domain: sdv scco

text:
Robert Lalonde
email: robert.lalonde@univ-rouen.fr

Catherine Strazielle

Motor performances of spontaneous and genetically modified mutants with cerebellar atrophy

Keywords: Cerebellum, Purkinje cells, Rotorod, Stationary Beam, Wire Suspension, Spinocerebellar ataxia

Chance discovery of spontaneous mutants with atrophy of the cerebellar cortex has unearthed genes involved in optimizing motor coordination. Rotorod, stationary beam, and suspended wire tests are useful in delineating behavioral phenotypes of spontaneous mutants with cerebellar atrophy such as Grid2 Lc , Grid2 ho , Rora sg , Agtpbp1 pcd , Reln rl , and Dab1 scm . Likewise, transgenic or null mutants serving as experimental models of spinocerebellar ataxia (SCA) are phenotyped with the same tests. Among experimental models of autosomal dominant SCA, rotorod deficits were reported in SCA1 to 3, SCA5 to 8, SCA14, SCA17, and SCA27 and stationary beam deficits in SCA1 to 3, SCA5, SCA6, SCA13, SCA17, and SCA27. Beam tests are sensitive to experimental therapies of various kinds including molecules affecting glutamate signaling, mesenchymal stem cells, anti-oligomer antibodies, lentiviral vectors carrying genes, interfering RNAs, or neurotrophic factors, and interbreeding with other mutants.

Introduction

Naturally mutated mice with cerebellar atrophy have been known for over half a century. [START_REF] Cendelin | From mice to men: lessons from mutant ataxic mice[END_REF][START_REF] Lalonde | Motor performance of spontaneous murine mutations with cerebellar atrophy[END_REF] Over the course of the last quarter century, technological advances in transgenic and targeted null mutations have yielded genetically modified mutants with cerebellar atrophy, notably to model autosomal dominant spinocerebellar ataxias (SCAs) numbered at the present time from SCA1 to SCA37 according to individually discovered genes. We review naturally mutated mice with cerebellar atrophy together with induced SCA mutations. Known for a longer time, the spontaneous mutants have been more thoroughly evaluated than the induced mutants and therefore serve as contrast with what remains to be determined in the newer models. Among the older models, we selected the most behaviorally characterized ones and those with more selective damage to the cerebellar cortex.

The main tests of motor coordination used to test natural mutations are stationary beam, suspended wire, and rotorod tests, all sensitive to cerebellar lesions. [START_REF] Lalonde | Motor performance of spontaneous murine mutations with cerebellar atrophy[END_REF][START_REF] Brooks | Tests to assess motor phenotype in mice: a user's guide[END_REF] Mice are placed on a stationary beam (Fig 1A) and either walk along it or remain still. In the suspended wire test, mice are placed upside-down on a horizontal wire and stay upside-down with two paws or four. If able to maintain their position with four paws, they can lift themselves upright on a sidebar of a coathanger (Fig 1B). On the rotorod (Fig 1C), mice are placed on a beam that rotates and must move in synchrony with it. In all three tasks, latencies before falling and number of falls are measured.

In stationary beam and coat-hanger tasks, movement time (MT) can be estimated as defined by the time taken to walk or slide across the beam or bar. Different types of MT have been used in the coat-hanger version to delineate more precisely difficulties encountered while moving along the horizontal bar and climbing atop the diagonal bar. Since animals support their body weight in an upside-down position, the suspended wire test requires muscle strength all the more, relevant in this context because one of the main features of cerebellar symptoms is hypotonia. In some lesioned animals, the loss in balance on the stationary beam can be compensated by remaining still, so that an additional measure can be used to disclose the deficit, namely the distance travelled along the beam or the time taken to reach the end of the beam. In contrast, the rotorod cannot be compensated by immobility unless the animal wraps its legs around the beam. After two successive rotations of this passive type, for example, the animal should be removed and the trial ended as if it fell.

Natural mouse mutations with cerebellar atrophy

Table 1 illustrates neurobehavioral characteristics of mice with natural mutations causing relatively selective degeneration of the cerebellar cortex and deficits in motor coordination tests. [START_REF] Lalonde | Motor performance of spontaneous murine mutations with cerebellar atrophy[END_REF][START_REF] Brooks | Tests to assess motor phenotype in mice: a user's guide[END_REF]

Grid2

The semi-dominant Lurcher mutation causes a gain-in-malfunction of Grid2 located on chromosome 6 and encoding the GluRδ2 ionotropic glutamate receptor associated with the amino-methyl-isoxazoleproprionate (AMPA) receptor. [START_REF] Zuo | Neurodegeneration in Lurcher mutant mice caused by mutation in the delta2 glutamate receptor gene[END_REF] Grid2 mRNA is predominantly expressed in cerebellar Purkinje cells. [START_REF] Takayama | Developmental changes in expression and distribution of the glutamate receptor channel delta2 subunit according to the Purkinje cell maturation[END_REF] The depolarized membrane potential of the Grid2 Lc encoded receptor is probably responsible for the nearly total loss of Purkinje cells from postnatal week 2 to 4. [START_REF] Caddy | Structural and quantitative studies on the normal C3H and Lurcher mutant mouse[END_REF] Two autosomal recessive hot-foot alleles (4J and nancy) cause different deletions in the coding sequences of Grid2. [START_REF] Lalouette | Hot-foot mutations affect the δ2 glutamate receptor gene and are allelic to Lurcher[END_REF][START_REF] Lalouette | Neurobiological effects of a null mutation depend on genetic context: comparison between two hotfoot alleles of the delta-2 ionotropic glutamate receptor[END_REF] For the ho-4J allele, the truncated GluRδ2 protein is expressed in Purkinje cell soma without being transported to the cell surface. [START_REF] Matsuda | Mutation in hotfoot-4J mice results in retention of δ2 glutamate receptors in ER[END_REF] The main neuropathological marker in Grid2 ho-nancy mutants is defective parallel fiber-Purkinje cell innervation and mild depletion of cerebellar granule cells, resulting in cerebellar ataxia and a hopping gait reminiscent of mice walking on a hot plate. [START_REF] Guastavino | Hot-foot murine mutation: behavioral effects and neuroanatomical alterations[END_REF] In contrast to the gain-in-malfunction subtending the Lc allele, ho alleles are non-functional, as indicated by the similar phenotype seen in the Grid2 targeted null mutation marked by reduced synaptic contacts on Purkinje cells, ataxia, and motor coordination deficits. [START_REF] Kashiwabuchi | Impairment of motor coordination, Purkinje cell synapse formation, and cerebellar long-term depression in GluR delta 2 mutant mice[END_REF] Thus, in Grid2 Lc mutants [START_REF] Lalonde | Rotorod sensorimotor learning in cerebellar mutant mice[END_REF][START_REF] Lalonde | Sensorimotor learning in three cerebellar mutant mice[END_REF][START_REF] Lalonde | Motor deficits in Lurcher mutant mice[END_REF][START_REF] Strazielle | Regional brain variations of cytochrome oxidase activity and motor coordination in Lurcher mutant mice[END_REF][START_REF] Hilber | Motor skills and motor learning in Lurcher mutant mice during aging[END_REF] and Grid2 ho-nancy mutants, [START_REF] Guastavino | Hot-foot murine mutation: behavioral effects and neuroanatomical alterations[END_REF][START_REF] Kashiwabuchi | Impairment of motor coordination, Purkinje cell synapse formation, and cerebellar long-term depression in GluR delta 2 mutant mice[END_REF][START_REF] Lalonde | Rotorod sensorimotor learning in cerebellar mutant mice[END_REF][START_REF] Krémarik | Regional brain variations of cytochrome oxidase activity and motor coordination in hot-foot mutant mice[END_REF] there is cerebellar ataxia as defined as a wide-spread gait and defective performances on stationary beam, coat-hanger, and rotorod tests relative to non-ataxic controls.

Rora

The autosomal recessive staggerer mutation deletes Rora located on chromosome 9, which encodes a retinoid-like nuclear receptor involved in neuronal differentiation and maturation highly expressed in Purkinje cells. [START_REF] Hamilton | Disruption of the nuclear hormone receptor ROR in staggerer mice[END_REF][START_REF] Nakagawa | Prominent expression of nuclear hormone receptor RORα in Purkinje cells from early development[END_REF] As in hot-foot, the lost function of the Rora sg mutation was confirmed by observing nearly identical neuropathological and behavioral phenotypes in targeted Rora null mutants. [START_REF] Doulazmi | A comparative study of Purkinje cells in two RORα gene mutant mice: staggerer and RORα[END_REF][START_REF] Steinmayr | Staggerer phenotype in retinoid-related orphan receptor α-deficient mice[END_REF] In Rora sg homozygotes, Purkinje cells decline in number before postnatal day 5, and, at the end of the first postnatal month, 75% of them disappear. [START_REF] Herrup | Regional variation and absence of large neurons in the cerebellum of the staggerer mouse[END_REF] Thus, the Purkinje cell loss begins at an earlier stage than Grid2 Lc (postnatal day 8) but is less complete. [START_REF] Caddy | Structural and quantitative studies on the normal C3H and Lurcher mutant mouse[END_REF] Even when present, metabotropic glutamate receptor type 1 (mGluR1)mediated signaling at cerebellar parallel fiber-Purkinje cell synapses was disrupted in Rora sg mutants. [START_REF] Mitsumura | Disruption of metabotropic glutamate receptor signalling is a major defect at cerebellar parallel fibre-Purkinje cell synapses in staggerer mutant mice[END_REF] As with Grid2 mutants, performances on stationary beam, coat-hanger, and rotorod tests were deficient in Rora sg mutants relative to non-ataxic controls. [START_REF] Kashiwabuchi | Impairment of motor coordination, Purkinje cell synapse formation, and cerebellar long-term depression in GluR delta 2 mutant mice[END_REF][START_REF] Lalonde | Rotorod sensorimotor learning in cerebellar mutant mice[END_REF][START_REF] Lalonde | Motor abnormalities in staggerer mutant mice[END_REF][START_REF] Deiss | Regional brain variations of cytochrome oxidase activity and motor co-ordination in staggerer mutant mice[END_REF]

Agtpbp

The autosomal recessive Purkinje cell degeneration (pcd) mutation affects Agtpbp1 located on chromosome 13 and encoding ATP/GTP binding protein 1. [START_REF] Fernandez-Gonzalez | Purkinje cell degeneration (pcd) phenotypes caused by mutations in the axotomy-induced gene, Nna1[END_REF] In normal mouse brain, Agtpbp1 mRNA is prominent in Purkinje cells, mitral cells of olfactory bulb, thalamic neurons, and retinal photoreceptors, all of them prone to degeneration in the mutant. Several alleles of the affected gene have been discovered, including 1J on C57BR/cdJ, 2J on SM/J, and 3J on BALB/cByJ backgrounds, respectively. The 1J allele was subsequently transferred to B6C3-a/a and C57BL/6J backgrounds and the 2J allele to C57BL/6J. The predominant pathology in Agtpbp1 pcd-1J mutants concerns almost total Purkinje cell loss from the third to the fourth postnatal week. [START_REF] Landis | The development and degeneration of Purkinje cells in pcd mutant mice[END_REF][START_REF] Mullen | Purkinje cell degeneration: a new neurological mutation in the mouse[END_REF] Agtpbp1 pcd/1J mutant mice were deficient relative to non-ataxic controls on coat-hanger and rotorod tests. [START_REF] Marec | Sensorimotor learning and retention during equilibrium tests in Purkinje cell degeneration mutant mice[END_REF] When three adult mutant models were placed on the same rectangular stationary beam, only Rora sg mutants 24 fell sooner than their non-ataxic controls, not Grid2 Lc mutants [START_REF] Lalonde | Sensorimotor learning in three cerebellar mutant mice[END_REF] or Agtpbp1 pcd/1J mutants. [START_REF] Marec | Sensorimotor learning and retention during equilibrium tests in Purkinje cell degeneration mutant mice[END_REF] This result may be explained by the almost total loss of Purkinje cells in Grid2 Lc and Agtpbp1 pcd/1J mutants but a 75% loss in Rora sg , so that the existence of dysfunctional Purkinje cells and their abnormal dendritic arborization appears to cause a worse phenotype than their absence. A second possibility is the earlier onset of Purkinje cell degeneration in Rora sg relative to the other two worsens the behavioral phenotype in the adult.

Reln and Dab1

The autosomal recessive reeler mutation disrupts Reln located on chromosome 5 and encoding reelin. [START_REF] D'arcangelo | A protein related to extracellular matrix proteins deleted in the mouse mutant reeler[END_REF][START_REF] D'arcangelo | Reelin is a ligand for lipoprotein receptors[END_REF] Two Reln rl alleles have been discovered: Jackson (J) and Orleans (Orl). The Jackson allele on the C57BL/6 or the hybrid B6C3 background has a deletion of the entire gene and Orleans on the Balb/c background a 220 bp deletion in the open reading frame, causing a frame shift. [START_REF] Hirotsune | The reeler gene encodes a protein with an EGF-like motif expressed by pioneer neurons[END_REF] Reelin is an extracellular matrix protein involved in neural adhesion and migration at critical stages of development. [START_REF] D'arcangelo | A protein related to extracellular matrix proteins deleted in the mouse mutant reeler[END_REF][START_REF] D'arcangelo | Reelin is a ligand for lipoprotein receptors[END_REF][START_REF] Hirotsune | The reeler gene encodes a protein with an EGF-like motif expressed by pioneer neurons[END_REF][START_REF] Hack | Reelin is a detachment signal in tangential chain-migration during postnatal neurogenesis[END_REF] Reln rl-J and Rel orl mutants display cell ectopias in cerebellum, hippocampus, and neocortex. [START_REF] Mariani | Anatomical, physiological and biochemical studies of the cerebellum from Reeler mutant mouse[END_REF][START_REF] Stanfield | The morphology of the hippocampus and dentate gyrus in normal and reeler mutant mice[END_REF][START_REF] Dräger | Observations on the organization of the visual cortex in the reeler mouse[END_REF] In line with elevated expression of Reln mRNA in granule but not Purkinje cells, [START_REF] Schiffmann | Reelin mRNA expression during mouse brain development[END_REF] the former constitutes the main depleted cell type in the cerebellum of the mutant. [START_REF] Caviness | Mechanisms of cortical development: a view from mutations in mice[END_REF] Among mutations upstream of the reelin signaling pathway, disruption of Dab1 located on chromosome 4 and encoding disabled homolog 1 characterizes the autosomal recessive Dab1 scm (scrambler) mutation with a similar phenotype to Reln rl ones. [START_REF] Sweet | Scrambler, a new neurological mutation of the mouse with abnormalities of neuronal migration[END_REF] Relative to non-ataxic controls, the performances on stationary beam, coat-hanger, and rotorod tests were deficient in Relrl orl mutants [START_REF] Lalonde | Neurobehavioral evaluation of Rel rl-Orl mutant mice and correlations with cytochrome oxidase activity[END_REF] and Dab1 scm mutants [START_REF] Jacquelin | Neurobehavioral performances and brain regional metabolism in Dab1 scm (scrambler) mutant mice[END_REF] , the latter being also deficient in climbing a vertical grid, a test better adapted to their motor capacity. [START_REF] Lalonde | Sensorimotor learning in Dab1 scm (scrambler) mutant mice[END_REF]

Genetically modified mouse mutations with cerebellar atrophy

To provide features of SCA subtypes (Tables 2 and3), genetically modified mice have been generated with human or animal versions of the mutated genes (transgenic) or with targeted deletion of the affected genes (null mutants).

SCA1

SCA1 is an autosomal dominant disorder caused by over 39 cytosine-adenine-guanine (CAG) trinucleotide repeats of the ATXN1 gene located on chromosome 6 and encoding ataxin-1, [START_REF] Clark | Spinocerebellar ataxia type 1-modeling the pathogenesis of a polyglutamine neurodegenerative disorder in transgenic mice[END_REF] found in Purkinje and other cells. [START_REF] Skinner | Ataxin-1 with an expanded glutamine tract alters nuclear matrix-associated structures[END_REF] The CAG or CAA triplet is specific for the amino acid glutamine, excessively repeated in several diseases designated as polyglutamine and including cerebellar or basal ganglia anomalies such as Huntington's disease. SCA1 causes adult-onset degeneration of cerebellar cortex and brainstem, leading to ataxia, dysarthria, and bulbar symptoms. Other SCA genes have been numbered and designated as "ataxins" (e.g. ataxin-2, ataxin-3, etc) when no previous function had yet been discovered.

To mimic human SCA1 disease, transgenic mice containing 82 CAG repeats in ATXN1 driven by the Purkinje-cell specific Pcp2 promoter were generated. [START_REF] Burright | SCA1 transgenic mice: a model for neurodegeneration caused by an expanded CAG trinucleotide repeat[END_REF] The cerebellar molecular layer of conditional ATXN1/Q82 transgenic mice is thinner and Purkinje cells are ectopic and shrunken, [START_REF] Clark | Purkinje cell expression of a mutant allele of SCA1 in transgenic mice leads to disparate effects on motor behaviors, followed by a progressive cerebellar dysfunction and histological alterations[END_REF] probably due to a gain-in-malfunction mutation, because Atxn1 null mutants display neither Purkinje cell pathology nor ataxia. [START_REF] Matilla | Mice lacking ataxin-1 display learning deficits and decreased hippocampal paired-pulse facilitation[END_REF] Unlike natural mutations described above whose ataxia is noticeable during the first three weeks of life, the ataxia seen in ATXN1/Q82 mice is only evident at 3 months. [START_REF] Clark | Purkinje cell expression of a mutant allele of SCA1 in transgenic mice leads to disparate effects on motor behaviors, followed by a progressive cerebellar dysfunction and histological alterations[END_REF] Nevertheless, the mutants were already impaired on rotorod and stationary beam tests as early as 5 weeks of age, when cytoplasmic vacuoles in Purkinje cells are their main histopathologic feature. At one year of age, ATXN1/Q82 transgenic mice became impaired in foot-print analyses. Compared to mutants with 82 CAG repeats, ATXN1/Q30 transgenic mice with 30 CAG repeats developed overt ataxia at a later onset and a milder form of Purkinje cell degeneration. [START_REF] Fernandez-Funez | Identification of genes that modify ataxin-1-induced neurodegeneration[END_REF] As seen in Rora sg mutants, 23 mGluR1-mediated signaling at cerebellar parallel fiber-Purkinje cell synapses was disrupted in ATXN1/Q82 transgenic mice. [START_REF] Shuvaev | Progressive impairment of cerebellar mGluR signalling and its therapeutic potential for cerebellar ataxia in spinocerebellar ataxia type 1 model mice[END_REF] Facilitated MGluR1 signaling after intracerebellar injection of the GABA B receptor agonist, baclofen, improved their rotorod performance. Contrariwise, intravermal injection of lentiviral vectors containing ATXN1/Q76 or ATXN1/Q30 with green fluorescent protein impaired rotorod performance in wild-type mice. The efficacy of intrathecally injected mesenchymal stem cells was also tested in ATXN1/Q82 mice. [START_REF] Matsuura | Mesenchymal stem cells ameliorate cerebellar pathology in a mouse model of spinocerebellar ataxia type 1[END_REF] The cells mitigated Purkinje cell ectopias, dendritic thinning, and rotorod deficits, prompting the possible use of this technique in other neurodegenerative conditions. [START_REF] Nakamura | Mesenchymal stem cells as a potential therapeutic tool for spinocerebellar ataxia[END_REF] Likewise, the injection of neural precursor cells derived from the subventricular zone into the cerebellar white matter of ATXN1/Q82 mice recovered Purkinje cell loss and ectopias as well as increasing grip strength and rotorod performance. [START_REF] Chintawar | Grafting neural precursor cells promotes functional recovery in an SCA1 mouse model[END_REF] Another strategy used to modify the ATXN1/Q82 phenotype comprises interbreeding with other mutants. Relative to the single transgenic, ATXN1/Q82 mice crossbred with Trp53 null mutant mice lacking the p53 protein involved in cell death counteracted Purkinje cell heterotopias and dendritic thinning as well as molecular layer shrinkage, though without affecting rotorod performance or formation of ataxin-1 nuclear inclusions. [START_REF] Shahbazian | Reduction of Purkinje cell pathology in SCA1 transgenic mice by p53 deletion[END_REF] When ATXN1/Q82 transgenic mice were crossbred with mice overexpressing the inducible form of rat Hsp70 encoding a molecular chaperone involved in protein folding, Purkinje cells had thicker and more arborized dendritic branches than the single transgenic but this time rotorod performance improved despite the absence of any change on ataxin-1 aggregates. [START_REF] Cummings | Over-expression of inducible HSP70 chaperone suppresses neuropathology and improves motor function in SCA1 mice[END_REF] In contrast, ATXN1/Q82 transgenic mice bred with Ube3a null mutants lacking E6-AP ubiquitin ligase had fewer ataxin-1 aggregates, but the Purkinje cell pathology was worse than the single transgenic, so that aggregates seem to have a protective role in this case. [START_REF] Cummings | Mutation of the E6-AP ubiquitin ligase reduces nuclear inclusion frequency while accelerating polyglutamine-induced pathology in SCA1 mice[END_REF] ATXN1/Q82 mice were also evaluated on whether RNA interference inhibits polyglutamine-induced neurodegeneration. [START_REF] Xia | RNAi suppresses polyglutamine-induced neurodegeneration in a model of spinocerebellar ataxia[END_REF] This time, midline cerebellar injection of recombinant adeno-associated virus vectors expressing short hairpin RNAs resolved ataxin-1 inclusions in Purkinje cells, restored cerebellar molecular thickness, and improved rotorod performance. From two of the models described above, [START_REF] Cummings | Over-expression of inducible HSP70 chaperone suppresses neuropathology and improves motor function in SCA1 mice[END_REF][START_REF] Xia | RNAi suppresses polyglutamine-induced neurodegeneration in a model of spinocerebellar ataxia[END_REF] different conclusions may be reached in regard to the role of ataxin-1 aggregates on sensorimotor coordination. In one model, [START_REF] Xia | RNAi suppresses polyglutamine-induced neurodegeneration in a model of spinocerebellar ataxia[END_REF] resolved ataxin-1 inclusions improved rotorod performance, leading to the conclusion that experimental therapies should aim at reducing the aggregates. But in another model, [START_REF] Cummings | Over-expression of inducible HSP70 chaperone suppresses neuropathology and improves motor function in SCA1 mice[END_REF] rotorod performance improved despite the absence of any change in ataxin-1 aggregates, leading to the conclusion that the aggregates are irrelevant in regard to function. These results underline the sensitivity of the rotorod test over a neuropathological marker as a target in evaluating experimental therapies.

An inducible tetracycline-regulated ATXN1/Q82 mouse model has also generated with a transgene controlled by the Pcp2 promoter with similar characteristics, including heterotopic Purkinje cells with cytoplasmic vacuoles and nuclear inclusions as well as reduced Purkinje cell dendritic arborization and lower cerebellar mGluR1 levels combined with rotorod deficits. [START_REF] Zu | Recovery from polyglutamine-induced neurodegeneration in conditional SCA1 transgenic mice[END_REF] Halting inducible ATXN1/Q82 expression with doxycycline in the drinking water reduced cytoplasmic vacuoles and nuclear inclusions and restored Purkinje cell dendritic arborization, cerebellar mGluR1 levels, and rotorod deficits, implicating the Purkinje cell-parallel fiber synapse on motor performance.

Other SCA1 models overexpress mouse not human gene mutations. In particular, Atxn1/Q154 knockin mice bear 154 CAG repeats of the mouse Atxn1 locus and express fulllength mutant ataxin-1 in its endogenous expression pattern and context, causing a loss in Purkinje cell number and dendritic branching, ataxia, paw-clasping, and rotorod deficits before the onset of ataxia. 58 In contrast, Atxn1/Q78 knockins had no obvious sign of cerebellar atrophy or ataxia but were still deficient on the rotorod. [START_REF] Lorenzetti | Repeat instability and motor incoordination in mice with a targeted expanded CAG repeat in the Sca1 locus[END_REF] To establish the role of the mutation, a cerebellar extract was injected from Atxn1/Q154 knockin or wild-type mice into deep cerebellar nuclei of Atxn1/Q78-Q2 knockins. [START_REF] Lasagna-Reeves | Ataxin-1 oligomers induce local spread of pathology and decreasing them by passive immunization slows spinocerebellar ataxia type 1 phenotypes[END_REF] The cerebellar extract from Atxn1/Q154 knockins increased oligomer formation in cerebellum and brainstem of Atxn1/Q78 knockins. As a form of experimental therapy, intraperitoneal injection of Atxn1/Q154 knockin mice with the antioligomer antibody, F11G3, reduced oligomers in Purkinje cells and improved rotorod acquisition.

In addition, Atxn1/Q154 crossed with a VEGFA transgenic line expressing human vascular endothelial growth factor-A increased calbindin staining intensity of Purkinje cells and molecular layer thickness as well as speeding up rotorod acquisition. [START_REF] Cvetanovic | Vascular endothelial growth factor ameliorates the ataxic phenotype in a mouse model of spinocerebellar ataxia type 1[END_REF] Moreover, Atxn1/Q154 knockins acquired the rotorod task more quickly after a lithium diet, 62 a manipulation affecting the neurochemistry of the cerebellum at the level of energy metabolism, purines, and unsaturated free fatty acids as well as aromatic-and sulphur-containing amino acids. [START_REF] Perroud | Pharmacometabolomic signature of ataxia SCA1 mouse model and lithium effects[END_REF] Mitochondria-targeted antioxidant MitoQ also ameliorated motor coordination in Atxn1/Q154 knockins. [START_REF] Stucki | Mitochondrial impairments contribute to spinocerebellar ataxia type 1 progression and can be ameliorated by the mitochondria-targeted antioxidant MitoQ[END_REF] Proteins interacting with ataxin-1 can promote SCA1-related neurodegeneration. In particular, pumilio1 regulates ataxin-1 levels in cells. [START_REF] Gennarino | Pumilio1 haploinsufficiency leads to SCA1-like neurodegeneration by increasing wild-type ataxin1 levels[END_REF] Pum1 null mutant mice lacking pumilio1 had higher ataxin-1 levels in cerebellum and cerebrum than wild-type, leading to loss of Purkinje cell number and dendritic arborization. This form of neuropathology likely contributes to delayed acquisition of the rotorod task, the presence of hind-paw clasping, more horizontal but less vertical activity in the open-field, and foot-print anomalies such as a wider stance, shorter stride length, and greater stride frequencies. Pum1 null mutants crossed with Atxn1/Q154 knockins increased the severity of the loss in Purkinje cell number and dendritic arborization and caused earlier onset of hind-paw clasping and mortality than the single transgenic. Contrariwise, Purkinje cell loss and dendritic arborization as well as rotorod deficits and paw-clasping in Pum1 null mutants were mitigated when intercrossed with ATXN1 null mutants.

We notice from these series of experiments that the rotorod test is the standard most often used to phenotype SCA1 models and assess the possible improvement in motor functions following various forms of experimental therapy, the stationary beam test being used only once. [START_REF] Clark | Purkinje cell expression of a mutant allele of SCA1 in transgenic mice leads to disparate effects on motor behaviors, followed by a progressive cerebellar dysfunction and histological alterations[END_REF] It remains to be determined to what extent the wire suspension test may contribute to phenotype more precisely SCA1 models.

SCA2

Like SCA1, SCA2 is an autosomal dominant disorder with CAG repeat expansions causing a gain-in-malfunction and leading to Purkinje cell degeneration, limb incoordination, and dysarthria. [START_REF] Alves-Cruzeiro | Motor dysfunctions and neuropathology in mouse models of spinocerebellar ataxia type 2: a comprehensive review[END_REF][START_REF] Imbert | Cloning of the gene for spinocerebellar ataxia 2 reveals a locus with high sensitivity to expanded CAG/glutamine repeats[END_REF] Disease occurs whenever CAG repeats rise above 31 in ATXN2 located on chromosome 12 and encoding ataxin-2. As with the Atxn1 knockout (KO), the Atxn2 mouse KO shows no sign of ataxia, prompting the conclusion that the disease leads to a gain-inmalfunction. [START_REF] Kiehl | Generation and characterization of Sca2 (ataxin-2) knockout mice[END_REF] ATXN2/Q58 mice contain 58 CAG repeats of the transgene driven by the Pcp2 promoter, leading to Purkinje cell loss, paw-clasping, reduced stride length, and rotorod deficits. [START_REF] Huynh | Nuclear localization or inclusion body formation of ataxin-2 are not necessary for SCA2 pathogenesis in mouse or human[END_REF] The mutants also had longer MTs and more foot-slips than wild-type on the stationary beam. [START_REF] Liu | Deranged calcium signaling and neurodegeneration in spinocerebellar ataxia type 2[END_REF] In experimental therapies, ATXN2/Q58 transgenic mice fed dantrolene, a stabilizer of intracellular calcium signaling, exhibited a lesser degree of Purkinje cell loss as well as faster MTs and fewer foot-slips on the stationary beam. Moreover, adenovirus-mediated expression of inositol 1,4,5phosphatase into deep cerebellar nuclei alleviated electrophysiological anomalies in Purkinje cells as well as accelerating MTs and reducing foot-slips on the stationary beam and increasing latencies before falling from the rotorod. [START_REF] Kasumu | Chronic suppression of inositol 1,4,5-triphosphate receptor-mediated calcium signaling in cerebellar purkinje cells alleviates pathological phenotype in spinocerebellar ataxia 2 mice[END_REF] These results indicate that inhibiting enzyme-mediated calcium signals may provide therapeutic benefits in patients. Purkinje cell loss and rotorod impairment were also alleviated after intraventricular transplantation of human mesenchymal stem cells. [START_REF] Chang | Mesenchymal stem cell transplantation ameliorates motor function deterioration of spinocerebellar ataxia by rescuing cerebellar Purkinje cells[END_REF] A second SCA2 transgenic murine model expresses the full-length human ATXN2 gene with 75 CAG repeats controlled by the self Atxn2 promoter. [START_REF] Aguiar | Ubiquitous expression of human SCA2 gene under the regulation of the SCA2 self promoter cause specific Purkinje cell degeneration in transgenic mice[END_REF] ATXN2/Q75 transgenic mouse brains contain Purkinje cell body shrinkage and loss of dendritic arborization and the mice have rotorod deficits. A third transgenic model with a longer expansion, ATXN2/Q127, was generated under control of the Pcp2 promoter. [START_REF] Hansen | Changes in Purkinje cell firing and gene expression precede behavioral pathology in a mouse model of SCA2[END_REF] ATXN2/Q127 mutants have cytoplasmic ataxin-2containing insoluble aggregates in Purkinje cells, Purkinje cell loss, thinning of the cerebellar molecular layer, and rotorod deficits.

A fourth model, the ATXN2/Q72-BAC transgenic mouse, was generated with 72 CAG repeats of the human ATXN2 gene controlled by the endogenous promoter on bacterial artificial chromosomes (BAC). [START_REF] Dansithong | Ataxin-2 regulates RGS8 translation in a new BAC-SCA2 transgenic mouse model[END_REF] ATXN2/Q72-BAC transgenic mice had smaller dendritic arborization and rotorod deficits. A knockin model of SCA2 disease was generated by homologous recombination of embryonic stem cells to replace the endogenous Atxn2 gene with a mutagenized mouse version containing 42 CAG repeats driven by the Atxn2 promoter. [START_REF] Damrath | ATXN2-CAG42 sequesters PABPC1 into insolubility and induces FBXW8 in cerebellum of old ataxic knock-in mice[END_REF] Atxn2/Q42 knockins had cytoplasmic ataxin-2-containing aggregates and rotorod deficits, though without displaying overt ataxia or any change in foot-print analyses, grip strength, or open-field activity. Thus, the rotorod deficit seen in Atxn2/Q42 knockins precedes overt ataxia, as reported in ATXN1/Q82 mice 46 described above.

And so the rotorod and perhaps other motor tests may be used in experimental therapies of SCA2 mutants before dysfunction appears under the naked eye.

SCA3

Like SCA1 and SCA2, SCA3, also called Machado-Joseph disease, is due to a polyglutamine expansion, this time an expansion of over 45 CAG repeats in ATXN3 located on chromosome 14 and encoding ataxin-3. [START_REF] Kawaguchi | CAG expansions in a novel gene for Machado-Joseph disease at chromosome 14q32.1[END_REF][START_REF] Padiath | Identification of a novel 45 repeat unstable allele associated with a disease phenotype at the MJD1/SCA3 locus[END_REF][START_REF] Schöls | Autosomal dominant cerebellar ataxias: clinical features, genetics, and pathogenesis[END_REF] SCA3 causes prominent cerebellar signs such as gait ataxia, dysmetria, and dysarthria, along with parkinsonian signs [START_REF] Coutinho | Autosomal dominant system degeneration in Portuguese families of the Azores Islands. A new genetic disorder involving cerebellar, pyramidal, extrapyramidal and spinal cord motor functions[END_REF] resulting from lesions in cerebellum, pontine nuclei, and substantia nigra pars compacta. [START_REF] Stevanin | Clinical and molecular advances in autosomal dominant cerebellar ataxias: from genotype to phenotype and physiopathology[END_REF] Ataxin-3 binds to the type 1 inositol 1,4,5-trisphosphate receptor, the intracellular calcium release channel. [START_REF] Chen | Deranged calcium signaling and neurodegeneration in spinocerebellar ataxia type 3[END_REF] Several SCA3 mouse models are available, [START_REF] Gould | Mouse models of spinocerebellar ataxia type 3 (Machado-Joseph disease)[END_REF] one of which featuring ATXN3/Q69 transgenic mice expressing the mutant gene driven by the Pcp2 promoter. [START_REF] Torashima | Lentivector-mediated rescue from cerebellar ataxia in a mouse model of spinocerebellar ataxia[END_REF] ATXN3/Q69 mice showed Purkinje cell ectopias and thinner dendrite branching along with overt ataxia and delayed rotorod acquisition, probably due to a gain-in-malfunction mutation, because Atxn3 null mutants display neither Purkinje cell pathology nor ataxia. [START_REF] Switonski | Mouse ataxin-3 functional knock-out model[END_REF] Purkinje cell ectopias and rotorod performance ameliorated in ATXN3/Q69 mice after midline cerebellar injection of lentiviral vectors expressing wild-type CRMP1 (collapsin response mediator protein, CRAG), which degrades polyglutamine proteins via the ubiquitin-proteasome pathway. [START_REF] Torashima | Lentivector-mediated rescue from cerebellar ataxia in a mouse model of spinocerebellar ataxia[END_REF] In addition, integrasedefective lentiviral vectors injected into the cerebellum decreased ataxin-3 aggregates and improved rotorod performance. [START_REF] Saida | One-year follow-up of transgene expression by integrase-defective lentiviral vectors and their therapeutic potential in spinocerebellar ataxia model mice[END_REF] Moreover, ATXN3/Q69 mice had fewer nuclear inclusions and higher cerebellar calbindin immunoreactivity along with thicker molecular and granular layers as well as higher latencies before falling off the rotorod, larger stride length in foot-print analyses, and higher horizontal activity in the open-field after intracerebellar injection of lentiviral vectors of short-hairpin RNAs encoding allele-specific silencing sequences. [START_REF] Nobrega | Silencing mutant ataxin-3 rescues motor deficits and neuropathology in Machado-Joseph disease transgenic mice[END_REF] A second SCA3 model employs the ATXN3/Q71 transgene driven by the murine Prp promoter, exhibiting ataxin-3 nuclear inclusions and substantia nigra pars compacta cell loss combined with ataxia, tremors, seizures, foot-print anomalies, grip strength weakness, open-field hypoactivity, and rotorod deficits. [START_REF] Goti | A mutant ataxin-3 putative-cleavage fragment in brains of Machado-Joseph disease patients and transgenic mice is cytotoxic above a critical concentration[END_REF] A third SCA3 transgenic model, ATXN3/Q79-Pcp2, expresses truncated ataxin-3 with an expanded polyglutamine stretch driven by the Pcp2 promoter. [START_REF] Ikeda | Expanded polyglutamine in the Machado-Joseph disease protein induces cell death in vitro and in vivo[END_REF] ATXN3/Q79-Pcp2 mice have Purkinje cell loss and overt ataxia. A fourth model with the same number of repeats, ATXN3/Q79-Prp, was generated with a transgene driven by the murine Prp promoter. [START_REF] Chou | Polyglutamine-expanded ataxin-3 causes cerebellar dysfunction of SCA3 transgenic mice by inducing transcriptional dysregulation[END_REF] ATXN3/Q79-Prp transgenic mice display mutated ataxin-3 nuclear inclusions in dentate, pontine, and nigral nuclei, an ataxic gait, forelimb clasping, open-field hypoactivity, and rotorod deficits. The fact that mutated ataxin-3 directly causes cell death is shown by findings in cultured cerebellar and nigral neurons. [START_REF] Chou | Polyglutamine-expanded ataxin-3 activates mitochondrial apoptotic pathway by upregulating Bax and downregulating Bcl-xL[END_REF] In these ATXN3/Q79-Prp transgenic mice, intraperitoneal injection of H1152, an inhibitor of rho-kinase (ROCK), decreased ataxin-3 protein concentrations at cerebellar, pontine, and neocortical sites as well as pontine cell death while counteracting open-field hypoactivity and rotorod deficits. [START_REF] Wang | H1152 promotes the degradation of polyglutamine-expanded ataxin-3 or ataxin-7 independently of its ROCK-inhibiting effect and ameliorates mutant ataxin-3-induced neurodegeneration in the SCA3 transgenic mouse[END_REF] Likewise, foot-print analyses of gait, open-field hypoactivity, and rotorod deficits improved after intraperitoneal injection of sodium butyrate, an inhibitor of histone deacetylase, an effect that reverses transcriptional downregulation. [START_REF] Chou | HDAC inhibitor sodium butyrate reverses transcriptional downregulation and ameliorates ataxic symptoms in a transgenic mouse model of SCA3[END_REF] Moreover, pontine nuclei degeneration and rotorod deficits were mitigated after oral administration of an adenosine A2A receptor agonist, T1-11 [N6-(4-Hydroxybenzyl) adenosine], extracted from Gastordia elata, a Chinese medicinal herb, or its synthetic analog, JMF1907 [N6- (3-Indolylethyl) adenosine]. [START_REF] Chou | T1-11 and JMF1907 ameliorate polyglutamine-expanded ataxin-3-induced neurodegeneration, transcriptional dysregulation and ataxic symptom in the SCA3 transgenic mouse[END_REF] An ATXN3 transgene with a longer Q94 expansion driven by the CMV promoter exhibited mutant ataxin-3 inclusions and shrunken cells in cerebellum, pons, and substantia nigra combined with open-field hypoactivity and deficits on rotorod but not vertical pole or foot-print assays. [START_REF] Silva-Fernandes | Motor uncoordination and neuropathology in a transgenic mouse model of Machado-Joseph disease lacking intranuclear inclusions and ataxin-3 cleavage products[END_REF] Thus, the ATXN3/Q94 transgenic resembles the Atxn2/Q42 knockin [START_REF] Damrath | ATXN2-CAG42 sequesters PABPC1 into insolubility and induces FBXW8 in cerebellum of old ataxic knock-in mice[END_REF] and ATXN1/Q82 transgenic [START_REF] Clark | Purkinje cell expression of a mutant allele of SCA1 in transgenic mice leads to disparate effects on motor behaviors, followed by a progressive cerebellar dysfunction and histological alterations[END_REF] which display rotorod deficits with no overt ataxia or anomalies in foot-print analyses.

Polyglutamatine repeats were further expanded with the ATXN3/Q135 transgene also regulated by the CMV promoter. 96 ATXN3/Q135 mutants have ataxin-3 nuclear inclusions, lower dentate nucleus volume and pontine nucleus number, as well as substantia nigra gliosis, causing ataxia, hindlimb clasping, tremor, shorter stride length, weaker grip strength, higher latencies before traversing the stationary beam, and lower latencies before falling off the hanging wire or rotorod. Pontine cell number and ataxin-3 nuclear inclusions as well as stationary beam and rotorod performance improved after intraperitoneal injections of 17-DMAG, inhibitor of Hsp90, a heat shock protein.

Unlike wild-type mice where ataxin-3 is present in the cytoplasm, ATXN3/Q70 and ATXN3/Q148-Prp transgenes controlled by the murine Prp promoter result in brain tissue containing ubiquitin-and ataxin-3-positive nuclear inclusions. [START_REF] Bichelmeier | Nuclear localization of ataxin-3 is required for the manifestation of symptoms in SCA3: in vivo evidence[END_REF][START_REF] Nguyen | Cerebellar soluble mutant ataxin-3 level decreases during disease progression in spinocerebellar ataxia type 3 mice[END_REF] The ATXN3/Q70 model showed shrunken Purkinje cell bodies and limb-clasping; both models displayed overt ataxia and tremor. [START_REF] Bichelmeier | Nuclear localization of ataxin-3 is required for the manifestation of symptoms in SCA3: in vivo evidence[END_REF] A similar ATXN3/Q148-Hd transgene driven by the rat Hd promoter was generated, exhibiting ataxin-3-positive nuclear inclusions in Purkinje and pontine cells along with rotorod deficits. [START_REF] Boy | A transgenic mouse model of spinocerebellar ataxia type 3 resembling late disease onset and gender-specific instability of CAG repeats[END_REF] They also showed home-cage hyperactivity at a young age but yet home-cage hypoactivity at an older age.

Atxn3/Q82 knockin mice were generated by introducing the murine gene with 82 polyglutamine repeats at the mouse locus, accumulating mutant ataxin-3 nuclear inclusions in Purkinje cells, pons, and substantia nigra. 100 However, 1-year old knockin mice did not differ from wild-type on stationary beam, rotorod, or open-field tests. In contrast, ATXN3/Q91 knockin mice generated by introducing the human gene with 91 polyglutamine repeats at the mouse locus exhibited astrogliosis in cerebellum and susbstantia nigra, Purkinje cell loss, and rotorod deficits. 101 Inducible double ATXN3/Q77 transgenic mice were generated by applying the tetracycline-off system under control of the hamster Prp promoter, causing nuclear inclusions, a thinner molecular layer and smaller Purkinje cells, home-cage hyperactivity, limb-clasping, smaller steps in foot-print analyses, and rotorod deficits. 102 The prion protein promoter drives the expression of a fusion protein tTA (tetracycline transactivator) consisting of the tetracycline repressor (TetR) and a transcription activation domain (AD) in the promoter construct. In the absence of tetracycline, the tTA protein can bind to the tetracycline responsive element (TRE) of a CMV minimal promoter in the responder construct, which activates ATXN3 transcription. When tetracycline (Tc) or one of its derivatives such as doxycycline binds to tTA and abolishes the binding of tTA to the responder construct, ATXN3 transcription is blocked. Crossbreeding the PrP promoter line with ATXN3 responder mouse lines yielded double transgenic mice. In responder line 2904 with 77 CAG repeats, nuclear aggregates staining positive for ataxin-3 formed along with a thinner molecular layer and smaller Purkinje cells. The rotorod deficits were reversed by doxycycline. To improve behavior with a n-methyl-d-aspartate (NMDA) glutamate receptor antagonist in ATXN3/Q77 mice, riluzole was dissolved in drinking water, which reduced soluble ataxin-3 levels but also calbindin expression in Purkinje cells and increased ataxin-3 positive accumulations without affecting home-cage hyperactivity and even worsened rotorod performance. 103 Three more SCA3-related models exist, this time with yeast artificial chromosomes (YACs), namely ATXN3/YAC-Q84, ATXN3/YAC-Q67 and ATXN3/YAC-Q15 transgenes under control of endogenous elements. 104 Unlike the expanded (CAG)84 allele, the expanded (CAG)76 allele contracted to 64, 67, and 72 repeats in the original 76 repeats. ATXN3/YAC-Q84 and ATXN3/YAC-Q67, but not ATXN3/YAC-Q15 transgenic mice, exhibited cell loss in pons and dentate nuclei with a wide-spread gait, fore-and hind-limb clasping, and tremor. A second report in ATXN3/YAC-Q84 transgenics revealed fewer than normal neuronal counts in pontine and substantia nigra, higher MTs and foot-slips on the stationary beam and shorter stride length on foot-print patterns. 82 ATXN3/YAC-Q84 transgenic mice fed dantrolene had faster MTs and fewer foot-slips on the stationary beam and elevated stride length on foot-print patterns. Moreover, ATXN3/YAC-Q84 mice had restored Purkinje cell firing and faster MTs on the stationary beam after intraperitoneal injection of an activator of potassium channels, SKA-31. 105 Likewise, ataxin-3 nuclear accumulation was mitigated in ATXN3/YAC-Q84 mice by artificial microRNA mimics targeting the 3'-untranslated region of ATXN3. 106 That mutant ATXN3 can directly affect normal mice is indicated by findings that injected lentiviral vectors encoding full-length human mutant ATXN3 into the mouse cerebellum of C57/BL6 mice causes intranuclear inclusions, neuropathological abnormalities, and neuronal death along with motor coordination deficits, a wide-based gait, and hyperactivity. 107 As in a SCA1 model described above, [START_REF] Clark | Purkinje cell expression of a mutant allele of SCA1 in transgenic mice leads to disparate effects on motor behaviors, followed by a progressive cerebellar dysfunction and histological alterations[END_REF] decreased ataxin-3 aggregates or concentrations occurred in conjunction with improved rotorod performance in four experimental therapies of three SCA3 models. [START_REF] Saida | One-year follow-up of transgene expression by integrase-defective lentiviral vectors and their therapeutic potential in spinocerebellar ataxia model mice[END_REF][START_REF] Nobrega | Silencing mutant ataxin-3 rescues motor deficits and neuropathology in Machado-Joseph disease transgenic mice[END_REF][START_REF] Wang | H1152 promotes the degradation of polyglutamine-expanded ataxin-3 or ataxin-7 independently of its ROCK-inhibiting effect and ameliorates mutant ataxin-3-induced neurodegeneration in the SCA3 transgenic mouse[END_REF][START_REF] Silva-Fernandes | Chronic treatment with 17-DMAG improves balance and coordination in a new mouse model of Machado-Joseph disease[END_REF] The reverse pattern was found in another model, when more ataxin-3 aggregates led to poorer rotorod performance. 103 Thus, ataxin-3 aggregates appear relevant in the context of improving motor functions in SCA3 models.

SCA5

SCA5 consists in autosomal dominant in-frame deletions and missense mutations in SPTBN2 located on chromosome 11 and encoding beta-III spectrin, involved in membrane protein adhesion in cytoplasm, a gene highly expressed in Purkinje cells. [108][109][110] The mutationlinked cerebellar cortical atrophy causes ataxia and dysarthria. SCA5 appears to result from a loss-of-function mutation in view of the ataxia of targeted null mutations of Sptbn2. One Sptbn2 KO murine model exhibited Purkinje cell loss accompanied by wider hindlimb gait, tremor, increased foot-slips on the stationary beam, and decreased latencies before falling off the rotorod, though not off the suspended wire, the latter test being more dependent on muscle strength. 111 A second Sptbn2 KO model exhibited a thinner than normal cerebellar molecular layer, shrunken Purkinje cells, ataxia, and myoclonic seizures as well as wire-suspension and rotorod deficits. 112 Tet-response element (TRE) and CMV promoter were cloned with SPTBN2 cDNA and the resulting mutants bred to tetracycline transactivator (tTA)/Pcp2 transgenic mice to generate conditional bigenics expressing recombinant transgenes specifically in Purkinje cells. 113 Conditional and inducible SPTBN2/double transgenics with a deletion mutation are characterized by a thinner molecular layer of the cerebellar cortex, overt ataxia, and rotorod deficits. The mutants also exhibited reduced mGluR1-alpha localization at Purkinje cell dendritic spines and decreased mGluR1-mediated responses, an indication that beta-III spectrin stabilizes mGluR1 at the cell membrane.

SCA6

SCA6 is another CAG expansion disease, this time caused by mutations of CACN1A located on chromosome 19 and encoding the P/Q-type voltage-gated alpha-1A subunit of a calcium channel, 114-115 causing a cerebellar syndrome marked by postural ataxia as well as limb and ocular dysmetria. 115 Exon 47 undergoes alternative splicing, leading to C-terminal isoforms containing either exon 47 alone or with polyglutamatine repeats. 116 Human C-terminal fragments ending at exon 46 (CT-short) or exon 47 (CT-long) ending with 27 polyglutamine residues were expressed in transgenic mice with CMV and ACTB promoters. 117 Both models had Purkinje cell loss but onset was more precocious in CACNA1A/Q27 mice than CACNA1A/CT-short mice.

CACNA1A/Q27 mice were deficient on stationary beam, inclined screen, and rotorod tests, whereas CACNA1A/CT-short mice were deficient only on the rotorod. Thus, the use of multiple tests helps delineate a more precise phenotype for two mutants of the same gene.

The C terminus of the alpha-1A subunit (alpha-1ACT) is a transcription factor that enhances expression of several Purkinje cell-expressed genes and partially rescued the phenotype of Cacna1a null mutants, whose electrophysiological properties resembled leaner and tottering spontaneous Cacna1a mutants. 118 The alpha1-ACT fragment was expressed via the Pcp2 promoter and Tet-off system yielding Pcp2-tTA/TRE-alpha1-ACT mutants bred with CACNA1A/Q33 mice expressing 33 CAG repeats to generate double transgenic mice.

CACNA1A/Q33 mutants have molecular layer thinning in cerebellum and gait disturbances.

Cacna1a/Q84 knockin mice were generated by introducing the mouse gene with 84 polyglutamine repeats at the mouse locus, causing cytoplasmic inclusions in Purkinje cells and rotorod deficits. 119 Cacna1a/Q84 knockins also displayed Purkinje cell axon anomalies in the form of torpedoes as found in SCA6 patients. 120 Cacna1a/Q84 knockin mice exhibited late-onset loss of Purkinje cell number but early-onset increase in foot-slips on the stationary beam and decreased latencies before falling off the rotorod despite the absence of any change in stride length or stance width. 121 As described in some SCA1, SCA2, and SCA3 mutants, Cacna1a/Q84 knockins displayed rotorod deficits prior to changes in foot-prints. Moreover, the same pattern was found for the first time with the stationary beam. Anomalies in Purkinje cell firing rate were discernable as early as the second postnatal week. 122 A model with longer repeats, Cacna1a/Q118 knockin mice, exhibited cytoplasmic inclusions in Purkinje cells, reduced Purkinje cell number and dendritic branching, short-stepped walking, and rotorod deficits, all of which being absent in a knockin with fewer repeats: Cacna1a/Q11. 123

SCA7

Like SCA types 1, 2, and 3, SCA7 is an autosomal dominant disorder caused by CAG trinucleotide repeats, this time concerning ATXN7 located on chromosome 3 and encoding ataxin-7. [124][125][126][127] Despite widespread distribution of ataxin-7, neurodegeneration mainly occurs in cerebellum, inferior olive, pons, and retinal photoreceptors. In one large cohort of patients, glutamine repeats ranged between 37 and 130, causing ataxia, dysmetria, dysarthria, blindness, and ophthalmoplegia as the principal clinical signs. 125 CAG repeat size was inversely correlated with disease onset, determining 71% of its variability. In patients with equivalent disease durations, CAG repeats were longer in those with blindness, ophthalmoplegia, and extensor plantar reflexes. Ataxin-7 neurodegeneration appears to involve the two main protein degradation pathways in mammalian cells: the ubiquitin-proteasome system and autophagy. 128 The ATXN7/Q52 transgene driven by the PDGFB (platelet-derived growth factor betachain) promoter was generated in mice. There also exists an ATXN7/BAC-Q92 murine model spatially and temporally regulated with cDNA flanked by loxP sites at the start site of translation in murine PrP of a bacterial artificial chromosome. [134][135][136] ATXN7/BAC-Q92 mice displayed Purkinje cell loss, molecular layer thinning, paw-clasping, and rotorod deficits 134,135 The rotorod deficits were reversed by polyglutamine ataxin-7 excision when the single transgenic was transformed into a triple transgenic carrying additional Pcp2-Cre and Gfa2-Cre transgenes. 134 Polyglutamine ataxin-7 excision and reversal of rotorod deficits also occurred after interbreeding ATXN7/BAC-Q92 mice with those expressing tamoxifen-inducible Cre recombinase and administered per os tamoxifen after the onset of motor abnormalities and neuropathology. 136 In addition, cerebellar nuclei injection of recombinant adeno-associated virus vectors expressing interfering RNAs reduced nuclear inclusions in the cerebellum, thickened the cerebellar molecular layer, improved rotorod performance, and mitigated paw-clasping responses in ATXN7/BAC-Q92 mutants. 135 Atxn7/Q266 knockin mice contain 266 CAG repeats into the murine Atxn7 locus with homologous recombination of embryonic stem cells. The resulting mice showed Purkinje cell shrinkage and ataxin-7 aggregates and photoreceptor degeneration combined with ataxia, tremors, myoclonic seizures, and rotorod deficits. 137 Purkinje cell shrinkage, low expression of cerebellar glutamate transporters, and rotorod deficits in Atxn7/Q266 knockin mice were attenuated after intercrossing with mice expressing HGF, that encodes hepatocyte growth factor driven by the NSE (neuron-specific enolase) promoter, the growth factor being normally expressed in Purkinje and granule cells. 138 Moreover, Atxn7/Q266 knockin mice improved after intraperitoneal injection of interferon-beta regarding ataxin-7 nuclear inclusions as well as stationary beam and horizontal ladder tests. 139 Thus, two sensorimotor tests were sensitive to a treatment in conjunction with mitigation of nuclear inclusions in a SCA7 model, as shown for the rotorod in SCA1 and SCA3 models.

In a viral-mediated model, lentiviral vectors express ATXN7 fragments in the mouse vermis. 140 Two months after injection, mutant ATXN7/Q100 but not wild-type ATXN7 driven by the PGK1 (phosphoglycerate kinase-1) promoter caused nuclear ataxin-7 aggregates in cerebellum, shrinkage of granule and molecular layers near the injection site, open-field hypoactivity, missteps on the horizontal ladder, and rotorod deficits reminiscent of the transgene approach.

SCA8

Unlike the polyglutamine-related SCAs described above, SCA8 pathogenesis involves a polyleucine CTG expansion in ATXN8 located on chromosome 13 and encoding ataxin-8. 141 SCA8 results in ataxia, dysmetria, and nystagmus. 142,143 Although several lines of evidence implicate a gain-in-malfunction, 144 Atxn8 null mutants limited to Purkinje cells exhibited a thinner cerebellar molecular layer and rotorod deficits. 145 The authors hypothesize that the human disease leads to a gain-in-malfunction in some areas but a lost in function of ATXN8 in Purkinje cells. Another SCA8 mouse model was generated in which fulllength human ATXN8 with 116 CAG repeats was transcribed via its endogenous promoter.

ATXN8/Q116 transgenic mice developed nuclear inclusions in Purkinje and basal pontine neurons, a loss in cerebellar molecular layer inhibition, and rotorod deficits. 146 3.8 SCA10 SCA10 pathogenesis is caused by autosomal dominant transmission of an intronic ATTCT pentanucleide repeat in ATNX10 located on chromosome 22 and encoding ataxin-10. 147,148 The gain-in-malfunction leads to ataxia, intention tremor, dysmetria, dysarthria, nystagmus, and seizures. [149][150][151] McFarland and Ashizawa 152 reviewed the two existing mouse models of SCA10 (Table 3). A model containing the ATNX10 transgene with 500 ATTCT repeats driven by the rat Eno2 (neuronal enolase) promoter suffered from reproductive unfitness. 153 A second transgenic model containing the ATNX10 transgene with 500 ATTCT repeats driven by the rat Prp promoter was fit for reproduction and showed pontine (though no cerebellar) neuropathology, ataxia, hindlimb clasping, seizures, open-field hypoactivity, and shorter steps as well as greater variability in step width on foot-print analyses but normal rotorod testing. 154 This constitutes the reverse pattern of SCA1, SCA2, SC3, SCA6, and SCA7 models when anomalies of rotorod testing occurred in nonataxic mice. It would be most useful to test this SCA10 model in other sensorimotor tests to gauge the sensitivity of these tests relative to the rotorod, often appearing as the standard in cerebellar models.

SCA13

SCA13 is caused by dominant-negative missense mutations in KCNC3 located on chromosome 19 and encoding Kv3.3, a potassium voltage-gated channel of the Kv3 subfamily. 155 SCA13 may be of childhood-or adult-onset characterized by ataxia, dysarthria, and seizures. 156 The F363L mutation in Kcnc3 of zebrafish, corresponding to human infant-onset F448L, caused axonal path-finding errors in primary motor neurons. 157 Lentiviral expression of Kcnc3 harboring the R424H missense mutation slowed down dendritic development and caused death of murine cultured Purkinje cells. 158 Null mutant mice lacking Kcnc3 had increased lateral deviation and foot-slips on the stationary beam. [159][160][161] The motor deficits were mitigated after selective restoration of Kv3.3 channels in Purkinje cells. 161

SCA14

 SCA14 is caused by autosomal dominant-negative missense mutations in PRKCG located on chromosome 19 and encoding the gamma isoform of protein kinase C (PKC-gamma). 162,163 SCA14 is characterized by cerebellar ataxia, dysarthria, and nystagmus. 164 The PRKCG transgene with the dominant-negative H101Y mutation driven by the CMV promoter yields mice with lower cerebellar PKC activity and Purkinje cell loss, a possible cause of their paw-clasping responses. 165 In another model, viral vectors containing the PRKCG-Green fluorescent protein transgene with the dominant-negative S119P mutation were injected into developing cerebellum, forming mutant aggregates in Purkinje cells and climbing fiber polyinnervation of Purkinje cells. 166 The first vector expresses tetracycline (tet) transactivator (tTA) under control of the Pcp2 promoter. The second set of vectors express respective genes driven by a tet-responsive promoter transactivated by tTA. Because mutant PKC-gamma colocalized with wild-type PKC-gamma, the mutation probably acts in a dominant-negative manner on wild-type PKC. Despite normal locomotion in their home-cage, one-week-old but not 1-month old PRKCG/S119P mutants had rotorod deficits.

SCA15

SCA15 is due to autosomal dominant deletions of ITPR1 located on chromosome 3 and encoding inositol 1,4,5-triphosphate receptor 1, permitting calcium release from intracellular stores. [167][168][169] SCA15 causes a relatively pure cerebellar syndrome particularly affecting the vermis and marked by ataxia, intention tremor, and nystagmus. 170,171 A targeted Itpr1 null mutation led to ataxia and tonic or tonic-clonic seizures. 172 In addition, two spontaneous murine mutants with Itpr1 deletions have been discovered. The opisthotonos (opt) mouse is characterized by low inositol 1,4,5-trisphosphate receptor 1 levels, abnormal calcium responses in Purkinje cells, ataxia, and convulsions. 173 A second spontaneous deletion of Itpr1 also decreases inositol 1,4,5-triphosphate receptor 1 levels in Purkinje cells and causes ataxia. 168 3.12 SCA17 SCA17 is an autosomal dominant disorder caused by over 43 CAG repeats in TBP located on chromosome 6 and encoding the TATA-binding protein, a transcription factor for many genes. 174,175 SCA17 is characterized by ataxia, nystagmus, dystonia, and, unlike other SCA diseases, prominent neuropsychiatric signs and dementia. 176 SCA17 murine models were reviewed. 177 Mice were generated with the TBP/Q transgene containing 109 or 69 CAG repeats driven by the Pcp2 promoter. 178 TBP/Q109 transgenic mice sustained losses in Purkinje cells and brainstem but also basal ganglia and neocortex, leading to gait anomalies on foot-print analyses, paw-clasping, open-field hyperactivity, and rotorod deficits. Mutated TBP aggregates were colocalized in Purkinje cells with ubiquitin and heat shock chaperone Hsc70. Purkinje cell loss, open-field hyperactivity, and rotorod deficits were mitigated by subcutaneous injection of granulocyte-colony stimulating factor to mobilize blood progenitor cells, its receptor being expressed on Purkinje cells. 178,179 Likewise, Purkinje cell misalignment and rotorod deficits were mitigated by intraperitoneal injection of an extract from the leaves of Ginkgo biloba, EGb 761. 180 Inducible TBP/Q105 knockin mice were generated by replacing exon 2 of the murine Tbp gene with exon 2 of human TBP carrying 105 CAGs. 181 The stop codon is flanked by two loxP sites as a targeting vector. The floxed mice were crossed with those carrying a rat Nes (nestin) promoter-driven Cre transgene and the stop codon removed via Cre-loxP recombination, resulting in knockin mice which express mutant and normal TBP under the influence of the endogenous promoter. TBP/Q105 knockins exhibited nuclear aggregates in Purkinje cells, Purkinje cell loss, home-cage hypoactivity, and rotorod deficits. To induce the expression of mutated mice at different ages, floxed heterozygous TBP/Q105 knockin mice were crossed with CreER transgenic mice expressing a fusion protein of Cre recombinase with an estrogen receptor ligand binding site driven by the chicken Actb (β-actin) promoter. The intraperitoneal injection of tamoxifen, an estrogen receptor ligand, binds the Cre recombinase fusion protein and makes it enter the nucleus to act on loxP sites, removing the stop codon. 182 Tamoxifen-inducible TBP/Q105 knockin mice exhibited Purkinje cell loss, reduced stride length on foot-print patterns, paw-clasping, slower MTs on the stationary beam, and rotorod deficits. The Purkinje cell loss was mitigated by intracerebellar injection of lentivral vectors expressing mesencephalic astrocyte-derived neurotrophic factor (MANF) or after interbreeding with MANF transgenic mice, the latter method also improving stationary beam, paw-clasping, and foot-print anomalies.

In addition to murine models, a rat model of SCA17 is available with 64 CAA/CAG repeats driven by the murine Prp promoter. 183 TBP/Q64 transgenic rats displayed nuclear aggregates in granule, Purkinje, and stellate cells, Purkinje cell loss, paw-clasping, and more foot-slips than wild-type on the stationary beam but no effect on rotorod acquisition. This result points again to the usefulness of the stationary beam as a test to be used in conjunction with the rotorod. Relative to wild-type, they also showed home-cage hyperactivity at a young age and hypoactivity at an older age, but with decreased rearing throughout.

SCA27

SCA27 is an autosomal dominant-negative disorder caused by mutations in FGF14 located on chromosome 14 and encoding fibroblast growth factor 14, an accessory subunit of voltage-gated sodium channels. [184][185][186] SCA27 causes a combination of cerebellar and basal ganglia symptoms including ataxia, dysarthria, nystagmus, tremors, and dyskinesia. 187 In normal mice, Fgf14 transcripts are expressed in cerebellar granule and dorsal striatal cells. 188 In the cerebellar slice preparation of Fgf14 null mutants, AMPA receptor-mediated excitatory postsynaptic currents evoked by parallel fiber stimulation were lower than those of wild-type. 188 Fgf14 null mutant mice exhibited overt ataxia, dyskinesia, paw-clasping, reduced grip strength, open-field hyperactivity, and slowed MT before reaching the top of an inclined screen as well as shortened latencies before falling from an inverted screen or rotorod. 189,190 The dyskinesia appears in the form of paroxysmal forelimb clonic spasms with hyperextended hindlimbs.

SCA28

SCA28 is an autosomal dominant-negative disorder caused by mutations in AFG3L2 located on chromosome 18 and encoding part of the mitochondrial ATPase-associated protease superfamily. 191,192 The neuropathology comprises a loss in cerebellar and brainstem volumes along with extraocular muscle atrophy. 193 SCA28 is characterized by ataxia, dysarthria, nystagmus, and ophthalmoparesis. 194 Homozygous Afg3l2 null mutant mice die on the second postnatal week. In heterozygous Afg3l2 null mutants, ATP synthesis in brain is defective due to insufficient assembly of respiratory complexes I and III. 195 Afg3l2 haploinsufficiency led to granule and Purkinje cell loss, an impaired negative geotaxic response (turning upward on an inclined grid), limb-clasping, more foot-slips than normal on the stationary beam, and rotorod deficits. 196 Purkinje cell degeneration and foot-slips diminished after interbreeding Afg3l2 haploinsufficient mice with Grm1 haploinsufficient mice lacking mGluR1, which prevents high calcium influx into Purkinje cells. 197

Concluding remarks

The present overview on SCA models illustrates the methodology used by researchers to determine behavioral phenotypes of cerebellar dysfunction. The cerebellum is intrinsically linked to ataxia and deficits in motor coordination since the 1917 paper by Holmes, who described the consequences of human gunshot wounds. 198 The main function of the cerebellum is to integrate sensory input with motor output, to modulate movements, not cause them, so that lesions of this area cause neither sensory deficit (hypoesthesia, blindness, or deafness) nor paralysis.

Spontaneous cerebellar mutants are featured prominently in the "Catalog of the neurological mutants of the mouse", a 1965 monograph by Sidman et al,199 because a new cerebellar mutation is all the easier to detect in a cage, even by uninformed laboratory personnel, as a result of the obvious lurching and swaying mice, in contrast to other neurologic signs that might go undetected or are even undetectable on casual viewing. Moreover, many mutations affecting the cerebellum are embryologically non-lethal and the mutant is not generally attacked by the non-mutant in their home cage. In a laboratory maximizing animal protection, the mutation permits a relatively long life span and the possibility of analyzing behavior from early development to old age. We pointed out the use of limb clasping 200 and myoclonic jumping 201 as additional features in phenotyping neurologic mutants, including those affecting the cerebellum.

In addition to qualitative observations of mutants, the need arose to provide quantified measurements of motor coordination, among which the rotorod gained prominence after the original 1962 paper by Plotnikoff et al 202 and the 1968 paper by Jones and Roberts. 203 As shown in the present review, the rotorod is the most common sensorimor test used to evaluate SCA models. Indeed, rotorod deficits were reported in 10 SCA models, namely SCA1 to 3, SCA5 to 8, SCA14, SCA17, and SCA27. Stationary beam deficits were described in 8 SCA models, namely SCA1 to 3, SCA5, SCA6, SCA13, SCA17, and SCA27, but wire suspension deficits in only SCA3 and SCA5. SCA1, SCA2, SCA3, SCA6, SCA7, and SCA14 models showed rotorod defects without overt ataxia or anomalous foot-print patterns. The reverse pattern of abnormal foot-print patterns and normal rotorod performance was found only in a SCA10 model. Moreover, a SCA6 model without ataxia was deficient on the stationary beam and a SCA17 model was deficient on the stationary beam but not on the rotorod, which should promote the use of the stationary beam as a test to be used in conjunction with the rotorod.

The rotorod task may be less time-consuming than other tasks in that ataxic mice are more likely to fall early, whereas early falls sometimes occur only from the narrowest stationary beam, which may cause mutants to wrap their limbs around it to prevent a fall. In the latter case, MT and distance travelled may be used as additional measures. The limb-wrapping strategy also occurs on the suspended wire, so that MT can be added on the coat-hanger version of the test.

Passive rotation on the rotorod can be counteracted by removing mice from the apparatus as soon as the behavior appears and considering it akin to a fall. [START_REF] Lalonde | Sensorimotor learning in Dab1 scm (scrambler) mutant mice[END_REF] All three tests are complimented by foot-print analyses to document the existence of an ataxic gait.

The question arises as to what extent stationary beam, suspended wire, and rotorod tests are selective for a cerebellar lesion relative to lesions elsewhere in the brain. The answer is that rotorod deficits have been reported after selective lesions of cerebellar afferent or efferent regions such as the inferior olive 204 and ventrolateral thalamus, 205 respectively, but also parts of the basal ganglia such as the substantia nigra pars compacta 206 or lateral pallidum. 207 Likewise, cerebellar 208,209 or dorsostriatal 210 lesions impair stationary beam performance, though the suspended wire has been less extensively investigated. 211 Therefore, current sensorimotor tests dependent on postural control seem meant to detect a central nervous system deficit rather than a specifically cerebellar-related one.

  129 ATXN7/Q52 mice displayed shrunken Purkinje cells with ataxin-7 nuclear inclusions and reduced dendritic arborization combined with overt ataxia, hindpaw clasping, open-field hypoactivity, and rotorod deficits. Two longer CAG expansions have been generated, ATXN7/Q92 transgenes expressed under control of either hamster Pcp2 or human RHO (rhodopsin) promoters to affect either Purkinje cells or photoreceptors but accumulating ataxin-7 immunoreactive aggregates in each respective cell type. 130 ATXN7/Q92-Pcp2 transgenic mice exhibited reduced Purkinje dendrite arborization without ectopias and rotorod deficits without overt ataxia. Another transgenic line expresses the same number of repeats but only in Bergmann glia of the cerebellum via the Gfa2 promoter. 131 ATXN7/Q92-Gfa2 mice developed Purkinje cell misalignment and dendrite thinning, ataxia, paw-clasping, and rotorod deficits, implicating glial dysfunction in polyglutaminemediated ataxia. Another murine model with 92 CAG repeats, this time driven by the Prp promoter was generated, ATXN7/Q92-Prp, expressing polyglutamine-expanded ataxin-7 throughout the CNS except in Purkinje cells. 132 Nevertheless, ATXN7/Q92-Prp developed shrunken Purkinje cells with diminished dendritic branches and rotorod deficits, indicating noncell autonomous cell degeneration in SCA7 pathogenesis. The accumulation of ataxin-7 immunoreactive neurons was observed in two other lines expressing 10 or 128 CAG repeats (ATXN7/Q10 or ATXN7/Q128) generated with the PDGFB promoter, the latter line exhibiting ataxia. 133

Figure legend Fig 1 a

 legend1 Figure legend

Table 1

 1 Neurobehavioral characteristics of natural mouse mutations with cerebellar atrophy

	Mutant	Chromosome	Neuropathology	Motor test	Genetic reference
	Grid2 Lc	6	Purkinje cell loss	!	Caddy and Biscoe 6
				stationary beam
				coat-hanger
				rotorod
	Grid2 ho	6	granule cell loss	!	Guastavino et al 10
				stationary beam
				coat-hanger
				rotorod
	Rora sg	9	Purkinje cell loss	!	Herrup and Mullen 22
				stationary beam
				coat-hanger
				rotorod
	Agtpbp1 pcd 13	Purkinje cell loss	!	Mullen et al 28
				coat-hanger
				rotorod
	Reln rl	5	granule cell loss	!	Mariani et al 34

Table 2

 2 Neurobehavioral characteristics of SCA1 to SCA8 mouse mutations with cerebellar atrophy on sensorimotor tests

	Mutant	Promoter	Neuropathology	Motor test	Identifier
					reference
	ATXN1/Q82	Pcp2	Purkinje cell loss	!	SCA1
				stationary beam	Burright et al 45
				rotorod
	ATXN1/Q30	Pcp2	Purkinje cell loss	?	SCA1
					Fernandez-
					Funez et al 48
	Inducible	Pcp2	ectopic Purkinje	!	SCA1
	ATXN1/Q82		cells	rotorod	Zu et al 57
	Atxn1/Q154	endogenous	Purkinje cell loss	!	SCA1
				rotorod	Watase et al 58
	Atxn1/Q78	endogenous	no atrophy	!	SCA1
				rotorod	Lorenzetti et al 59
	ATXN2/Q58	Pcp2	Purkinje cell loss	!	SCA2
				stationary beam	Huynh et al 69
				rotorod
	ATXN2/Q75	ATXN2	Purkinje soma	!	SCA2
			shrinkage	rotorod	Aguiar et al 73
	ATXN2/Q127	Pcp2	Purkinje cell loss	!	SCA2
				rotorod	Hansen et al 74
	ATXN2/Q72-BAC	ATXN2	reduced Purkinje	!	SCA2
			cell dendrite	rotorod	Dansithong et
			branches		al 75
	Atxn2/Q42	Atxn2	ataxin-2	!	SCA2
			aggregates	rotorod	Damrath et al 76
	ATXN3/Q69	Pcp2	Purkinje cell	!	SCA3
			ectopias, thin	rotorod	Torashima et
			dendrite branches		al 84
	ATXN3/Q71	Prp	nuclear inclusions	!	SCA3
				rotorod	Goti et al 88
	ATXN3/Q79-Pcp2	Pcp2	Purkine cell loss	?	SCA3
					Ikeda et al 89
	ATXN3/Q79-Prp	Prp	nuclear inclusions	!	SCA3
				rotorod	Chou et al 90
	ATXN3/Q94	CMV	shrunken cells	!	SCA3
				rotorod	Silva-Fernandes

Table 3

 3 Neurobehavioral characteristics of SCA10 to SCA28 mouse mutations with cerebellar atrophy on sensorimotor tests

	Mutant	Promoter	Neuropathology	Motor test	Identifier reference
	ATNX10 KO	not applicable pontine nuclei loss	?	SCA10
					White et al 154
	Kcnc3 KO	not applicable	?	!	SCA13
				stationary beam	McMahon et al 159
	PRKCG/H101Y	CMV	Purkinje cell loss	?	SCA14
					Zhang et al 165
	Inducible	Pcp2	climbing fiber	!	SCA14
	PRKCG/S119P		poly-innervation of	rotorod	Shuvaev et al 166
			Purkinje cells
	Itpr1 KO	not applicable	?	?	SCA15
					van de Leemput et
					al 168
	Itpr1 KO	not applicable	?	?	SCA15
					Matsumoto et al 172
	opt	not applicable	abnormal calcium	?	SCA15
			responses in		Street et al 173
			Purkinje cells
	TBP/Q109	Pcp2	Purkinje cell loss	!	SCA17
				rotorod	Chang et al 178
	Inducible	endogenous	Purkinje cell loss	!	SCA17
	TBP/Q105			stationary beam	Huang et al 181
				rotorod
	TBP/Q64	Prp	Purkinje cell loss	!	SCA17
				stationary beam	Kelp et al 183
				NE rotorod
	Fgf14	not applicable	parallel fiber	!	SCA27
			anomalies	inclined screen	Wang et al 189
				inverted screen
				rotorod
	Afg3l2	not applicable	granule and	!	SCA28
			Purkinje cell loss	inclined grid	Maltecca et al 196
				stationary beam
				rotorod