Neuropathology in Huntington’s Disease: A Balancing Act between Neurodegeneration and Aggregates

Abstract

Neuropathology of Huntington’s disease (HD) presents with progredient neuronal cell loss mainly in the striatum, but also in multiple other brain areas suggesting HD as a multisystem neurodegenerative disorder. Mutant huntingtin aggregates are the characteristic hallmark of HD. The aggregates are misfolded proteins varying in location, form, size and structural composition indicating a complex involvement in neurotoxicity. The question if and how the aggregates and many interacting protein partners may lead to cell death is continuously a matter of debate. The role of mutant huntingtin is more than ever of paramount importance as present genetic therapeutic approaches try to target downregulation of the Huntingtin gene expression and/or lowering the corresponding protein. In this context—and these aspects are focussed—it is of crucial interest to elucidate the regional distribution as well as the cellular and subcellular localization of aggregates in established animal models of HD and in affected HD brains.

1. Introduction

The autosomal dominantly transmitted Huntington’s disease (HD) is caused by an expanded cytosine-adenine-guanine (CAG) trinucleotide repeat in exon 1 of the Huntingtin gene (HTT) resulting in an abnormally long polyglutamine tract in the protein huntingtin (Htt; [1]). Patients with 36–39 CAG repeats have an increasing risk to develop HD characteristic symptoms and repeats of 40 and more will result in onset of the disease within a normal lifespan [2]. In about 90% of adult-onset HD patients, the mean age of onset is between 35 and 50 years with marked individual variations; duration of the illness is usually 15–20 years. There is also a correlation between the CAG repeat length and the age of onset in HD [3]. Manifest patients ≤20 years were classified as juvenile-onset HD patients with an estimated prevalence of up to 15%, associated with CAG repeats >60 leading to early death [4, 5]. Core clinical symptoms are cognitive decline, progredient motor impairments and psychiatric alterations—the latter often preceding the onset of the other symptoms.

Neuropathologically, HD shows progredient neuronal cell loss most pronounced in the neostriatum, but also in many other cortical and non-cortical brain areas with considerable regional differences between the HD individuals reflecting the high variability of clinical symptoms. Currently, there is no cure for HD, and only symptoms can be treated.

HD-affected brains show misfolded proteins in form of mutant Huntingtin (mHtt) aggregates, which may be toxic or protective, and their pathomechanism is far from being understood. Aggregates are detected in the nucleus of neurons, the cytoplasm, cell processes and the neuropil. Notably, new therapies address lowering the mHTT gene production and/or mHtt protein expression to slow down or even stop disease progression [6, 7]. Therefore, localization of mHtt in HD-affected brains is of major interest in the interplay between the pathogenesis and therapeutic approaches.

In this chapter, we start with some general aspects on neurodegeneration in the human HD brain, then review the distribution and composition of mHtt aggregates and inclusions in two selected rodent models and in human HD brains and conclude with an outlook to future studies to further elucidate the controversial discussion about aggregates and their toxicity.

2. Neurodegeneration in human HD brains

Degeneration of human HD brains has been reported long before the causative gene was detected [8, 9]. The diagnosis was initially performed according to family history, characteristic choreiform movements, cognitive decline and the progredient course of the disease. First post-mortem studies focussed on bilateral striatal atrophy that has always been the most pronounced and consistent macroscopic alteration of the HD brain. The striatal atrophy, which occurs in 95% of all examined HD post-mortem brains, has led to the grading system of Vonsattel, still the most used tool when neurodegeneration of the human HD brain is classified [10, 11]. Based on a high number of post-mortem brains, Vonsattel evaluated the degeneration of striatal areas including the caudate nucleus and putamen as dorsal (motor) and the accumbens as ventral (limbic) neostriatum and the globus pallidus with its external and internal segment as paleostriatum, the latter belonging to the diencephalon. The caudate nucleus and putamen are first affected with a progredient caudo-rostral, medio-lateral and dorso-ventral shift, followed by the accumbens ventrally to the head of the caudate nucleus. Vonsattel determined the temporospatial striatal atrophy into five grades (0–4), also involving the pathohistology of the affected areas. An example of the striatal atrophy at grade 3–4 is documented in Figure 1A. At this advanced stage of degeneration, the medial striatal outline bordering the ventricle is straight, the caudate nucleus can hardly be identified and the putamen and globus pallidus are enormously shrunken. The neostriatal atrophy is due to the progredient loss of medium-sized projection neurons that are with about 90–95% the most abundant neuronal cell type in all neostriatal areas (Figure 1C). With progredient degeneration, most neurons are dysmorph or lost in the caudate nucleus and putamen, especially in the dorsal parts, accompanied by a marked increase in glial cells (Figure 1B). In HD, astrocytosis was reported to be inversely proportional to neuronal cell loss with a later beginning, whereas the density of oligodendrocytes was already increased at the early Vonsattel grades 0, 1 and 2 [12]. An increase in oligodendrocytes was also observed in the tail of caudate nucleus in HD mutation carriers already prior to the onset of symptoms [13]. A grade-dependant increase of activated microglia was also found in the dorsal neostriatum and globus pallidus [14]. In contrast, ventral striatal areas only show mild neuronal cell loss and moderate gliosis, even at advanced stage of degeneration [11]. The pronounced vulnerability of medium-sized projection neurons is still unclear [15, 16] and the fate of local aspiny striatal interneurons which make up about 5–10% of all striatal neurons is still controversial. They appear to be less and later affected in HD [17]. The striatal interneurons consist of different subpopulations all of which may undergo a different grade-dependant degeneration pattern [18]. Notably, striatal atrophy is also correlated with HD repeat size, younger age of onset and age of death [19].

Figure 1.

(A) Frontal HD brain section at advanced degeneration stage (Vonsattel grade 3–4) and a control brain at comparable striatal level with the anterior commissure (ac). Atrophy of the caudate nucleus (Cd) adjacent to the internal capsule (ic) leads to straight outline (white arrowheads) bordering the enlarged lateral ventricle (V) not observed in striatal areas of the control brain. Putamen (Pu) and globus pallidus (GP) also display severe atrophy. Note the shrinkage of white matter (wm) and the corpus callosum (cc) in the HD brain. (B) Cresyl violet-stained paraffin section (15 μm) of the HD putamen displays loss of most medium-sized striatal neurons with some neuronal cell bodies left (black arrowheads) and a pronounced gliosis as detected by abundant small cell nuclei. (C) Section of the control brain shows normal distribution of medium-sized striatal neurons (black arrowheads). Bar in A = 5 cm; bar in C for B and C = 150 μm.

The globus pallidus as the main neostriatal projection area also undergoes severe atrophy in HD (Figure 1A). Pallidal degeneration starts later at Vonsattel grade 2 up to a volume loss of around 50% at grade 4 with the external (the major output target area of the dorsal neostriatum) more affected than the medial segment [20]. Interestingly, the pallidal volume shrinkage is mainly due to loss of neuropil [11, 21], which could represent the reduction of projection axons emerging from neostriatal neurons und their synaptic terminals densely contacting the large pallidal neurons and their proximal dendritic shafts.

Beyond the striatum cortical, other diencephalic and brainstem areas are also affected though highly variable in expression. Morphometric studies of the telencephalon detected that all four lobes showed cortical atrophy more expressed in parietal and occipital than in frontal and temporal areas [22]. These observations were confirmed by MRI-based studies [23]. Imaging studies can be applied in large cohorts of patients in pre-symptomatic, early, middle and late stages and are therefore valuable tools for investigating the development of atrophy in HD brains. A recent MRI study in HD patients carried out annually over a time period of 10 years also confirmed the greatest atrophy in parietal and occipital cortical areas [24]. Remarkably, neuronal cell loss is considerably variable between HD subjects as detected in selected cortical areas [25]. Furthermore, loss of neurons in the primary motor cortex is related to motor symptoms, whereas loss of neurons in the anterior cingulate cortex is related to mood disturbances [26]. To date, many cortical areas have not yet been examined.

In addition to pallidal studies, diencephalic investigations focussed on thalamic and hypothalamic affection. A voxel-based morphometry-based study detected a co-variation between atrophy and cognitive performance suggesting impairment in executive functions [27]. Atrophy is described in the centromedian/parafascicular thalamic complex [28]. The centromedian nucleus is involved into the sensorimotor-associated basal-ganglia-thalamo-cortical feedback loop. The mediodorsal nucleus, which is involved in the corresponding limbic loop, also shows significant neuronal cell loss [29]. Thus, thalamic nuclei involved in HD-associated functionally important feedback loops appear to be severely affected in HD.

Interestingly, the hypothalamus shows a significant loss of grey matter signals already in prodromal HD individuals [30]. Some non-motor dysfunctions are discussed to be associated with changes in neuropeptidergic cell populations disturbing hypothalamic circuitry [31]. Dysfunctions include daily hormone excretion pattern and circadian rhythm disorders, which could also be a target for therapeutic treatment in the disease.

As HD patients show cognitive decline such as planning deficits and short-term memory impairments often already in prodromal phases of the disease, hippocampal involvement should also be considered. A mild but significant atrophy of the hippocampus formation was observed by Lange and Aulich [22] and later confirmed by MRI-based morphometric studies [23]. Accordingly, Vonsattel et al. [11] detected loss of neurons and gliosis in numerous HD cases. However, to really evaluate the hippocampal impact on cognitive impairments, more specific studies on the different subdivisions and cell populations in correlation with clinical symptoms are necessary.

Consistent neuronal loss was also detected in brain stem areas such as substantia nigra, superior and inferior olive, pontine and vestibular nuclei [32]. Regional brain stem affection may contribute to better understanding of vestibular and oculomotor dysfunctions in HD, the latter being one of the main clinical features of HD.

Vonsattel et al. [11] reported on the basis of more than 1000 post-mortem brains that the cerebellum is only slightly smaller in grade 3 and 4 HD brains than in controls. He also detected that the mainly segmental loss of Purkinje cells is inconsistent across the HD brains examined. In contrast, Rüb et al. [33] found Purkinje cell loss in the cerebellum and loss of neurons in the four cerebellar nuclei. In a recent study, significant Purkinje cell loss was correlated with motor impairments, whereas no loss was associated with a major mood-phenotype in HD [34]. Notably, cerebellar atrophy is particularly pronounced in juvenile-onset HD individuals accompanied with neuronal loss and gliosis [35].

White matter degeneration is most obvious in telencephalic areas including the corpus callosum and internal capsula (Figure 1A) indicating a severe affection of interhemispheric commissural connections and projection fiber tracts between cortical and noncortical brain regions. White matter alterations occur early in HD as supported by post-mortem [36] and magnetic resonance studies [37, 38]. Fiber tracts that are less in HD focus are also early affected. The fornix connecting the hippocampus with mammillary bodies displays a reduction of 34% already in prodromal cases and 41% in manifest HD [39]. This study also shows that white matter pathology is partly due to myelin breakdown and reduction of oligodendrocyte genes.

All in all, for many cortical and noncortical areas including white matter and fiber tracts, detailed information is still limited and needs more specific interdisciplinary investigations to provide a better understanding of the regional pathology and respective functional impairments. Of note, HD may also be related to other neurodegenerative diseases, for example Alzheimer (AD) and Parkinson disease that could influence regional degeneration [19]. However, the frequency of the coexistence of AD in HD is similar to AD in general population [11]. These aspects have to be considered when evaluating the neuropathology of HD, especially at older ages. Finally, the variation in neuronal degeneration among different HD patients also reflects the heterogeneity in functional impairments and pathogenesis.

3. Mutant Huntingtin aggregates

Misfolded proteins are common in many neurodegenerative diseases such as amyloid plaques or neurofibrillary tangles in Alzheimer and Lewy bodies in Parkinson disease. However, Alzheimer and Parkinson diseases comprise a group of disorders with similar symptoms, but may be caused by various reasons. HD is caused by a single gene; therefore, studies on HD-specific aggregates are particularly useful, as they are well comparable among different HD-affected brains. The CAG trinucleotide repeat in the HTT gene leads on translation to a polyglutamine stretch at the N-terminus of the protein Htt. Misfolded fragments of the protein are detected as aggregated forms differing in size, shape and composition within the cell nucleus, soma, cell processes or optionally also in the intercellular space. The pathogenesis of aggregates is still unclear and the interplay between neurodegeneration and aggregation far from being understood.

In HD brains, misfolded proteins were first described as so-called inclusion bodies by conventional electron microscopy [40]. They were detected in the nucleus of neurons as membrane-less round structures that could be distinguished due to lighter homogenous appearance from the surrounding caryoplasm. This observation must have been more of an incidental finding, as intranuclear inclusions are relatively rare in adult-onset human HD brains and extremely difficult to detect without immunohistochemical staining. Similar intranuclear structures with fibrillar and granular composition were also detected in the first transgenic HD animal model, the R6/2 mouse, as documented in Figure 2, by conventional transmission electron microscopy. R6/2 mice express exon 1 of the human HD gene with 115–150 CAG repeats, develop symptoms very early with some features reflecting juvenile-onset HD and exhibit a widespread distribution of intranuclear inclusions in all brain areas [41, 42]. Next, in human HD brains, the presence of mHtt aggregates was confirmed in nuclei and axons by light- and electronmicroscopic mHtt- and Ubiquitin-immunohistochemistry [43]. Generation of the EM48 antibody, which is specific to N-terminal fragments of mHtt [44], confirmed and extended the localization of mHtt aggregates/inclusions in neuronal cytoplasm, dendrites, axons and synapses [45]. Since then, the presence of aggregates and/or inclusions became the characteristic hallmark for histopathology in human HD brains and the increasing number of small and large animal models. From the morphological viewpoint, the terms for aggregates and inclusions are heterogenously used and therefore confusing. Common descriptions are aggregates in the neuropil and inclusions localized intranuclear. Considering the heterogenous size and form in human HD brains [43], it is difficult to distinguish one from another. In this chapter, we try to use both simultaneously, if possible.

Figure 2.

Transmission electron microscopy of a cortical pyramidal neuron in R6/2 mouse. (A) The intranuclear inclusion (INI) is a membrane-less structure clearly distinguished from the surrounding chromatin in the caryoplasm and the nucleolus (N). (B) Enlargement of the INI reveals loosely arranged fibrillar (white arrowheads) and granular structures. Postembedding Ubiquitin immunogold staining exhibits particles (15 nm) localized in the INI (black arrowheads), not in caryoplasm. Bar in A = 1 μm; bar in B = 0.5 μm.

Studying mHtt aggregates/inclusions in the broad spectrum of HD animal models, it becomes apparent that the regional, cellular and subcellular localization is as diverse as the large number of HD models themselves. The use of different antibodies makes the assessment of comparable results still more difficult, furthermore, if strong retrievals are used prior to antibody incubation.

Nevertheless, it is undisputed that animal models are valuable to elucidate crucial aspects of underlying pathomechanisms, help to understand the neurological dysfunction and psychiatric alterations and are undispensable for the development of preclinical therapeutic approaches. The choice of an HD animal model will always depend on the underlying question. However, it has to be considered carefully to which extent the respective animal model could answer the respective question in human HD. To elucidate differences and similarities of mHtt aggregates, two established rodent models were presented here in more detail.

3.1 Aggregates in R6/2 mouse

The R6/2 mouse presents with behavioural and motor dysfunctions very early and shows severe other symptoms as progredient weight loss with affection of many peripheral organs leading to early death at 12–15 weeks of age [41]. According to their rapid and reproducible phenotype, they were early transferred to commercial breeding from where they are accessible by all interested scientists. The easy availability has also contributed to the fact that the R6/2 mouse has become one of the most extensively studied HD animal model.

Neuropathologically, the R6/2 mouse displays the greatest density of aggregates/inclusions, which makes this model extremely valuable when aggregates/inclusions are used for follow-up studies and/or in vitro and in vivo investigations of the misfolded proteins themselves. Therefore, the R6/2 mouse with abundant aggregates, the early and severe symptoms and a short life span has become a standard model for testing preclinical therapeutic approaches.

When investigating by conventional electron microscopy (Figure 2), intranuclear inclusions are easily detected in adult R6/2 mice in all brain areas inspected. As in human HD brains, the membrane-less intranuclear inclusion is clearly distinguished from the surrounding caryoplasm and comprises homogenously distributed fibrillar and granular structures (Figure 2A and B). Postembedding immunogold staining with Ubiquitin confirms intranuclear inclusion.

Immunostaining with EM48 antibody reveals an overall distribution of mHtt aggregates in all R6/2 brain areas. At cellular level, many neurons exhibit reactivity throughout the caryoplasm and a dense inclusion (Figure 3A). Immunoelectron microscopy confirmed nuclear distribution of EM48 reactivity loosely distributed in the caryoplasm and the dense intranuclear inclusion (Figure 3B). This observation extends the general assumption, that intranuclear mHtt is mainly localized as inclusion body. In R6/2 mice, the whole nucleus may harbour aggregates with varying expression. Nucleoli are always spared (Figure 3B). Of note, neurons with immunopositive caryoplasm show signs of degeneration as the irregular invaginated nuclear envelope starts to collapse indicating that mHtt may cause the cellular dysfunction finally leading to cell death (Figure 3B). The cytoplasm lacks mHtt reactivity. Single immunopositive spots were also detected in the surrounding neuropil. Taken together, the aggregates/inclusions in R6/2 mice are distributed across all brain areas, focussed on the caryoplasm and sparsely localized in the neuropil.

Figure 3.

EM48-immunohistochemistry in the striatum of R6/2 mouse. (A) Vibratome section (50 μm) displays brown-stained nuclei (black arrowheads) many of which with a black inclusion body. Single positive spots are distributed in the neuropil (white arrowheads). (B) Transmission immunoelectron microscopy confirms EM48 reactivity in the caryoplasm (C) densely arranged in the intranuclear inclusion (INI). Nucleoli (N) are spared. Immunopositive nuclei show irregular nuclear envelope. Adjacent glial cell nucleus (G) lacks EM48 reactivity. Bar in A = 100 μm; bar in B = 1 μm.

3.2 Aggregates in the tgHD rat

The transgenic rat model of HD (tgHD rat) carries a truncated htt cDNA fragment with 51 CAG repeats under control of the native rat promotor [46]. In contrast to the R6/2 mouse, the tgHD rat presents with slowly progressive motor and behavioural impairments reflecting the adult-onset phenotype of human HD individuals. Interestingly, the tgHD rat shares neuropathological similarities in regional distribution and subcellular composition of aggregates with human HD brains. In the tgHD rat und in human HD brain, aggregates are focussed on the ventral striatum and the extended amygdala [47, 48, 49] areas that are crucial for elucidating psychiatric aspects of the disease. In the tgHD rat, detailed transmission immunoelectron microscopy detected that aggregates are localized in medium-sized striatal neurons as small patches in neuronal cytoplasm, mitochondria, myelinated and unmylinated axons, synaptic terminals and, most frequently, loosely distributed or as large compact inclusions in dendrites and dendritic spines [48].

Aggregates are also localized in the nucleus (Figure 4). In contrast to the R6/2 mouse, the tgHD rat caryoplasm only exhibits very few small EM48-positive spots and occasionally a single inclusion (Figure 4A and B). Signs of degeneration are rarely observed in the striatal neurons. In sum, the tgHD rat shows a more regional mHtt distribution focussed on basal forebrain systems. On subcellular level, aggregates/inclusions may be detected in many parts of medium-sized striatal neurons.

Figure 4.

EM48 immunohistochemistry in the neostriatum of tgHD rat. (A) Transmission electron microscopy shows an intranuclear inclusion (INI) in a normal appearing medium-sized neuron of a 23 months old tgHD rat. Some positive spots (black arrowheads) are also detected in the neuronal caryoplasm, cytoplasm and dendrites (D). (B) At higher enlargement the INI exhibits fibrillar (white arrowheads) and granular structures. Ly, lysosomes; bar in A = 5 μm; bar in B = 1 μm.

3.3 Aggregates in human HD brains

There are multiple studies on aggregates in HD animal models, but the localization of mHtt in human HD brains is less extensively investigated. In the studies of DiFiglia et al. and Gutekunst et al. [43, 45], important aspects of regional, cellular and subcellular aggregate localization were worked out in detail. Brains investigated included various Vonsattel grades as well as juvenile- and adult-onset HD brains. MHtt immunopositive intranuclear aggregates consistently called inclusion bodies by DiFiglia et al. [43] were more frequently detected in cortical layers of juvenile- than in adult-onset HD brains, in which they were predominantly detected in the neuropil in neuronal cell processes (called dystrophic neurites). The pronounced cortical localization in layers V and VI—especially in adult-onset HD individuals—was confirmed by Gutekunst et al. [45] for various cortical areas. Our investigations of selected frontal, parietal, temporal and occipital cortical areas by peroxidase EM48-immunohistochemistry also show that layers V and VI display the highest amount of aggregates/inclusions in varying degrees (unpublished results). The striatum, which is the first focus of studies in HD animal models, only displays a limited amount of aggregates in human HD brains, more expressed in the ventral than in the dorsal neostriatum [43, 45]. This observation was extended by our investigation, as we found aggregates/inclusions focussed to the accumbens and the extended amygdala [49], both functional-anatomical entities acting as interface between motor, limbic and olfactory-associated basal forebrain areas.

All human brains investigated in our cohort showed a heterogenous spectrum of aggregates differing in size, form and composition (Figure 5A). Confocal EM48 immunofluorescence counterstained with DAPI detected that most aggregates are localized in the neuropil, and only a few nuclei are associated with small positive spots (Figure 5B). Of note, it is relatively easy to localize aggregates in the nucleus and cytoplasm; however, the exact assignment to neuronal cell processes in the neuropil is difficult and awaits further detailed investigations using various techniques. One of the techniques to elucidate the fine structural composition of aggregates is transmission electron microscopy. Large inclusions often display an immunopositive rim with granular and vesicular structures and a mainly immunonegative core with densely arranged fibrillar structures (Figure 5C). In sum, in human HD brains aggregates/inclusions are predominantly localized in cortical areas, and—less expressed—in selected basal limbic-associated forebrain systems. Localization of aggregates/inclusions in many subcortical areas is less investigated and awaits further and more detailed investigation. Particularly, correlation studies between aggregate distribution and neurological dysfunctions are almost completely lacking in human HD.

Figure 5.

EM48-immunohistochemistry in human HD brain (CAG 54/20). (A) Vibratome section (80 μm) displays distribution of mHtt aggregates varying in form and size localized in layer V and VI of the anterior cingulate cortex. (B) Confocal immunofluorescence of a cryosection (15 μm) with EM48 (green) counterstained with DAPI (blue) shows mainly neuropil aggregates. Some nuclei are associated with tiny positive dots (white arrowheads). (C) Transmission electron microscopy exhibits a large aggregate with an immunopositive rim (Ri) and a mainly immunonegative core (Co). Arrowheads mark the border of the neuropil structure harbouring the aggregate. M, mitochondrium; T, synaptic terminal; bar in B for A and B = 50 μm; bar in C = 1 μm.

A breakthrough to understand the mHtt structure at close to native cellular level was performed by the recently developed high-resolution cryo-electron tomography [50, 51]. This methods allows a three-dimensional imaging of cytosolic inclusions and aggregates [51]. Hela cells and mouse neurons transfected with GFP-tagged Htt exon 1 comprising 97 Q displayed inclusions which were identified by live cell imaging and further treated for cryo-electron tomography. Large mHtt inclusions are composed of organized centrally located fibrils, which interact with the membranes of the endoplasmic reticulum and deform their normal organization. This observation elucidates the subcellular machinery of mHtt aggregates and suggests a destructive effect of the inclusions. Comparing Bäuerlein’s results with the large inclusions detected in the human HD brains investigated here by transmission electron microscopy (Figure 5C), it may look similar with the granular and vesicular structures in a more loosely arranged rim area and tightly packed fibrillar structures within the core. The question remains how far Bäuerlein’s results reflect the broad spectrum of mHtt inclusions/aggregates in the human HD brain. Nevertheless provide these results important insights into cellular impairments by mHtt inclusions/aggregates and are encouraging findings, which show that the interdisciplinary research on subcellular level is currently on the way to complement one another.

For therapeutic approaches that target lowering of mHtt levels in the brain, it is of major interest to develop non-invasive tools as biomarkers to visualize mHtt during disease progression. Imaging agents visible by positron emission tomography would be extremely helpful to identify and track mHtt distribution prior and during the therapy. Recently, a high-affinity Fluorine-18 radioligand was developed for imaging mHtt aggregates in HD animal models and also human post-mortem HD brain tissue [52]. This PET-imaging agent showed sufficient brain uptake in rodents and non-human primates to be monitored in vivo. Furthermore, autogradiography with the ligand displayed specific binding on human post-mortem HD brain sections, which may correspond to aggregate accumulation as further indicated by mHtt-immunohistochemistry on adjacent sections. Even if such non-invasive studies are still at the very beginning, the application of radioligands as PET-imaging tracer in human HD brains to monitor alterations of mHtt localization would be of enormous benefit for controlling the course of therapy. It is also of crucial importance to carefully evaluate the validity of ligands as imaging biomarkers especially in mHtt-rich human brain areas, as localization of mHtt may be completely different from the before investigated animal model.

4. Conclusions

In HD, neurodegeneration is most expressed in striatal areas, but cortical and other noncortical areas are also severely affected including the grey and—especially in early HD—also the white matter. Notably, neurodegeneration is varying across the different HD brains, which may reflect the diversity in functional impairments of HD patients. For many brain areas, detailed information about macroscopic and microscopic affection is still limited and needs more specific interdisciplinary investigations to provide a better understanding of the regional neuropathology and related dysfunctions.

Aggregates/inclusions are the characteristic histopathological hallmark of HD. In HD animal models, the regional, cellular and subcellular localization is as diverse as the large number of models themselves with differently pronounced similarities to the human HD aggregation pattern. To date, the exact role of the Htt protein has not yet been clarified. The mechanism of formation and maturation of aggregates is currently intensively studied in living cell cultures providing first insights into the dynamic of mHtt and the toxic influence of aggregates and inclusion bodies. Furthermore, high-resolution techniques and improved tissue preservation are necessary to transfer the results on living cells to the aggregation process in human HD brains. So far, the controversial discussion about gain of function and toxicity in the interplay between aggregates and neurodegeneration is going on.

Acknowledgments

We are grateful to Anne Schlichting, Luzie Augostinowski, Katja Rumpf, Sabine Peuckert, Robert Nadgrabski, Claudia Schneider (Institute of Anatomy, Medical Faculty, Ruhr-University Bochum, Germany) for their excellent technical assistance.

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Conflict of interest

There are no conflicts of interest.