An in vitro compartmental system underlines the contribution of mitochondrial immobility to the ATP supply in the NMJ

Abstract

The neuromuscular junction (NMJ) is the largest, most complex synapse in the human body. Motor neuron (MN) diseases, such as amyotrophic lateral sclerosis (ALS), specifically target MNs and the NMJs. However, little is known about the reasons for MN selective neuronal and synaptic vulnerability in MN diseases. Here, utilizing a compartmental microfluidic in vitro co-culture system, we provide a possible explanation why the NMJ, other than its unusual dimensions, differs from other synapses. Using live imaging techniques, we discovered that cultured MNs display higher axonal and synaptic mitochondrial immobility compared with sympathetic neurons (SNs), leading to a profound enrichment of mitochondria only in the MN NMJ. Furthermore, employing a synaptic ATP sensor, we show that mitochondrial respiration is the key contributor to ATP production in MN NMJs but not in SN synapses. Taken together, our data suggest that mitochondrial localization underlies the unique and specific qualities of MN NMJs. Our findings shed light on the role of mitochondria in MN and NMJ maintenance, and possibly indicate how mitochondria may serve as a source for selective MN vulnerability in neurodegenerative diseases.

Axonal TDP-43 Drives NMJ Disruption through Inhibition of Local Protein Synthesis

“…As we show, the NMJ relies on mitochondria activity (Fig. 5A-D) 31 and on local synthesis ( Fig. 5E-G).…”

“…3B vs. 3D). As NMJs are enriched with mitochondria 31,48,49 , and due to mitochondrial dependency on local synthesis 21,50 (Fig 4G-H), the enhanced local synthesis in NMJs can be attributed to its mitochondrial density. This suggests that the high polarization of MNs leads to higher dependency on local synthesis of mitochondrial proteins.…”

“…To study the effect of TDP-43 mislocalization on the NMJ in a precise and controlled environment, we used a neuromuscular co-culture setup in microfluidic chambers (MFCs) that we recently developed [28][29][30][31][32] . This platform can model several pathological features of ALS, such as axon degeneration, NMJ dysfunction and MN death 29,[33][34][35] .…”

“…This platform can model several pathological features of ALS, such as axon degeneration, NMJ dysfunction and MN death 29,[33][34][35] . The fluidic separation between the motor neuron cell-body and axon allows the formation of functional NMJs exclusively at the distal compartment 29,[31][32][33][34][35][36] . Utilizing this system, it is possible to study and track spatiotemporal events such as localized protein synthesis at the subcellular level.…”

Abnormal Mitochondrial Quality Control in Neurodegenerative Diseases

“…In addition, Altman et al (2019) showed that, compared with sympathetic neurons, cultured MNs displayed higher axonal and synaptic mitochondrial immobility. This finding suggests that the accumulation of mitochondria in the neuromuscular junction (NMJ) may play a significant role in MN function (Altman et al, 2019).…”