C16 PRION DISEASES: A POTENTIAL LESSON FOR UNDERSTANDING THE PATHOGENESIS OF ALS
KOPITO R
Stanford University, Palo Alto, CA, USA
Email address for correspondence: [email protected]
Keywords: prion, aggregation
Sequence-specific nucleated protein aggregation underlies the pathogenesis of most neurodegenerative diseases and constitutes the molecular basis of prion formation. Nevertheless, prion disorders have been distinguished from classical neurodegenerative diseases by virtue of their ability to be transmitted between individuals. In this lecture I will argue that prion-like propagation of pathogenic aggregated forms can explain the well-documented stereotypical spread of disease pathology in neurodegenerative disorders such as Huntington's, Lou Gehrig's, Alzheimer's and Parkinson's diseases. I will present data demonstrating that fibrillar polyglutamine aggregates like those associated with Huntington's disease can be internalized by mammalian cells in culture where they gain access to the cytosolic compartment and become co-sequestered in aggresomes together with components of the ubiquitin-proteasome system and cytoplasmic chaperones. I will also present recent unpublished data examining the biochemical and biophysical properties of protein aggregates and cell membranes that are necessary for cytoplasmic intrusion of aggregates and the implications for the pathogenesis and management of this class of conformational disease.
C17 MOLECULAR PATHOGENESIS AND TRANSLATIONAL RESEARCH IN SPINAL AND BULBAR MUSCULAR ATROPHY (SBMA)
SOBUE G1, KATSUNO M1, ADACHI H1, BANNO H1,2, SUZUKI K1
1Nagoya University Graduate School of Medicine, Nagoya, Japan, 2Institute for Advanced Research, Nagoya University, Nagoya, Japan
Email address for correspondence: [email protected]
Keywords: SBMA, polyglutamine, androgen receptor
Spinal and bulbar muscular atrophy (SBMA) is an adult-onset lower motor neuron disease characterized by slowly progressive muscle weakness and atrophy. The cause of this disease is the expansion of a trinucleotide CAG repeat, which encodes the polyglutamine tract, within the first exon of the androgen receptor (AR) gene. Expanded polyglutamine tracts have been found to cause several neurodegenerative diseases including SBMA, Huntington's disease, several forms of spinocerebellar ataxia, and dentatorubral-pallidoluysian atrophy. In these disorders, known as polyglutamine diseases, the CAG repeat has a strong tendency to further expand, accelerating the disease onset with in successive generations.
SBMA exclusively occurs in adult males, whereas both heterozygous and homozygous females are usually asymptomatic. As for a transgenic mouse model of SBMA expressing the full-length human AR containing 97 CAGs (AR-97Q), neuromuscular symptoms are markedly pronounced and accelerated in the male mice, but either not observed or far less severe in the female counterparts. Androgen deprivation through surgical castration substantially improved the symptoms, histopathological findings, and nuclear accumulation of the pathogenic AR in the male AR-97Q mice. In contrast, subcutaneous injection of testosterone causes significant aggravation of symptoms, histopathological features, and nuclear localization of the pathogenic AR in the female AR-97Q mice. Since the nuclear translocation of AR is ligand-dependent, testosterone appears to show toxic effects by accelerating nuclear translocation of the pathogenic AR. Lending support to the ligand-dependent hypothesis are the clinical observations that manifestation of symptoms is minimal even in the females homozygous for an expanded CAG repeat in the AR gene, and that testosterone administration exacerbates neuromuscular symptoms in a patient with SBMA. In a large-scale randomized-controlled trial of clinical trial, leuprorelin treatment was associated with a greater reduction in barium residue than was placebo in patients with a disease duration less than 10 years difference between groups.
The ligand-dependent accumulation of the pathogenic AR, an initial step in the neurodegenerative process in SBMA, is followed by several downstream molecular events such as transcriptional dysregulation, axonal transport disruption, and mitochondrial insufficiency, indicating that both upstream and downstream molecular events should be targeted to therapy development. Although the precise mechanism by which motor neurons die remains unclear, activation of cellular defence reactions, ubiquitin-proteasome system, autophagy and heat shock proteins, has been shown to alleviate disease progression in animal models of SBMA. Restoration of transcriptional activity through histone acetylation is also capable of suppressing neurodegeneration in SBMA mice. Development of combination therapies will be the key for the translational research in SBMA.
C18 MECHANISMS OF PERIPHERAL AXONAL NEURODEGENERATION IN SPINAL MUSCULAR ATROPHY
FARRAR M1,2, VUCIC S1, LIN C1, PARK S1, KIERNAN M1
1Neurosciences Research Australia and University of New South Wales, Randwick, NSW, Australia, 2Sydney Children's Hospital, Randwick, NSW, Australia
Email address for correspondence: [email protected]
Keywords: spinal muscular atrophy, axon, excitability
Background: Spinal muscular atrophy (SMA) is a disorder of spinal motor neurons characterized clinically by the development of muscle weakness and atrophy. The ‘up-front’ clinical course suggests a substantial early loss of motor neurons followed by increasing stability of the surviving neurons with slow or no clinical deterioration and is unusual for a neurodegenerative disease. The mechanisms underlying this spectrum of differential survival and potential compensation of motor neurons in SMA remain unknown.
Objectives: To gain insights into axonal biophysical properties, disease pathogenesis and potential adaptations in SMA, the present study utilised clinical and functional assessments, combined with axonal excitability studies.
Methods: Axonal excitability studies were undertaken in 25 genetically characterized adolescent and adult SMA patients, stimulating the median motor nerve at the wrist. Multiple excitability indices (stimulus-response curve, strength-duration time constant, threshold electrotonus, current-threshold relationship and recovery cycle) were compared with 50 age-matched controls. Neurophysiological parameters were correlated with clinical and functional measures of disease severity, namely the MRC sum score and Spinal Muscular Atrophy Functional Rating Scale (SMAFRS) in SMA patients.
Results: In SMA patients there were reductions in CMAP amplitude (P < 0.0005) associated with reduction in stimulus response slope (P < 0.0005), confirming significant axonal loss. In the mild or ambulatory SMA patients, there was reduction of peak amplitude without alteration in axonal excitability; in contrast, in the non-ambulatory or severe SMA cohort prominent changes in axonal function were apparent. Specifically, there were steep changes in the early phase of hyperpolarisation in threshold electrotonus (P < 0.0005) that correlated with clinical severity. Additionally there were greater changes in depolarizing threshold electrotonus (P < 0.0005) and prolongation of the strength-duration time constant (P = 0.001). Mathematical modelling of the excitability changes in severe SMA patients supported a mixed pathology comprising features of axonal degeneration and regeneration.
Discussion and conclusions: The present study has established dysfunction of axonal K+and Na+ conductances and alterations in passive membrane properties in SMA patients, supporting a mixed pathology of degeneration and regeneration. Importantly, excitability changes were most abnormal in the most clinically affected patients, critically indicating that the excitability changes relate to the process neurodegeneration and compensatory partial regeneration.
C19 ROLE OF SMN PROTEIN IN MOTOR NEURON DEGENERATION AND PROTECTION
TURNER B1, ALFAZEMA N2, DAVIES K2, HORNE M1, TALBOT K2
1Florey Neuroscience Institutes, Melbourne, Australia, 2MRC Functional Genomics Unit, University of Oxford, Oxford, UK
Email address for correspondence: [email protected]
Keywords: survival motor neuron, spinal muscular atrophy, SOD1
Background: SMA results from insufficent levels of survival motor neuron (SMN) protein in spinal motor neurons and skeletal muscle. Reduced levels of SMN protein have been reported in mutant SOD1 models of ALS and recently spinal cords of sporadic ALS patients, in line with genetic association studies showing that low SMN levels may increase susceptibility to and/or severity of ALS. These findings raise the interesting prospect that SMN upregulation may be therapeutic in ALS which we tested here in transgenic ALS model mice.
Objectives: To investigate the effect of SMN upregulation on disease progression and neurodegeneration in transgenic SOD1G93A mice.
Methods: Transgenic SOD1G93A mice on either B6SJL or B6 congenic backgrounds were crossed with mice overexpressing human SMN driven by the prion protein promoter (PrP-SMN). Double transgenic SOD1G93A PrP-SMN mice and control genotypes SOD1G93A and PrP-SMN were examined for weight loss, motor function, disease progression and survival. Spinal cords were analysed by motor neuron counts, SMN immunohistochemistry and biochemical subcellular fractionation for SMN levels.
Results: Transgenic full-length SMN was overexpressed 2-fold in postnatal brains and spinal cords of PrP-SMN mice. In SOD1G93A PrP-SMN mice, human SMN overexpression was maintained up to 60 days, but depleted from 90 days, in accordance with endogenous Smn in SOD1G93A mice. SMN upregulation significantly delayed body weight decline, disease onset and preserved spinal motor neurons in SOD1G93A PrP-SMN animals, despite limited effects on symptom progression. Elevated SMN levels in cytoplasmic and nuclear fractions were confirmed in spinal cords of doubly transgenic mice. In contrast to PrP-SMN mice, SMN-positive gems were drastically reduced in motor neurons in SOD1G93A and SOD1G93A PrP-SMN animals.
Discussion and conclusions: These results suggest that SMN protein depletion in spinal motor neurons may be a determinant of neuronal vulnerability and loss in models of ALS. SMN upregulation using a transgenic approach protects against early phases of disease and neurodegeneration in mutant SOD1 mice. The mechanisms underlying protection of motor neurons by overexpressed SMN in this model is currently under investigation.