**6. Mitochondrial pathology in ALS**

Mitochondria constitute approximately 25% of the cytoplasmic volume in most eukaryotic cells and produce cellular energy in the form of ATP via electron transport and oxidative phosphorylation. During electron transfer in the inner membrane of the organelle, electrons spontaneously leak from the electron transport chain and react with available O2 to produce superoxide, which makes mitochondria the major cellular sources of ROS. Mitochondria have been recognized as target organelles for the regulation and execution of cell death under pathological conditions [172-173]. There are many mitochondria in the motor neurons because of the high rate of metabolic demand therein, which implies that motor neurons are susceptible to functional or morphological alteration in mitochondria. Mtochondrial abnormality may play a crucial role in the pathologic mechanism of motor neuron diseases and of ALS. Studies with ALS patients and animal ALS models have been performed to examine both the mor‐ phologic and functional abnormalities of the mitochondria [174]. Morphological abnormality in the organelle that includes a fragmented network, swelling, and increased cristae has been observed in the soma and proximal axons of ventral motor neurons of sporadic ALS (sALS) patients [39]. In ALS patients, a reduction in complex IV of the electron transport chain activity was evident and has been associated with mutations in mitochondrial DNA [175-176]. Although SOD1 is mainly localized in cytosols, it is also resilient in other subcellular com‐ partments such as the mitochondria [45,177-178] and even the endoplasmic reticulum [182]. The aggregates of mutant SOD1 were shown within the mitochondria of the spinal cord of SOD1G93A mice before the onset of symptoms [46-47] and were implicated in increased oxidative damage, decreased respiratory activity of mitochondria [48], and the appearance of mitochondrial swelling and vacuolization [47]. Dissociated cytochrome c from the interaction of mitochondria with mutant SOD1 activates apoptosis [44]. Mitochondria function as reservoirs of intracellular Ca2+, as ER. Once overloaded in cytosol, the accumulated Ca2+ in the mitochondria prepares the organelle to undergo permeability transition, and then swells and ruptures in their outermembrane, which in turn produces free radicals from them and oxidizes their lipids and DNA [179-180]. Ca2+-induced mitochondrial damage can also result in mitochondrial release of cytotoxic substances such as cytochrome c [181] and can affect caspase cascade. The homeostasis at the intracellular Ca2+ level was also disturbed in motor neurons of SOD1G93A mice [182]. Moreover, increased Ca2+ uptake into the mitochondria of motor neurons easily occurred after exposure to the glutamate agonist AMPA or kinate, and triggered increased ROS generation [183]. ALS-linked SOD1 has been shown to slow down fast axonal transport of mitochondria. The axonal mitochondria transport was primarily reduced in the anterograde direction, which suggests that the energy supply in the presynaptic terminals of the motor endplates is compromised [184]. Multiple functions of the mitochondria over cellular injury and the apearance of mitochondrial dysfunction in the presymptomatic stage may contribute to various routes of neuronal death in ALS. More recently, in mice that expressed human TDP-43 only in neurons that included motor neurons, massive accumulation of mitochondria in TDP-43-negative cytoplasmic inclusions in the motor neurons were reported and the lack of mitochondria in the motor axon terminal was observed [185]. In addition, the transgenic mice that overexpressed human TDP-43 driven by the mouse prion promoter demonstrated motor deficits, early mortality, and mitochondrial aggregation [186]. These results imply that TDP-43 is indirectly involved in mitochondrial dysfunction in neurodege‐ nerative diseases such as ALS.

proteasome system and the autophay-lysosome pathway [190-191]. Enhancing the latter with physiological characteristics prevents motor neuron dysfunction in vivo [192-193]. Defects in the autophagy pathway have a principal disease-causing role in human pathologies that include neurodegeneration [189,194]. Studies of the spinal motor neurons of ALS patients [195] and ALS transgenic mice [196] have delineated the abnormality in autophagy, which is probably correlated with the pathogenesis of the disease [192-193,197]. A growing number of studies support the concept that autophagy makes diseased motor neurons healthy by clearing the aggregated mutant SOD1, which was accomplished by inducing autophagy, as illustrated by the increased number of autophagosomes and the higher level of autophagy markers such as Beclin-1, ATG5-ATG12 complex, and LC3-II [192-193]. It is also possible, however, that blunt autophagy in neurodegenerative conditions was accompanied by the abnormal accumulation of autophagosomes and excessive markers, which might have killed the neurons [197-198] and which indicates the compensatory role of autophagy in inherited ALS. Thus, the detailed molecular mechanism of the development of autophagy-mediated diseases must be explained.

Multiple Routes of Motor Neuron Degeneration in ALS

http://dx.doi.org/10.5772/56625

47

The parallel pathway of oxidative stress and Fas-mediated apoptosis in motor neuron death in SOD1G93A mice was previously focused on [96]. This study provided the first evidence that combination therapy that targets oxidative stress and apoptosis together also delays the onset and progression of motor dysfunction and extends the survival time of ALS transgenic mice. Evidence was accumulated that shows that oxidative stress and apoptotic insults cause neuronal death through distinctive pathways and with unique morphological changes. The neurotrophins' nerve growth factor, the brain-derived neurotrophic factor (BDNF), neurotro‐ phin 3 (NT-3), and NT-4/5, and the insulin-like growth factors IGF-I and IGF-II, promote neuronal survival by preventing programmed cell death or apoptosis, but they significantly enhance necrotic degeneration of neurons exposed to oxidative stress or deprived of oxygen and glucose [199-200]. Neurotrophins can induce oxidative stress by upregulating NADPH oxidase, which leads to neuronal cell necrosis [201]. Surprisingly, the insulin-like growth factor 1 (IGF-1) prevented neuronal cell apoptosis and protected spinal motor neurons in ALS mice [199,202], but markedly potentiated neuronal cell necrosis induced by hydroxyl radicals or glutathione depletion [203]. Given that oxidative stress and apoptosis play a central role in motor neuron degeneration and can contribute to neuronal death through distinctive routes in ALS, it was hypothesized that a therapeutic approach that targets both oxidative stress and apoptosis would have additive effects on neuronal survival and the motor function. To pharmacologically prevent oxidative stress and apoptosis, Neu2000, a novel anti-oxidant, and

, a well-known anti-apoptosis agent, were used. The former, a chemical derivative of aspirin and sulfasalazine, was developed to protect neurons from oxidative stress with greater potency and safety, and has been shown to be a potent and secure anti-oxidant in vitro and in animal

has been shown to prevent apoptosis through mecha‐

**8. Therapeutic strategy for ALS**

models of hypoxic ischemia [204]. Li+

Li+

**8.1. Separate routes of motor neuron degeneration in ALS**

### **7. Autophagy in ALS**

Autophagy is a degradative mechanism that is involved in the recycling and turnover of longlife proteins and organelles [187]. Autophagy is basically induced by lack of nutrients and energy or by various toxicants. Although its primary role is adaptation to scarcity, this degradative process is also critical for the normal turnover of cytoplasmic contents that include neurons. Genetic ablation of autophagy-related genes provokes neurodeneration even with lack of disease-like mutant proteins [188]. Recent studies verified the importance of the autophagy pathway in various pathological conditions that include neurodegenerative diseases [189]. Interestingly, the catabolic process is both beneficial and detrimental to cells, depending on its context and specific stimuli. The lethality of mutated SOD1 is the result of abnormal protein aggregates, which impair the degradation machinery such as the ubiqutinproteasome system and the autophay-lysosome pathway [190-191]. Enhancing the latter with physiological characteristics prevents motor neuron dysfunction in vivo [192-193]. Defects in the autophagy pathway have a principal disease-causing role in human pathologies that include neurodegeneration [189,194]. Studies of the spinal motor neurons of ALS patients [195] and ALS transgenic mice [196] have delineated the abnormality in autophagy, which is probably correlated with the pathogenesis of the disease [192-193,197]. A growing number of studies support the concept that autophagy makes diseased motor neurons healthy by clearing the aggregated mutant SOD1, which was accomplished by inducing autophagy, as illustrated by the increased number of autophagosomes and the higher level of autophagy markers such as Beclin-1, ATG5-ATG12 complex, and LC3-II [192-193]. It is also possible, however, that blunt autophagy in neurodegenerative conditions was accompanied by the abnormal accumulation of autophagosomes and excessive markers, which might have killed the neurons [197-198] and which indicates the compensatory role of autophagy in inherited ALS. Thus, the detailed molecular mechanism of the development of autophagy-mediated diseases must be explained.
