Live Cell Imaging of ATP Levels Reveals Metabolic Compartmentalization within Motoneurons and Early Metabolic Changes in FUS ALS Motoneurons

Abstract

Motoneurons are one of the most energy-demanding cell types and a primary target in Amyotrophic lateral sclerosis (ALS), a debilitating and lethal neurodegenerative disorder without currently available effective treatments. Disruption of mitochondrial ultrastructure, transport, and metabolism is a commonly reported phenotype in ALS models and can critically affect survival and the proper function of motor neurons. However, how changes in metabolic rates contribute to ALS progression is not fully understood yet. Here, we utilize hiPCS-derived motoneuron cultures and live imaging quantitative techniques to evaluate metabolic rates in fused in sarcoma (FUS)-ALS model cells. We show that differentiation and maturation of motoneurons are accompanied by an overall upregulation of mitochondrial components and a significant increase in metabolic rates that correspond to their high energy-demanding state. Detailed compartment-specific live measurements using a fluorescent ATP sensor and FLIM imaging show significantly lower levels of ATP in the somas of cells carrying FUS-ALS mutations. These changes lead to the increased vulnerability of diseased motoneurons to further metabolic challenges with mitochondrial inhibitors and could be due to the disruption of mitochondrial inner membrane integrity and an increase in its proton leakage. Furthermore, our measurements demonstrate heterogeneity between axonal and somatic compartments, with lower relative levels of ATP in axons. Our observations strongly support the hypothesis that mutated FUS impacts the metabolic states of motoneurons and makes them more susceptible to further neurodegenerative mechanisms.

Glycolic acid and D-lactate—putative products of DJ-1—restore neurodegeneration in FUS - and SOD1-ALS

“…In addition, it has been recently shown that increased demand for NAD + relative to ATP induces aerobic glycolysis (Luengo et al, 2021). We recently showed that a boosted metabolic turnover of the glycolytic pathway improved the viability of FUS-ALS MN, whereas blocking glycolysis reduced their viability (Zimyanin et al, 2023). These data altogether suggest that the beneficial treatment effect of GA and DL in FUS-and SOD1-ALS MNs might be because of a metabolic rescue by restoring the NAD(P)H reservoir.…”

“…DJ-1 is integral for maintaining mitochondrial potential, Ca 2+ homeostasis and ATP production. Of note, loss of DJ-1 did not affect mitochondrial respiration but increased ROS production and mitochondrial permeability transition pore opening (Giaime et al, 2012), phenotypes which we recently identified also in FUS-ALS MNs (Zimyanin et al, 2023). GA can support the mitochondrial membrane potential and neuronal survival (Toyoda et al, 2014), improve mitochondrial energy production, thereby increasing the levels of NAD(P)H (Bour et al, 2021 Preprint) and can also reduce oxidative stress via a glutathione-mediated pathway (Diez et al, 2021).…”

Increased Vulnerability to Ferroptosis in FUS-ALS

“…FUS-ALS pathophysiology was associated with significant mitochondrial dysfunction. Specifically, axonal mitochondrial transport was shown to be impaired, as well as mitochondrial depolarization, which was obvious when observed together with the impacted metabolic state of FUS-mutated neurons [33,34]. We first investigated whether this mitochondrial phenotype is also seen in HeLa cells engineered to carry either wild-type FUS-eGFP or P525L FUS-eGFP.…”