P215 ASSESSING COGNITIVE CHANGES IN ALS: FROM A NOVEL BEHAVIORAL TEST TO ITS NEURAL CORRELATES
YA Yunusova1,2
S Gillingham1,4
J Ansari2
CJM Scott2
A Ganda2
D Stuss1,3
S Black1,2
L Zinman1,2
aUniversity of Toronto, Toronto, Canada
bSunnybrook Research Institute, Toronto, Canada
cOntario Brain Institute, Toronto, Canada
dRotman Research Institute, Baycrest, Toronto, Canada
Email address for correspondence: [email protected]
Keywords: cognitive testing, MRI, frontal lobe function
Background: ALS is viewed as a multisystem disorder (Citation1), yet its cognitive changes are difficult to identify in the clinic largely due to the relatively mild nature of the deficits, limitations of the standardized neuropsychological assessments in patients with severe motor impairments, and the lack of agreement regarding the nature and origin of the cognitive decline.
Objectives: The goals of this study are: A) To assess sensitivity of a novel Computerized Frontal Battery (ALS-CFB), which links cognitive processes to specific brain regions (Citation2), against a standard neuropsychological battery; and B) To measure the association between cognitive impairment and structural changes in the brain.
Methods: Eighteen patients with ALS and the same number of matched healthy controls were recruited. Ten patients with ALS returned for a repeat assessment in approximately 8 months. Structural MR included T1W (3D) and interleaved 2D dual TSE PD/T2. A novel Semi-Automated Brain Region Extraction (SABRE) approach (Citation3) was used to perform a volumetric analysis in the gray (GM) and white (WM) matter as well as sulcal and ventricular CSF and subcortical hyperintensities of 30 segmented regions. Cognitive testing included ALS-CFB, composed of tests sensitive to frontal processes such as energization (i.e., ability to initiate and sustain a response), executive functioning, emotion regulation and metacognitive processing, as well as the standardized neuropsychological battery.
Results: ALS-CFB detected changes in cognition when the standardized testing failed to do so. Significant atrophy was observed in the GM of the inferior frontal (IF) and anterior temporal (AT) regions as compared to the controls (p < 0.05). Longitudinally, the atrophy was seen in the GM in the medial inferior frontal cortex bilaterally and in the WM in the middle frontal (MF), inferior frontal (IF) and medial middle frontal (MMF) regions on the right. Sulcal CSF increased in the same regions. Significant correlations were observed between GM atrophy in the IF region and reaction time and anti-saccades testing and AT atrophy correlated with digit forward task across participants. Longitudinally, IF and MF atrophy was correlated with changes in verbal fluency and reaction time.
Discussion and conclusions: ALS-CFB detected impairments in energization, motor inhibition as measured by saccades, and executive function in the ALS group. These changes were missed by the standard neuropsychological battery. Additionally, MR imaging revealed volumetric changes in the GM, WM and sulcal CSF predicted by the frontal lobe model loci (IF, MF) (Citation2). The results demonstrate the sensitivity and utility of the ALS-CFB in detecting region specific frontal lobe dysfunction, which is correlated to structural changes on neuroimaging.
References
- Strong MP. . Amyotroph Lateral Scler 2009;10: 131–146.
- Stuss DT. J. Int. Neuropsych. Soc, 2011;17, 759–65.
- Ramirez J. . Neuroimage, 2011; 54(2): 963–73.
P216 FRONTOTEMPORAL PATTERN OF NEURODEGENERATION IN AMYOTROPHIC LATERAL SCLEROSIS: A CORTICAL THICKNESS 3T MRI STUDY
F Trojsi1,2
A D'Ambrosio1,2
A Gallo1,2
F Esposito2,3
D Corbo2
A Sagnelli1
G Piccirillo1
G Tedeschi1,2
MR Monsurrò1,2
eDepartment of Neurological Sciences, Second University of Naples, Naples, Italy
fMRI Research Center SUN-FISM, Neurological Institute for Diagnosis and Care “Hermitage Capodimonte”, Naples, Italy
gDepartment of Neuroscience, University of Naples “Federico II”, Naples, Italy
Email address for correspondence: [email protected]
Keywords: MRI, cortical thickness, frontotemporal lobar degeneration (FTLD)
Background: The extensive application of advanced magnetic resonance imaging (MRI) techniques has undoubtedly improved our knowledge of ALS pathophysiology. Nevertheless, the actual spread of the neurodegenerative process throughout the central nervous system is not fully understood.
Objectives: The aim of the present study was to assess the spatial distribution of cortical damage in ALS, by using a cortical thickness (Cth) measurement approach.
Methods: Automatic surface-based Cth measurements were performed on structural 3T MRI data of 20 ALS patients and 18 matched healthy controls (HC) in a case-control study design. Clinical scores of disability and disease progression were correlated with Cth measures.
Results: Comparing patients with controls, we observed significant cortical thinning mainly in frontotemporal areas, bilaterally, including the primary motor, lateral frontal and prefrontal, medial frontal, temporal and parieto-occipital cortices. Furthermore, cortical thinning was significantly related to i) ALS Functional Rating Scale-Revised (ALSFRS-R) score (an index of disease disability) in a left lateral frontal area (p = 0.0356), ii), disease progression rate in a left medial temporal area (p = 0.0265), and iii) to disease duration in a right medial frontal area (p = 0.0072).
Discussion and conclusions: Cortical thinning of the primary motor cortex might be a diagnostic marker for upper motor neuron degeneration in ALS. The correlations found between cortical thinning of frontotemporal areas and clinical measures of disability and disease progression as well as disease duration might provide a further evidence of the progressive development in ALS of a multisystem disorder within the spectrum of frontotemporal lobar degeneration (FTLD).
P217 CORTICAL THICKNESS IN AMYOTROPHIC LATERAL SCLEROSIS AND FRONTO-TEMPORAL DEMENTIA
C Schuster1
E Kasper1
J Machts4
N Naue4
D Bittner5
J Kaufmann5
R Benecke2
S Teipel1,3
S Vielhaber5,4
J Prudlo2,1
hGerman Center of Neurodegenerative Diseases (DZNE), Rostock, Germany
iDepartment of Neurology, University Hospital Rostock, Rostock, Germany
jDepartment of Psychiatry and Psychotherapy Rostock, Rostock, Germany
kGerman Center of Neurodegenerative Diseases (DZNE), Magdeburg, Germany
lUniversity Hospital Magdeburg, Magdeburg, Germany
Email address for correspondence: [email protected]
Keywords: cortical thickness, cognitive impairment, ALS-FTD
Background: A continuum between amyotrophic lateral sclerosis (ALS) and fronto-temporal dementia (FTD) is encouraged by pathological, biochemical and genetic characteristics. Overlapping cognitive and behavioural profiles in patients with ALS and FTD have been demonstrated by neuropsychological studies. Imaging studies showed that FTD patients have a distinct pattern of cortical thinning in the temporal lobe which is furthermore associated with progression of clinical deficits (Citation1).
Objectives: Our objective was to compare the cortical thickness of ALS patients with different cognitive stages ranging from normal cognitive performance up to dementia (ALS-FTD). We expected a similar pattern of cortical thinning wherein the ALS patients with no cognitive impairment (ALSnci) should have a stronger thinning in motor areas and ALS-FTD patients in frontal and temporal regions. The patients with cognitive impairment (ALSci) should show a pattern involving motor, frontal and temporal regions.
Methods: An age and gender matched sample of 69 ALS patients and 34 healthy controls was investigated. The ALS group was divided by their cognitive status and according to current consensus criteria (Citation2, Citation3): 33 ALSnic, 28 ALSci, 8 patients were diagnosed as ALS-FTD. The cortical thickness was measured using FreeSurfer. A vertex-wise comparison was performed using a GLM and results are reported according to the Desikan-Killiany atlas.
Results: A vertex-wise comparison between all four groups revealed the following significant results (for all p < 0.05, FDR corrected): ALS-FTD patients had a thinner cortex as the ALSci who had a thinner cortex as the ALSnci patients (ALS-FTD < ALSci < ALSnci) in left and right superior temporal gyrus, superior frontal gyrus, left caudal middle frontal gyrus, lateral orbital frontal gyrus (for ALSnci > ALSci both hemispheres). Besides, a thinner cortex of ALSci patients compared to ALSnci patients was found in the left pars opercularis and rostral middle frontal gyrus. Additionally, a thinner cortex of ALS-FTD patients compared to ALSci patients could be demonstrated in the posterior cingulate cortex. Only the comparison between healthy controls and ALSci patients revealed significant results in terms of significant thinning in the left and right precentral gyrus, pars opercularis, insula, superior and middle temporal gyrus, posterior cingulated cortex and left middle and superior frontal gyrus and medial orbital frontal cortex.
Conclusion: As expected, alterations in the brain of patients with cognitive impairment could be found in the temporal and frontal regions. The decreasing thickness of the fronto-temporal cortex from ALSnci via ALSci to ALS-FTD appears to be an indicator for the continuum between ALS and FTD. Further research is needed to explore cortical thickness as prognostic marker.
References
- Rohrer JD, Warren JD, Modat M . Neurology 2009;72:1562–1569.
- Strong MJ, Grace GM, Freedman M . ALS 2009;10:131-46.
- Phukan J, Elamin M, Bede P . JNNP 2012;83: 102–108.
P218 DIFFERENTIAL INVOLVEMENT OF CORTICOSPINAL TRACT FIBERS IN UMN-PREDOMINANT ALS PATIENTS: A DIFFUSION TENSOR IMAGING AND TRACTOGRAPHY STUDY
V Rajagopalan1
G Yue2
E Pioro1
mCleveland Clinic, OH, USA
nKessler Foundation Research Center, West Orange, NJ, USA
Email address for correspondence: [email protected]
Keywords: tractography, hyperintensity, truncation
Background: Diagnosis of amyotrophic lateral sclerosis (ALS) is dependent on clinical evidence of combined upper motor neuron (UMN) and lower motor neuron degeneration, although either can predominate at disease onset. Some UMN-predominant ALS patients display bilateral hyperintensity of the corticospinal tract (CST) on T2- and proton density (PD)-weighted MRI sequences. Interestingly, other UMN-predominant ALS patients, phenotypically indistinguishable, do not have such CST hyperintensity. The reason for this variability between presence or absence of CST hyperintensity and the essentially identical UMN-predominant clinical features is unclear.
Objectives: Using diffusion tensor imaging (DTI) with diffusion tensor tractography (DTT) to quantitatively assess the intracranial CST of UMN-predominant ALS patients with or without hyperintensity could possibly reveal differences in DTI abnormalities along the CST of both patient subgroups.
Methods: DTI data were obtained at 1.5T in 47 UMN-predominant ALS patients with (n = 21) CST hyperintensity (ALS-CST + ) or without (n = 26) CST hyperintensity (ALS-CST-) and in neurologic controls (n = 12). Fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (AD), and radial diffusivity (RD) were compared between patients and controls along the CST. ALSFRS-R score, disease duration, disease progression rate, and El Escorial score were also obtained in ALS patients.
Results: Significant abnormalities (p < 0.05) were detected in FA, AD, or RD in CST primarily at the internal capsule (IC) level in ALS patients, especially in the ALS-CST + subgroup. AD and RD values were different between ALS-CST + and ALS-CST- patients suggesting distinct pathologies between these subgroups. Furthermore, DTT revealed subcortical truncation of fibers projecting from the precentral gyrus in some ALS-CST + and ALS-CST- patient subgroups but in no controls; fibers projecting to the adjacent postcentral gyrus were spared. This suggests an abnormality of CST fibers arising from primary motor cortex of UMN-predominant ALS patients and not of fibers projecting to/from the primary sensory cortex. Significantly shorter disease duration (p = 0.02) and faster disease progression rate (p = 0.03) were observed in ALS patients with CST fiber truncation than those without, whereas no significant differences were noted for ALSFRS-R and El Escorial scores.
Discussion and conclusions: Abnormalities in DTI metrics at the IC level suggest axonopathy in UMN-predominant ALS patients, which can be quantitatively distinguished between those with CST hyperintensity and those without. Our DTT finding of subcortical fiber truncation in patients arising from the primary motor cortex and not those projecting to/from the primary sensory cortex suggests specificity of this finding, and is in keeping with relative sparing of the sensory system in ALS. Correlation of CST fiber truncation with shorter disease duration and faster disease progression rate in these UMN- predominant ALS patients suggests that DTI/DTT can identify a more severe pathologic process in some patients and be used to distinguish between patient subgroups.
P219 THE RESTING STATE DEFAULT MODE NETWORK (DMN) IS PATHOLOGICALLY HYPERACTIVE IN AMYOTROPHIC LATERAL SCLEROSIS
F Tietz1
V Hartung1
T Prell1
S Penzlin1
B Ilse1
M Bokemeyer2
OW Witte1
J Grosskreutz1
oDepartment of Neurology
pInstitute for Diagnostically and Interventiol Radiology, Jena University Hospital, Jena, Germany,
Email address for correspondence: [email protected]
Keywords: resting state network, default mode network, dual regression
Background: Resting state networks are activated in the human brain when the subjects is at rest (closed eyes, non sleeping and asked to think about nothing). They can be assessed using BOLD effect based functional MRI-techniques. The default mode network (DMN) is active in the medial and lateral part of the prefrontal and temporolateral cortex. The DMN intensifies its activity at rest; its function is thought to be related to daydreaming, comprehending emotions of other people, building autobiographic memory and others functions. In ALS, the DMN has been shown to be activated in larger areas than normal in sensorimotor regions, but premotor and frontal involvement was not decisive.
Objective: To determine the extent of the DMN in ALS patients MRI where T1 spatial normalization was optimized to reduce misrepresentation artefacts.
Methods: We compared 40 Patients with ALS against 40 age matched healthy controls, using the FSL Software package (http://www.fmrib.ox.ac.uk/fsl/index.html). We performed detailed spatial normalization and skull stripping using SPM and an independent component analysis using the FSL Dual Regression Tool (http://www.fmrib.ox.ac.uk/analysis/dualreg) for the comparison of the network activity on group level. All Subjects underwent an assessment of clinical questionnaires (ALSFRS, SF36, EuroQual5D, Frontal Assessment Battery, Mini-Mental Status Examination and Edinburgh Handedness Inventory).
Results: We found a distinctly different behaviour of the DMN of ALS patients when compared to controls. In particular the DMN in ALS significantly increased in both the frontal and temporal regions of the DMN highly suggestive of a loss of intracortical inhibition.
Discussion: These findings may represent compensation for lost underlying subcortical neuronal network capacities or indeed the loss of anatomically long tract connectivity itself when intrinsic intracortical inhibition is postulated to be dependent on anatomically intact long tract connectivity (as opposed to U-fibre connectivity). Previously, a similar analysis (Citation1) was performed, but they found much less hyperactivity in fewer regions of the DMN in patients. Using adequate preprocessing (i.e., optimized skull stripping) greatly improves detection of the extent of DMN hyper- and hypoactivity, and thus the value of rsfMRI to describe ALS related alterations of brain connectivity.
References
- Mohammadi B J.expneurol. 2008.
P220 VOXEL-BASED INTENSITOMETRY OF T1 MRI CAN DIAGNOSE ALS AND PREDICTS CLINICAL DETERIORATION
V Hartung1
C Gaser2
F Tietz1
T Prell1
S Penzlin1
B Ilse1
M Bokemeyer3
OW Witte1
J Grosskreutz1
qDepartment of Neurology
rDepartment of Psychiatry and Psychotherapy
sInstitute for Diagnostic and Interventional Radiology, Jena University Hospital, Jena, Germany
Email address for correspondence: [email protected]
Keywords: MRI, biomarker, diagnostic tool
Background: We introduce a novel approach to advanced voxel-based MRI analysis called Voxel-based intensitometry (VBI). Utilizing known approaches like VBM we developed VBI as a technique to reveal and assess white matter damage in ALS, which is more consistent and more easily accessible than grey matter alteration.
Objectives: To submit T1 MRI data of 30 patients (mean age 63, SD 11; mean ALSFRS-R 37, SD 6; mean disease duration 24 months, SD 18) and 37 matched healthy volunteers to in-house developed preprocessing algorithms that allow for direct comparability of intensity information and assess its feasibility for diagnosing cerebral involvement in motor neuron disease.
Methods: Patients and controls were subjected to T1 MRI and comprehensive clinical assessment, including ALSFRS-R. VBM-like preprocessing algorithms were adapted to normalize all datasets in space and average intensity. Group comparison ANCOVA was used to identify significant ALS-related clusters of altered T1 signal, in which mean intensity was calculated. Regression analysis was conducted to find regions where intensity and ALSFRS-R scores correlated.
Results: Group comparisons revealed areas of significantly different intensity between patients and controls. These regions comprehensively overlapped with the known changes in the white matter of ALS patients’ brains, such as pyramidal tract, corpus callosum, frontal regions and known regions of extra motor involvement. In these clusters, mean intensity allowed to separate patients and controls with 89% specificity at 100% sensitivity. Furthermore regression analysis identified ROIs in which averaged intensity significantly correlated with ALSFRS-R scores (Spearmans rho = 20.89**).
Discussion: In our cohort VBI identified patients vs. controls. Additionally the correlation between intensity and clinical scores suggests predictability of clinical status of the patients on behalf of their MR scans. If proven true, VBI would be the first objective tool to diagnose ALS and verify clinical status in addition to clinical assessment and ALSFRS-R. These results have to be verified in larger cohort multi center trials to prove VBI as a competent biomarker.
P221 A LONGITUDINAL FMRI STUDY OF THE ACTIVATION PATTERN IN MOTOR AND EXTRA-MOTOR AREAS IN ALS
S Vielhaber1,2
C Stoppel1
N Naue2
J Machts2
S Petri3
K Kollewe3
H-J Heinze1,2
R Dengler3
A Schoenfeld1
tUniversity of Magdeburg, Magdeburg, Germany
uGerman Center for Neurodegenerative Diseases, Magdeburg, Germany
vMedical School Hannover, Hannover, Germany
Email address for correspondence: [email protected]
Keywords: fMRI, hippocampus, biomarker
Background: Amyotrophic lateral sclerosis (ALS) is increasingly recognized to be a multisystem disease associated with cognitive dysfunction. Conversely, functional neuroimaging studies revealed activation changes in motor-related areas, but sometimes also within fronto-temporal regions. Typically, activity elicited by a certain task is compared in ALS vs. healthy controls. However, prospective studies investigating the functional changes related to the disease-progression of ALS in a longitudinal within-subject design are lacking.
Objectives: In the present study, we investigated the time course of changes in fMRI activation patterns and their potential contribution to the understanding of ALS pathophysiology.
Methods: Motor- and novelty-processing related brain activity was analyzed in patients with ALS (n = 14) in two sessions separated by a 3-month interval. To assess motor-related activity we employed a Go/NoGo-task, in which task-irrelevant novel stimuli (inside and outside scenes) were additionally presented, offering the possibility to investigate the neural processing of stimulus novelty relative to frequently presented stimuli.
Results: The comparison of the results of the first and second measurement revealed that the activity in the motor cortex decreased during the time course of 3 months. This might reflect the progressive major neural loss in the motor system and functionally connected areas. Interestingly, the activation-decrease in the cerebellum correlated positively with the patients’ ALSFRS-Score and in motor cortex with their MRC-Megascore. Importantly, novelty-processing related activity in the hippocampus was higher in the second compared to the first measurement. This activation-increase showed a negative correlation with the patients’ ALSFRS-Score. This pattern most likely reflects compensatory activity in order to overcome dysfunctions typically observed at the beginning of lesions.
Conclusions: This longitudinal fMRI study captures progression of motor and extra-motor degeneration in ALS. The functional pattern points clearly to an involvement of the hippocampus. Compared to the motor system lesion, the hippocampal dysfunction occurs at a later disease stage which might explain why memory impairment is hardly detected in a clinical setting.
P222 PATIENTS WITH ALS SHOW ALTERED ADIPOSE TISSUE DISTRIBUTION IN WHOLE BODY MRI ANALYSIS
J Kassubek1
H-P Müller1
E Lindauer1
L Dupuis2,3
H Neumann4
AC Ludolph1
wDepartment of Neurology, University of Ulm, Ulm, Germany
xInserm U692, Laboratoire de Signalisations Moléculaires et Neurodégénérescence, Strasbourg, France
yUniversité de Strasbourg, Faculté de Médecine, UMRS692, Strasbourg, France
zInstitute of Neural Information Processing, University of Ulm, Ulm, Germany
Email address for correspondence: [email protected]
Keywords: adipose tissue, MRI, automated analysis
Introduction: Since neurodegenerative diseases affect body weight, assessment of the body fat distribution in the course of the disease might act as a surrogate marker. Body mass index and blood lipids have already been reported to correlate with survival and functional status of ALS patients (Citation1, Citation2). However, these techniques provide no information on the regional distribution of adipose tissue in ALS patients but there are important metabolic differences between fat depots. In this study, whole body MRI was used to analyze fat tissue volume differences between ALS patients and controls.
Methods: Whole body MRI data were acquired on a 1.5 T scanner by acquisition of 6 to 8 consecutive T1-weighted volumes. Sixty-two patients with definite or probable ALS according to revised El Escorial criteria (age 60 ± 12 years, ALSFRS 36.3 ± 7.5, mean disease duration 22 months) and 62 age- and gender-matched controls were examined by a standardized automatic image postprocessing protocol (Citation3), subcutaneous and visceral fat volumes were determined by the Automatic Tissue Labelling Analysis algorithm. Data were diffusion filtered prior to application of the fat determination algorithm ARTIS (Adapted Rendering for Tissue Intensity Segmentation).
Results: The region-of-interest-based approach could be restricted to parts of the body where major fat volume changes were expected in the course of the disease so that MRI-based fat volume analysis was able to differentiate between visceral and subcutaneous fat tissue. Total fat volume of ALS patients was not changed as compared with controls. ALS patients displayed decreased subcutaneous and increased visceral fat content and thus an increased ratio of visceral to subcutaneous fat. Multiple regression analysis showed that gender and ALS-FRS-R were significant predictors of fat volumes.
Conclusion: The potential in determination of subcutaneous and visceral fat volume by MRI based analysis was demonstrated. Fat distribution was altered in ALS patients with increased visceral fat compared to controls. These findings demonstrated that adipose tissue was affected in its topography in ALS and calls for further functional studies on this key metabolic tissue.
References
- Dorst J, Kühnlein P, Hendrich C, Kassubek J, Sperfeld AD, Ludolph AC J Neurol. 2011;258:613–7.
- Dupuis L, Pradat PF, Ludolph AC, Loeffler JP Lancet Neurol. 2011;10:75–82
- Müller HP, Raudies F, Unrath A, Neumann H, Ludolph AC, Kassubek J NMR Biomed 2011;24:17–24.
P223 A LONGITUDINAL STUDY OF DIFFUSION TENSOR IMAGING IN ALS PATIENTS
S Ajroud-Driss
A Mansour
K Herrmann
E Parks
P Casey
J Allen
R Sufit
T Siddique
A Apkarian
Northwestern University Feinberg School of Medicine, Chicago, IL, USA
Email address for correspondence: [email protected]
Keywords: DTI, fractional anisotropy, corticospinal tracts
Background: DTI measures the direction of the flow of water along axon tracts. Fractional anisotropy (FA) derived from DTI serves as an in vivo marker of the integrity of these tracts. Degeneration of these tracts will result in a reduced FA value.
Objective: To assess changes in white matter tracts integrity in ALS patients compared to controls, early in the disease and at 6 months.
Methods: 12 patients with probable and definite ALS with FVC > 70% and ALSFRS > 35 and 12 healthy controls were enrolled. Subjects were scanned at baseline and 6 months later. ALSFRS-R, FVC and manual muscle testing were performed at baseline and at 6 months. Patients and controls underwent two structural MRI scans at baseline and at 6 months that included a high resolution T1 and a DTI scan. We calculated voxel wise fractional anisotropy (FA) Voxel-wise statistical analysis of FA data was carried out using the tract-based spatial statistics. A mean FA skeleton was created for each group. Also, the T1-anatomical brain images were used to calculate cortical gray matter volume.
Results: All ALS patients exhibited a significant progression in their disease over a period of 6 months. When comparing the whole brain skeletal FA between ALS patients and controls we found that the region corresponding to the bilateral corticospinal tracts caudal to the thalamus had significantly lower FA in ALS patients. Over 6 months the anisotropy for this region did not exhibit a significant change, but showed a decreasing trend. This decrease in FA correlated with the decline in FVC. The anisotropy of part of the superior longitudinal fasciculus that was not affected at baseline seemed to decrease in ALS patients but not in controls over 6 months. For the area where ALS patients showed a significant difference from control baseline, there was a difference in radial but not in axial diffusivity suggesting that disrupted myelin tracts may be at the root of the drop in FA noted in the patients. The correlation between peripheral grey matter and mean skeletal FA observed in the healthy controls was disrupted in ALS patients.
Discussion and conclusion: The significant decrease in FA values noted in ALS patients very early in the disease process suggest that DTI is a sensitive tool for detecting early corticospinal tract degeneration and may help establish early diagnosis. Surprisingly longitudinal analysis did not reveal significant change in FA values overtime. Longitudinal decline in FA has been found by some investigators but not by others. DTI could be used as a surrogate marker of UMN degeneration but further studies need to be done before using it as a marker of disease progression. We also found that our patients had difficulties tolerating repeated MRI scan as their disease progressed.
P224 CORTICAL HYPEREXCITABILITY PRECEDES THE CLINICAL ONSET OF SPORADIC ALS
P Menon1
MC Kiernan2,3
S Vucic1,2
aaWestmead Hospital, University of Sydney, Sydney, NSW, Australia
abNeuroscience Research Australia, Sydney, NSW, Australia
acPrince of Wales Hospital, University of NSW,, Sydney, NSW, Australia
Email address for correspondence: [email protected]
Keywords: cortical hyperexcitability, precedes clinical onset, sporadic ALS
Objective: The pathophysiological mechanisms underlying the development of sporadic amyotrophic lateral sclerosis (ALS) remains unresolved. Cortical hyperexcitability was noted to precede the clinical onset of familial ALS in previous studies. This issue remains a matter of debate in sporadic ALS and was addressed in the present study which assessed cortical excitability from a clinically and electro-physiologically normal region in early sporadic ALS patients.
Methods: Cortical excitability studies were undertaken in 12 sporadic, Awaji Criteria probable, ALS patients (6 Male; 6 Female, mean age 56 years) using the threshold tracking technique of Transcranial Magnetic Stimulation (TMS).Motor Evoked Potentials (MEP) were recorded from a clinically normal abductor pollicis brevis (APB) muscle which was also assessed by qualitative and quantitative electromyography (EMG). Results were compared to 66 age matched healthy controls.
Results: Short-interval intracortical inhibition (SICI) was significantly reduced in ALS patients when compared to controls (averaged SICI ALS 0.3 ± 0.8%; controls 10.5 ± 0.7%, P < 0.0001) along with a reduction in the resting motor thresholds (P < 0.05) and cortical silent period (P < 0.05) and an increase in intracortical facilitation (P < 0.05), all indicative of cortical hyperexcitability. Importantly, there was no neuro-physiological evidence of LMN dysfunction, within the target APB muscle. Motor unit architecture represented by the mean motor unit potential (MUP) amplitude, duration and polyphasia was normal.
Conclusion: The present study confirms that cortical hyperexcitability precedes the onset of lower motor neuron dysfunction in sporadic ALS, thereby suggesting a central origin of sporadic ALS with cortical hyperexcitability underlying motor neuron degeneration.
P225 SERIAL MOTOR UNIT NUMBER INDEXING (MUNIX) STUDIES IN THE BICEPS MUSCLE OF ALS PATIENTS
P Barkhaus1
S Nandedkar2
adMedical College of Wisconsin/ Froedtert Hospital, Milwaukee, WI, USA
aeCareFusion, Middleton, WI, USA
Email address for correspondence: [email protected]
Keywords: motor unit, motor unit number estimation, motor unit number index
Background: Motor unit number estimation (MUNE) studies have been used to study loss of motor units (MUs) in patients with motor neuron disease (MND). MUNE is usually performed in distal muscles of hand, e.g., thenar or hypothenar. Proximal muscles are difficult to study due to the multiple nerve stimuli required. A new method called ‘motor unit number index (MUNIX)‘ requires only a single supramaximal stimulation with optimized surface electrode placement, and uses surface EMG interference pattern (SIP) for analysis. Hence it can be used to study large proximal muscles.
Objective: To investigate the change in the number and size of MUs in biceps using the MUNIX method.
Methods: Studies were performed in fourteen patients with MND at 3 month intervals. All patients had 5–7 such studies over a 12–18 month period. The compound muscle action potential (CMAP) was recorded using supramaximal intensity. Several active electrode positions were tested to record the response with highest amplitude. The SIP was recorded using isometric contraction at force levels ranging from slight to maximum. MUNIX was computed using the area and power of the CMAP and SIP signals (Citation1). Motor unit size index (MUSIX) was computed by dividing CMAP amplitude by MUNIX.
Results: Patients were divided into 2 groups based on the disease onset in upper limb (group 1) or lower limb/bulbar (group 2). In Group 1 (5 patients), the MUNIX decreased by 54% while CMAP decreased by 42%. MUSIX increased by 40% over the study period. Three patients had reduced CMAP and MUNIX with increased MUSIX. In group 2 (9 patients) the CMAP and MUNIX decreased by 17% and 18%, respectively, and MUSIX increased by only 2%. One patient had reduced CMAP and MUNIX and with increased MUSIX. Another two patients had borderline abnormalities.
Discussion: This study demonstrates greater MU loss in the biceps muscle of patients whose disease onset was in the upper limb. The reduced MUNIX was partially compensated by reinnervation which gave increased MUSIX with a smaller drop in the CMAP amplitude. In the second patient group, the biceps showed fewer changes in CMAP, MUNIX and MUSIX. Most patients had normal findings in each of 5–7 investigations. This reflects less involvement and progression of the biceps. MUNIX was easy to perform, requiring less than 5 minutes. Recording the CMAP with maximal amplitude was easier when the response from the previous study is available for comparison.
Conclusion: MUNIX can be used to study MU loss and remodeling in MND in proximal muscle such as the biceps.
References
- Nandedkar SD, Barkhaus PE, Stalberg EV Muscle Nerve 2010;42:796–807.
P226 BIOELECTRICAL ACTIVITY IN MUSCLES OF PATIENTS WITH MOTOR NEURON DISEASE: QUANTITATIVE MUP ANALYSIS, REPETITIVE NERVE STIMULATION AND SINGLE FIBER EMG
Ü Kokes
MB Baslo
HA Idrisoglu
Electroneurophysiology Programme in Istanbul University, Institute of Health Science, Department of Neurosience in Istanbul, Istanbul, Turkey
Email address for correspondence: [email protected]
Keywords: Single fiber EMG, repetitive nerve stimulation, quantitative motor unit potential analysis
Background: Electrodiagnostic examination reveals the findings of lower motor neuron (LMN) involvement in ALS which is characterized by ongoing denervation and reinnervation. The rate of disease progression and outcome depend on the dominant process either denervation or reinnervation. As well as the typical routine EMG findings of ALS, the tests which evaluate neuromuscular junction (NMJ) might be pathologic reflecting the crippled transmission caused by immature axonal sprouts and end-plates.
Objective: In this study, bioelectrical activity of motor units in MND was evaluated by Q-MUPA, RNS, SFEMG.
Methods: Thirty two patients (13 F, 19 M; 19–74 years old) who had been diagnosed as MND by clinical and electrophysiological findings were enrolled. Repetitive nerve stimulation of ulnar and accessory nerves as well as Q-MUPA and SFEMG in m. EDC had been performed.
Results and discussion: Mean disease duration was 36.9 ± 60.3 months. In Q-MUPA, MUP durations were prolonged in 28 patients and 7 of them had also high amplitude values. In RNS, 18 patients showed significant decrement (13 patients on m.TRP, 3 patients on m.ADM, 2 patients on both m.ADM and m.TRP). However, 13 of the patients didn’t have a significant decrement. SFEMG revealed high jitter values in 29 patients but 2 patients with normal strength in their EDC muscles revealed normal jitter. In 1 patient, both RNS and SFEMG could not be performed.
Conclusion: These findings demonstrate an ongoing reinnervation in MND by means of long duration - MUPs and faulty transmission at NMJ caused by collateral sprouting. The MUP changes and NMJ abnormality could be detected by Q-MUPA and RNS and/or SFEMG, respectively.
P227 EEG SOURCE ANALYSIS IN AMYOTROPHIC LATERAL SCLEROSIS PATIENTS
E Cuspineda-Bravo1
A Olivares-Torres1
J Iglesias-Fuster2
T Zaldívar-Vaillant1
J Gutiérrez-gil1
A Soto-Lavastida1
J Sánchez-López1
C Pérez-gesen4
D Del Río-Bazán3
C Beltrán1
A Puerta-Armas1
C Machado-Curbelo1
G Lara-Fernández1
afInstitute of Neurology and Neurosurgery, Havana City, Havana, Cuba
agCuban Center for Neuroscience, Havana City, Havana, Cuba
ahHospital Hermanos Ameijeiras, Havana City, Havana, Cuba
aiNational Intitute of Endocrinology, Havana City, Havana, Cuba
Email address for correspondence: [email protected]
Keywords: qEEGt, source analysis, brain electrical tomography
Background: Traditional electroencephalography (EEG) analysis has been considered of scant importance in Amyotrophic Lateral Sclerosis (ALS) because it does not show any particular slowing or specific abnormal activity in this disease, nevertheless the few quantitative EEG(QEEG) studies carried out in ALS has shown evidences of subtle EEG changes in alpha frequencies. No EEG sources analysis studies have been performed in ALS patients.
Objectives: This work focuses on the application of quantitative electric tomography (qEEGT) analysis for determine and localize cortical sources of subtle, not visually detected, changes in EEG activity (for all frequency bands) in ALS patients without dementia.
Methods: conventional basal EEG (19 electrodes, 10/20 system) were recorded from 15 ALS patients in a relaxed wakefulness stage with eyes closed. Bayesian model averaging (BMA) approach was used to estimate electrical sources of EEG recorded activity. High resolution source Z-spectra and 3- dimensional images (Brain Electrical Tomography) of Z values for all the sources at each frequency (0.5–30 Hz) were obtained for all cases. To estimate statistically significant increments and decrements of brain electric activity within the frequency spectra, the t-Student vs. Zero test was performed.
Results: Evident changes were found at the level of EEG cortical sources in all ALS patients independent of time disease evolution. Significant decrement of alpha activity was found at the pre and potcentral gyrus, and in the occipital cortex, cuneus and precuneus areas of both hemispheres. Significant increment of theta activity was found in the precentral and postcentral gyrus, supplemented motor area, superior and middle frontal gyrus of both hemispheres.
Conclusions: This study documents the presence of widespread cortical dysfunction suggesting pathological involvement beyond the motor areas in ALS patients, including those with recent onset of their disease. qEEGt analysis is likely to be a powerful tool for detecting and localize subtle EEG changes in ALS patients.
P228 THE CLINICAL AND NEUROPHYSIOLOGIC STUDY OF UPPER MOTOR NEURON-DOMINANT AMYOTROPHIC LATERAL SCLEROSIS
Y Xu
L Tang
N Zhang
D Fan
Peking University Third Hospital, Beijing, China
Email address for correspondence: [email protected]
Keywords: PLS, upper motor neuron-dominant ALS, characteristics
Objective: To investigate the clinical and electrophysiological characteristics of upper motor neuron-dominant amyotrophic lateral sclerosis (UMN-D ALS).
Methods: The clinical and electrophysiological characteristics were analyzed retrospectively in 76 patients with UMN-D ALS and 19 patients with primary lateral sclerosis (PLS).
Results: There were 84 UMN-D ALS patients after 8 PLS patients shifted to this group. In UMN-D ALS patients, the females increased in those with ages more than 40 (male: female = 1: 1.37), 32 patients (38.1%) were bulbar onset. UMN-D ALS patients acquired EMG evidence of denervation in a median of 30 months after symptom onset, and clinical lower motor neuron (LMN) signs 6 months later, 77 patients (91.6%) developed LMN sign by 4 years from symptom onset. ALSFRS-R changed from 40 ± 3 to 32 ± 4 in UMN-D ALS patients in 4 years (t = 1.83, P < 0.05). The amplitude of motor unit action potential (MUAP) of the first interosseus dorsalis was higher and the duration of MUAP was longer in UMN-D ALS patients than in PLS patients ((582.5 ± 30.7) μv vs (353.5 ± 21.5) μv; (t = 1.87, P < 0.05));((19.8 ± 2.3) ms vs (9.6 ± 1.3) ms; (t = 1.85, P < 0.05)).
Conclusions: UMN-D ALS patients characterized by more females, more cases with bulbar onset and faster progression than PLS patients, and electromyographic evidence of focal denervation.
P229 ALS ENTERS THE WORLD OF BIG DATA: INITIAL DESCRIPTION OF AND RESULTS FROM THE PRO-ACT PLATFORM
M Leitner1
A Sherman2
D Schoenfeld2
N Atassi2
JD Berry2
N Zach1
E Sinani2
J Walker2
I Katsovskiy2
M Cudkowicz2
ajPrize4Life, Cambridge, MA, USA
akMass General Hospital, Boston, MA, USA
Email address for correspondence: [email protected]
Keywords: research resource, dataset, clinical data
Background: Large datasets are critical for identifying statistically significant and biologically relevant observations, particularly for diseases resulting from the intricate interplay of genetic and environmental factors. The Pooled Resource Open-access ALS Clinical Trials (PRO-ACT) platform provides an unprecedented opportunity to increase our understanding of the ALS patient population and the natural history of the disease.
Objective: To build, analyze, and make publicly available the world's largest merged dataset of patient clinical data from completed Phase II/III ALS clinical trials.
Design and methods: We obtained 6700+patient records from 11 completed industry-sponsored trials and 1000 records from 7 government/non-profit sponsored trials. These data include the placebo arms from all trials and the drug arms from the majority of trials, including the riluzole trials. Utilizing empirical and theoretical knowledge from the literature, common data elements developed by NINDS and NEALS, and with input from clinical investigators, we have developed an ALS specific common data structure. We have used this common data structure to enable the merging of disparate datasets for the first time into a unified searchable dataset.
Results: The dataset contains at minimum an estimated 6 million datapoints, including never before described data, collected over a time period of 6–22 months (average 13 months). We have begun statistically analyzing this dataset so as to describe its contents and increase its utility for the research community. Baseline characteristics, survival distribution, and identified prognostic factors will be described. The rate of functional decline was compared between the subset of the industry-sponsored trials and the government/foundation-sponsored trials that assessed ALSFRS to ensure that the respective patient populations were similar. The full dataset will be made freely accessible to the research community for analysis and download (anticipated release Dec 2012 to coincide with ALS/MND), and a sample of the data will be released this summer for the ALS Prediction Prize challenge promoting the development of quantitative methods to predict future ALS progression.
Discussion: The PRO-ACT platform is the result of a unique collaboration among industry, academic, and foundation partners. Creation of this platform will enable a wide variety of analyses that were previously not possible. Use of this large merged ALS clinical dataset by the research community will help us better understand the natural history of the disease, further efforts to successfully stratify ALS patients, and provide insights into the design and interpretation of future clinical trials. Academic institutions and companies are encouraged to both contribute and analyze data for new insights into ALS. Pooling and analyzing clinical trial data is currently being explored for a wide variety of related diseases including Alzheimer's disease, Parkinson's disease, Multiple Sclerosis, and Frontotemporal Dementia, and the PRO-ACT platform will serve as a model for these future efforts.
P230 GRAY MATTER DEMYELINATION IS UNEXPECTEDLY COMMON IN NEURODEGENERATION: AMYOTROPHIC LATERAL SCLEROSIS
Y Li1
J Zhang1
S Kang1
D Bergles1
J Rothstein1,2
alJHU School of Medicine, Baltimore, MD, USA
amJHU Neuroscience and the Brain Science Institute, Baltimore, MD, USA
Email address for correspondence: [email protected]">[email protected]">[email protected]
Keywords: NG2 glia, oligodendrocyte, myelination
Background: ALS is a non-cell autonomous disease as astroglia and microglia play a role in disease progression in rodent models. However, whether oligodendrocyte lineage is involved in ALS pathogenesis is not clear. Our recent study has shown that NG2 glia, also known as oligodendrocyte progenitor, proliferate and undergo dramatic changes in the lumbar spinal cord gray matter in G93A SOD1 mice at endstage. This suggests that oligodendroglia might degenerate during disease progression, which could be causing NG2 glial proliferation. More importantly, in ALS patients, although it is well known corticospinal tract demyelination occurs, it is largely unknown whether there are oligodendrocyte lineage changes in motor cortex and spinal cord gray matter where motor neurons degenerate.
Objectives: Given that NG2 glia are restricted to oligodendrocyte differentiation in ALS mouse lumbar spinal cord gray matter, we asked whether oligodendrocyte degenerate as disease progresses as measured by myelination in mouse. In addition, we investigated whether oligodendrocyte lineage undergoes similar changes in the CNS gray matter of ALS patients.
Methods: Immunohistochemistry (IHC) was used to determine NG2 glia cell morphology. IHC staining, luxol fast blue myelin staining and erichrome cyanine myelin staining were performed to evaluate myelin expression. Western Blotting was used to quantitatively evaluate myelin protein expression. MRI in vivo imaging was carried out to determine mouse lumbar spinal cord myelin signals.
Results: In ALS mouse lumbar spinal cord, loss of myelin basic protein expression/demyelination was seen as early as disease onset. Repeated longitudinal mouse MRI studies also suggested that gray matter demyelination occurred in vivo at the onset of clinical disease. Further myelin staining showed clear gray matter demyelination and distorted myelin structures in the lumbar spinal cord gray matter. In ALS patient motor cortex gray matter, NG2 glia showed increased immunoreactivity compared to those without neurological diseases. In addition, patchy demyelination in the gray matter of motor cortex and spinal cords was reliably and repeatedly observed, with preservation of neurons in the some regions of demyelination . Furthermore, we found that in ALS patients the decrease in myelin protein expression was correlated to oligodendrocyte progenitor marker, PDGFαR, expression, which is consistent with our animal studies.
Discussion: Both animal and human studies unexpectedly showed gray matter oligodendrocyte degeneration as measured by different myelin stains and in vivo MRI imaging in ALS. Since oligodendrocytes not only facilitate saltatory conduction of action potential, but also support neuronal functions such as lactate metabolism, through oligodendroglial specific MCT1, the injury to oligodendroglia in ALS may have pathogenic consequences.
Conclusions: Oligdendrocytes degenerate in the gray matter in ALS.
P231 SERUM LEVELS OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN AMYOTROPHIC LATERAL SCLEROSIS. RELATION TO ITS INCREASED EXPRESSION OF SKIN
H Ishikawa
S Ono
Teikyo University Chiba Medical Center, Ichihara, Chiba, Japan
Email address for correspondence: [email protected]
Keywords: skin, serum, vascular endothelial growth factor
Background: Vascular endothelial growth factor (VEGF), first discovered as a tumor-secreted protein that promotes vascular permeability, is an important angiogenesis factor. VEGF is expressed in many tissues and is rapidly upregulated during hypoxia, which occurs during acute or chronic vascular disease, pulmonary disease, or cancer. The precise mechanisms governing this upregulation remain unknown, but oxygen deprivation is a key regulator of VEGF gene expression with cells rapidly accumulating hypoxia-inducible transcription factors (HIFs) in response to decreased oxygen tension. A central role of VEGF in the pathophysiology of motor neuron disease has been raised by studies in mice where the hypoxia-response element of the VEGF promoter has been deleted. The VEGF165 isoform stimulates normal and abnormal vessel growth and promotes survival of motor neurons during hypoxia, VEGF gene expression is stimulated mainly through binding of HIF to a defined hypoxia-response element in the VEGF promoter.
Objectives: We measured serum levels of VEGF in ALS patients and controls, and studied the relationship between serum levels of VEGF and the density of VEGF immunoreactivity of skin in ALS patients.
Methods: Skin biopsy samples were taken from the left upper arm of 20 patients with ALS (mean age 62.1 years) and from 20 controls with other neurodegenerative diseases matched for sex and age (mean age 59.7 years). Routine formalin-fixed paraffin-embedded 6 μm sections were immunostained according to standard techniques. The sections were incubated with anti-VEGF antibody. The sections were stained by ABC kit. The immunoreactivity was quantified with an image-analysis system. Blood samples were collected from the same 20 ALS patients and the same 20 controls. Samples were analyzed using commercially available ELISA kits for VEGF.
Results: Serum VEGF levels were significantly higher (p < 0.02) in ALS patients (365.4 ± 140ng/l) than in controls (203.6 ± 125.9ng/l). There was a moderate positive relationship (r = 0.51) between duration of the disease and VEGF levels in serum from ALS patients. The immunoreactivity of VEGF was strongly positive in the epidermis and in some blood vessels and glands of the reticular dermis in all ALS patients. Its optical density in ALS patients (6.22 ± 2.91) was significantly higher (p < 0.001) than in controls (1.65 ± 0.61). Furthermore, there was a significant positive relationship (r = 0.84, p < 0.001) between the immunoreactivity and duration of illness in ALS patients. There was a significant positive relationship (p < 0.02) between serum VEGF concentrations and the optical density of VEGF in ALS patients.
Conclusions: The increased amount of serum VEGF may reflect in part the increased VEGF immunoreactivity of skin in ALS and may be involved in the pathogenesis of ALS.
P232 A COMPARATIVE STUDY OF BIOMARKERS FOR DYSPHAGIA IN AMYOTROPHIC LATERAL SCLEROSIS
E Bearce
A Pinnegar
M Collins
M Sell
M Konczal
L Al-Khashti
V Gaiser
JR Coates
T Lever
University of Missouri, Columbia, MO, USA
Email address for correspondence: [email protected]
Keywords: dysphagia, swallowing, animal models
Background: Swallowing impairment (dysphagia) is common in amyotrophic lateral sclerosis (ALS), resulting in malnutrition, dehydration, and pulmonary aspiration. These symptoms are associated with a poor quality of life and contribute to death. No effective treatments currently exist for dysphagia in ALS.
Objectives: Our lab is dedicated to studying animal models of ALS to hasten the identification of novel treatments for dysphagia. Previous work in our lab showed that, like people with ALS, the SOD1-G93A transgenic mouse develops symptoms of dysphagia. Dogs with degenerative myelopathy (DM) exhibit a spontaneous SOD1 mutation that results in ALS-like symptoms of the limbs similar to humans and transgenic mice. Unspecified swallowing impairment was recently reported in end-stage dogs with DM. Our goal is to identify common biomarkers of dysphagia in ALS across species (mouse, dog, and human).
Methods: We have developed 4 dysphagia assays that have the potential for universal application in mammals: 1) Video analysis (VA) of oral ingestive behaviors, 2) Fluoroscopic (x-ray) analysis (FA) of swallowing, 3) Laryngeal brainstem response (LBR), and 4) Histological analysis (HA) of the brainstem nuclei involved in swallowing (trigeminal, facial, ambiguus, hypoglossal, dorsal motor nucleus of the vagus, and nucleus tractus solitarius). Using VA methodology, we investigated the lick rate of SOD1-G93A mice (n = 14) and nontransgenic littermates (n = 12) of either sex at 3, 5, and 7 weeks of age. For HA, brainstems of formalin-fixed end-stage DM (n = 4) and control (n = 4) dogs, and ALS (n = 2) and control (n = 2) humans were paraffin processed and stained using hematoxylin & eosin to identify brainstem nuclei and general histopathology by light microscopy. Researchers were blinded to genotype for all assays.
Results: At all time points, SOD1-G93A mice demonstrated a significantly slower lick rate compared to controls (p < 0.05). We are now establishing biomarkers of dysphagia in this strain using FA, LBR, and HA methods. In all canine DM and human ALS samples, subtle evidence of neurodegeneration was identified within each of the brainstem nuclei involved in swallowing. This finding is markedly different from our previous report of prominent vacuolization in the brainstem nuclei of SOD1-G93A mice.
Discussion and conclusions: Early-onset impairment of lick rate in SOD1-G93A mice suggests this strain may be a model of bulbar rather than spinal onset ALS, making it uniquely suitable for translational research to humans with bulbar onset ALS (˜30% of ALS cases). However, our histological findings suggest that neurodegeneration in canine DM may more closely recapitulate human ALS. We are currently testing this hypothesis using VA and FA methods to establish a clinicopathological correlation of dysphagia in canine DM for comparison with SOD1-G93A mice. Identification of common biomarkers of dysphagia in ALS across species is a critical step toward the identification of novel therapeutics suitable for pre-clinical trials.
P233 NOVEL MARKERS OF MITOCHONDRIAL DYNAMICS IN AXONS ARE DISRUPTED IN MUTANT SOD1 TRANSGENIC MICE
B Gentil
H Durham
MNI McGill University, Québec, Canada
Email address for correspondence: [email protected]
Keywords: mitochondrial fusion, axon, SOD1
Background: Distal axonopathy with loss of integrity of the neuromuscular junction occurs early in the pathogenesis of a familial form of ALS caused by mutations in SOD1 (ALS1). A strong and early mitochondrial phenotype also is observed in culture and transgenic mouse models, including altered morphology, fission/fusion, transport and bioenergetics. The dramatic rounding of mitochondria observed in cultured motor neurons expressing the ALS1-causing mutant, SOD1G93A, correlated with inhibition of mitochondrial fusion. This led us to investigate the role of post-translational modifications of profusion proteins and how they are affected by mutant SOD1.
Objectives: To assess differences in mitochondrial dynamics in neuronal cell bodies and axons and the impact of mutant SOD1 on these processes.
Methods: Explant cultures of murine dorsal root ganglia (DRG) were used for this study because of the ability to analyze cell bodies and axons separately by biochemical techniques. Mitochondrial fusion was assayed by co-expressing photoactivable EGFP targeted to the mitochondrial matrix. Profusion proteins were examined by Western analysis of extracts prepared from cell body and axonal compartments of the cultures and from brain, spinal cord and sciatic nerves of wildtype mice and those carrying SOD1WT or SOD1G93A transgenes.
Results: In cultured neurons, the rate of mitochondrial fusion was higher in the cell body than in the axon, suggesting a local regulation of profusion proteins which could be affected in ALS1. Full length Mfn2 was detected on Western blots as a 83 kDa band. However, in extracts of DRG axons and of mouse sciatic nerve and brainstem, this band was diminished and two smaller bands were labeled by antibody to Mfn2, migrating at 70 kDa and 13 kDa. This suggested the smaller forms (sMfn2) arose from proteolytic cleavage of full length Mfn2. Preferential cleavage in axons was indicated by relative absence of the sMfn2 in DRG cell bodies and in regions of nervous tissue dominated by cell bodies and dendrites (i.e., lumbar spinal cord, cortex or cerebellum). In silico analysis identified a presenilin-like consensus sequence in the predicted cleavage region. Interestingly, these sMfn2 were absent from sciatic nerve of SOD1G93A transgenic mice at 120 days of age, just prior to appearance of overt symptoms.
Discussion: The data provide strong evidence of specialized processes controlling mitochondrial dynamics in neuronal cell bodies and axons. Cleavage of the profusion protein, Mfn2, in axons correlated with reduced rate of mitochondrial fusion. Absence of Mfn2 cleavage in sciatic nerve of fALS1 transgenic mice implicates abnormal protease activity in the axonopathy that is an early feature of the disease. We propose that sMfn2 could serve as a biomarker for evaluation of therapeutic efficacy.