Imaging Net Retrograde Axonal Transport In Vivo: A Physiological Biomarker

Abstract

Objective The objective of this study is to develop a novel method for monitoring the integrity of motor neurons in vivo by quantifying net retrograde axonal transport. Methods The method uses single photon emission computed tomography to quantify retrograde transport to spinal cord of tetanus toxin fragment C (125I‐TTC) following intramuscular injection. We characterized the transport profiles in 3 transgenic mouse models carrying amyotrophic lateral sclerosis (ALS)‐associated genes, aging mice, and SOD1G93A transgenic mice following CRISPR/Cas9 gene editing. Lastly, we studied the effect of prior immunization of tetanus toxoid on the transport profile of TTC. Results This technique defines a quantitative profile of net retrograde axonal transport of TTC in living mice. The profile is distinctly abnormal in transgenic SOD1G93A mice as young as 65 days (presymptomatic) and worsens with disease progression. Moreover, this method detects a distinct therapeutic benefit of gene editing in transgenic SOD1G93A mice well before other clinical parameters (eg, grip strength) show improvement. Symptomatic transgenic PFN1C71G/C71G ALS mice display gross reductions in net retrograde axonal transport, which is also disturbed in asymptomatic mice harboring a human C9ORF72 transgene with an expanded GGGGCC repeat motif. In wild‐type mice, net retrograde axonal transport declines with aging. Lastly, prior immunization with tetanus toxoid does not preclude use of this assay. Interpretation This assay of net retrograde axonal transport has broad potential clinical applications and should be particularly valuable as a physiological biomarker that permits early detection of benefit from potential therapies for motor neuron diseases. ANN NEUROL 2022;91:716–729

Neuroscience highlights in 2022: cytoskeletal transport

“…Through injection of a radiolabelled, non-toxic fragment of tetanus neurotoxin into the mouse gastrocnemius muscle, it was recently shown that single photon emission computed tomography applied to the lumbar spinal cord can be used to quantify net retrograde axonal transport of endosomes in vivo. 8 Confirming evidence obtained using intravital imaging approaches, 1,9 this technique identified trafficking decline with aging, as well as disruptions in mice bearing mutations in the ALS/frontotemporal dementia (FTD) genes SOD1, C9ORF72 and PFN1. Notably, transport disruption was rescued in gene-edited mutant SOD1 mice, with the benefit being detectable well before behavioural improvements.…”

Engineering Neurotoxin-Functionalized Exosomes for Targeted Delivery to the Peripheral Nervous System

“…Conversely, the non-targeted Un-Exo group shows uneven distribution of Exos across the tissue and aggregation in a particular location along the nerve fiber. These observations suggest that TTC-Exos travel via a retrograde axonal transport mechanism along the PNS by binding to polysialoganglioside and presynaptic receptors on neurons, facilitating inter-neuronal transfer across peripheral nerves [ 46 , 47 ].…”

Neuroanatomical Tract Tracers: Not Just for Neural Tracing Anymore

“…injection. 82 Three transgenic amyotrophic lateral sclerosis (ALS) mouse models were used in this experiment to obtain insight into the potential of this technique for the early detection of neurodegenerative pathology. Extending studies, which utilized TTC displacement assays to determine the binding affinity of TTC to its receptors expressed in synaptosomes, such as realtime monitoring of radiolabeled NaTT retrograde transport from muscle to motor neurons allow for the quantitative assessment of transduction efficiency.…”

“…To improve this, radiolabeled TTC ( 125 I-TTC) was used in conjunction with SPECT to quantify 125 I-TTC retrograde transport from a limb muscle to the spinal cord via the sciatic nerve following i.m . injection . Three transgenic amyotrophic lateral sclerosis (ALS) mouse models were used in this experiment to obtain insight into the potential of this technique for the early detection of neurodegenerative pathology.…”