Mutations in superoxide dismutase (SOD1) are causative for inherited amyotrophic lateral sclerosis. A proportion of SOD1 mutant protein is misfolded onto the cytoplasmic face of mitochondria in one or more spinal cord cell types. By construction of mice in which mitochondrially targeted enhanced green fluorescent protein is selectively expressed in motor neurons, we demonstrate that axonal mitochondria of motor neurons are primary in vivo targets for misfolded SOD1. Mutant SOD1 alters axonal mitochondrial morphology and distribution, with dismutase active SOD1 causing mitochondrial clustering at the proximal side of Schmidt-Lanterman incisures within motor axons and dismutase inactive SOD1 producing aberrantly elongated axonal mitochondria beginning pre-symptomatically and increasing in severity as disease progresses. Somal mitochondria are altered by mutant SOD1, with loss of the characteristic cylindrical, networked morphology and its replacement by a less elongated, more spherical shape. These data indicate that mutant SOD1 binding to mitochondria disrupts normal mitochondrial distribution and size homeostasis as early pathogenic features of SOD1 mutant-mediated ALS.

TDP-43 has been found in inclusion bodies of multiple neurological disorders, including amyotrophic lateral sclerosis, frontotemporal dementia, Parkinson's disease and Alzheimer's disease. Mutations in the TDP-43 encoding gene, TARDBP, have been subsequently reported in sporadic and familial ALS patients. In order to investigate the pathogenic nature of these mutants, the effects of three consistently reported TARDBP mutations (A315T, G348C and A382T) were tested in cell lines, primary cultured motor neurons and living zebrafish embryos. Each of the three mutants and wild-type (WT) human TDP-43 localized to nuclei when expressed in COS1 and Neuro2A cells by transient transfection. However, when expressed in motor neurons from dissociated spinal cord cultures these mutant TARDBP alleles, but less so for WT TARDBP, were neurotoxic, concomitant with perinuclear localization and aggregation of TDP-43. Finally, overexpression of mutant, but less so of WT, human TARDBP caused a motor phenotype in zebrafish (Danio rerio) embryos consisting of shorter motor neuronal axons, premature and excessive branching as well as swimming deficits. Interestingly, knock-down of zebrafisfh tardbp led to a similar phenotype, which was rescued by co-expressing WT but not mutant human TARDBP. Together these approaches showed that TARDBP mutations cause motor neuron defects and toxicity, suggesting that both a toxic gain of function as well as a novel loss of function may be involved in the molecular mechanism by which mutant TDP-43 contributes to disease pathogenesis.

Tay-Sachs disease is a severe lysosomal disorder caused by mutations in the HexA gene coding for the α-subunit of lysosomal β-hexosaminidase A, which converts G(M2) to G(M3) ganglioside. Hexa(-/-) mice, depleted of β-hexosaminidase A, remain asymptomatic to 1 year of age, because they catabolise G(M2) ganglioside via a lysosomal sialidase into glycolipid G(A2), which is further processed by β-hexosaminidase B to lactosyl-ceramide, thereby bypassing the β-hexosaminidase A defect. Since this bypass is not effective in humans, infantile Tay-Sachs disease is fatal in the first years of life. Previously, we identified a novel ganglioside metabolizing sialidase, Neu4, abundantly expressed in mouse brain neurons. Now we demonstrate that mice with targeted disruption of both Neu4 and Hexa genes (Neu4(-/-);Hexa(-/-)) show epileptic seizures with 40% penetrance correlating with polyspike discharges on the cortical electrodes of the electroencephalogram. Single knockout Hexa(-/-) or Neu4(-/-) siblings do not show such symptoms. Further, double-knockout but not single-knockout mice have multiple degenerating neurons in the cortex and hippocampus and multiple layers of cortical neurons accumulating G(M2) ganglioside. Together, our data suggest that the Neu4 block exacerbates the disease in Hexa(-/-) mice, indicating that Neu4 is a modifier gene in the mouse model of Tay-Sachs disease, reducing the disease severity through the metabolic bypass. However, while disease severity in the double mutant is increased, it is not profound suggesting that Neu4 is not the only sialidase contributing to the metabolic bypass in Hexa(-/-) mice.

The antiproliferative action of nitric oxide (NO) has been well established and increased production was reported in the infarcted rat heart. Concomitantly, increased DNA synthesis and hyperplasia of cardiac myocytes were documented in the hypertrophied myocardium. Despite these observations, the effect of NO on DNA synthesis in hypertrophied cardiac myocytes remains unexamined. Hypertrophy of the non-infarcted left ventricle (NILV) in 1-week post-MI rats was characterized by the increased prepro-ANP and reduction of α-myosin heavy chain protein expression. Inducible NO synthase was expressed in the NILV and associated with a concomitant attenuation of MnSuperoxide dismutase protein content. The latter data suggest that an antiproliferative action of NO in the hypertrophied NILV may proceed via either a cyclic GMP-dependent pathway and/or facilitated by a peroxynitrite-dependent mechanism. In neonatal rat ventricular myocytes (NNVM), the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) promoted a dose-dependent attenuation of DNA synthesis via a cyclic GMP-independent pathway. The permeable superoxide dismutase mimetic and peroxynitrite scavenger MnTBAP abrogated SNAP-dependent attenuation of DNA synthesis in NNVM. MnTBAP failed to inhibit SNAP-mediated recruitment of extracellular signal regulated kinase 1/2 (ERK1/2) but partially attenuated p38 phosphorylation. In hypertrophied NNVM induced by norepinephrine, SNAP-mediated peroxynitrite-dependent inhibition of DNA synthesis, ERK1/2 and p38 phosphorylation were significantly attenuated. Collectively, these data suggest that despite a favourable environment for NO and subsequent peroxynitrite generation in the NILV, hypertrophied cardiac myocytes may be partially refractory to their biological actions.