disease, which significantly affect daily activities and quality of life, therefore cognitive training interventions present an exciting therapeutic intervention possibility for HD. We aimed to determine if specific cognitive training, in an operant task of attention, modifies the subsequent behavioural and neuropathological phenotype of the Hdh(Q111) mouse model of HD. Three testing groups comprising both Hdh(Q111) mice and wildtype controls were used. The first group received cognitive training in an operant task of attention at 4 months of age. The second group received cognitive training in a comparable non-attentional operant task at 4 months of age, and the third group were control animals that did not receive cognitive training. All groups were then tested in an operant task of attention at 12 months of age. Relative to naïve untrained mice, both wildtype and Hdh(Q111) mice that received cognitive training in the operant task of attention demonstrated an increased number of trials initiated, greater accuracy, and fewer ‘time out’ errors. A specific improvement in response time performance was observed in Hdh(Q111) mice, relative to naïve untrained Hdh(Q111) mice. Relative to the group that received comparable training in a non-attentional task, both wildtype and Hdh(Q111) mice that received attentional training demonstrated superior accuracy in the task and made fewer ‘time out’ errors. Despite significant behavioural change, in both wildtype and Hdh(Q111) mice that had received cognitive training, no significant changes in neuropathology were observed between any of the testing groups. These results demonstrate that attentional cognitive training implemented at a young age significantly improves attentional performance, at an older age, in both wildtype and Hdh(Q111) mice. Attentional cognitive training also improved motor performance in Hdh(Q111) mice, thus leading to the conclusion that cognitive training can improve disease symptoms in a mouse model of HD

Translation of cell therapies to the clinic: characteristics of cell suspensions in large diameter injection cannulae

With the use of cell replacement therapies as a realistic prospect for conditions such as Parkinson's and Huntington's diseases, the logistics of the delivery of cell suspensions to deep brain targets is a topic for consideration. Because of the large cannulae required for such procedures, we need to consider the behaviour of cell suspensions within the cannulae if we are to ensure that the injected cells are distributed as intended within the target tissue. We have investigated the behaviour of primary embryonic cell suspensions of neural tissue, in cannulae of different diameters, using a protocol designed to mimic the handling and injection of cells during clinical application. Internal cannula diameter had a large effect on the distribution of cells during their dispensation from the syringe. In vertical, or near vertical cannulae, cells settled towards the tip of the needle, and were dispensed unevenly, with the majority of cells emerging in the first 10-20% of the injectate. In horizontal or near-horizontal cannulae we observed the opposite effect, such that few cells were dispensed in the first 80% of the injectate, and the majority emerged in the final 10-20%. Use of a glass cannula showed that the results obtained using the horizontal cannula were caused by settling and adherence of the cells on the side of the cannulae, such that during dispensation, the overlying, cell-free solution was dispensed first, prior to the emergence of the cells. We show that the behaviour of cells in such cannulae is affected by the cannula diameter, and by the material of the cannula itself, and in horizontal cannulae, uneven expulsion of cells from the needle can be ameliorated by regular rotation of the cannula during the procedure. We discuss the potential impact of these observations on the translation of cell therapies to the clinic

SMAD transcription factors are altered in cell models of HD and regulate HTT expression

Transcriptional dysregulation is observable in multiple animal and cell models of Huntington's disease, as well as in human blood and post-mortem caudate. This contributes to HD pathogenesis, although the exact mechanism by which this occurs is unknown. We therefore utilised a dynamic model in order to determine the differential effect of growth factor stimulation on gene expression, to highlight potential alterations in kinase signalling pathways that may be in part responsible for the transcriptional dysregulation observed in HD, and which may reveal new therapeutic targets. We demonstrate that cells expressing mutant huntingtin have a dysregulated transcriptional response to epidermal growth factor stimulation, and identify the transforming growth factor-beta pathway as a novel signalling pathway of interest that may regulate the expression of the Huntingtin (HTT) gene itself. The dysregulation of HTT expression may contribute to the altered transcriptional phenotype observed in HD

Unilateral lesions of the dorsal striatum in rats disrupt responding in egocentric space

Rats were trained in a specially designed, multichoice operant chamber on a visual choice reaction time task designed to assess performance on each side of the rat’s body. The task required animals to sustain a nose poke in a central hole, until a brief light stimulus was presented in either of two holes that were located on the same side of the box. Once the rats were trained to perform the task to both sides independently they received unilateral injections of quinolinic acid into the dorsal striatum. Postoperatively, lesioned animals were impaired when performing the task on the side contralateral to the lesion. The time taken to initiate contralateral responses was increased. Contralateral responses were also exclusively biased toward the nearer of the two response locations, regardless of the location of the stimulus. This was interpreted as a specific impairment in generating responses in contralateral space. In contrast, no comparable deficit was seen when the animals performed the task on the side ipsilateral to the lesion. Additional postoperative challenges, in which response options were presented bilaterally, showed this response deficit to be defined in egocentric coordinates, with the severest response deficits for the most contralateral locations

Huntingtin subcellular localisation is regulated by Kinase signalling activity in the StHdhQ111 model of HD

Huntington’s disease is a neurodegenerative disorder characterised primarily by motor abnormalities, and is caused by an expanded polyglutamine repeat in the huntingtin protein. Huntingtin dynamically shuttles between subcellular compartments, and the mutant huntingtin protein is mislocalised to cell nuclei, where it may interfere with nuclear functions, such as transcription. However, the mechanism by which mislocalisation of mutant huntingtin occurs is currently unknown. An immortalised embryonic striatal cell model of HD (StHdhQ111) was stimulated with epidermal growth factor in order to determine whether the subcellular localisation of huntingtin is dependent on kinase signalling pathway activation. Aberrant phosphorylation of AKT and MEK signalling pathways was identified in cells carrying mutant huntingtin. Activity within these pathways was found to contribute to the regulation of huntingtin and mutant huntingtin localisation, as well as to the expression of immediate-early genes. We propose that altered kinase signalling is a phenotype of Huntington’s disease that occurs prior to cell death; specifically, that altered kinase signalling may influence huntingtin localisation, which in turn may impact upon nuclear processes such as transcriptional regulation. Aiming to restore the balance of activity between kinase signalling networks may therefore prove to be an effective approach to delaying Huntington’s disease symptom development and progression