Inter- and intracellular resistance and vulnerability in motor neuron diseases

Abstract

Amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA) are fatal diseases presenting with degeneration and loss of lower motor neurons in the brainstem and spinal cord. However, oculomotor and trochlear motor neurons that control eye movement, are resistant until late stages of disease. Moreover, individual motor neurons degenerate in a distinct pattern. Firstly, the distal synapse between motor neuron and muscle, the neuromuscular junction (NMJ), becomes denervated. Subsequently, the neuron degenerates in a retrograde manner, until the cell body in the spinal cord is lost later in disease. This highlights the distal axon and NMJ as the most vulnerable subcellular compartment of motor neurons.

We set out to investigate the mechanisms that underlie the resistance of oculomotor neurons, as well as the differential vulnerability within motor neurons themselves. We first validated the resistance of oculomotor neurons in two mouse models of ALS and SMA in Paper I. Next, we performed a large transcriptomic screen of differentially vulnerable motor pools in SMA mice throughout disease progression in Paper II. We used laser capture microdissection (LCM) to dissect motor neurons from different pools in the brainstem and spinal cord. Here we revealed pathways and transcripts that were enriched only in the resistant oculomotor neurons that serve to prevent them from going into apoptosis, as well as promote their regeneration. We showed that applying one of the oculomotor-enriched transcripts, Gdf15, to vulnerable spinal motor neurons derived from stem cells improved their survival. Next, in Paper III we utilised in vitro approaches and generated a model of oculomotor resistance in ALS by deriving both spinal and oculomotor neurons from mouse embryonic stem cells. Here we found that in vitro oculomotor neurons are more resistant to excitotoxic stress and have increased levels of prosurvival Akt-signaling, which also held up in LCM-dissected post-mortem human spinal and oculomotor neurons.

To investigate the vulnerability of the distal axon, we generated a technique called Axon-Seq in Paper IV, that allows transcriptomic profiling of isolated motor axons by culturing motor neurons in microfluidic chambers. We subsequently used this technique in Paper V to show that human stem cell-derived motor neurons carrying ALS-causative mutations in FUS and TDP-43 showed unique transcriptome dysregulation in somas and axons. This implies that different pathways of degeneration are at play between subcellular compartments, and provides novel targets for therapeutic intervention directed at different compartments of the motor neuron.