Automated in vivo drug screen in zebrafish identifies synapse-stabilising drugs with relevance to spinal muscular atrophy

Abstract

Synapses are particularly vulnerable in many neurodegenerative diseases and often the first to degenerate, for example in the motor neuron disease spinal muscular atrophy (SMA). Compounds that can counteract synaptic destabilisation are rare. Here, we describe an automated screening paradigm in zebrafish for small-molecule compounds that stabilize the neuromuscular synapse in vivo. We make use of a mutant for the axonal C-type lectin chondrolectin (chodl), one of the main genes dysregulated in SMA. In chodl−/− mutants, neuromuscular synapses that are formed at the first synaptic site by growing axons are not fully mature, causing axons to stall, thereby impeding further axon growth beyond that synaptic site. This makes axon length a convenient read-out for synapse stability. We screened 982 small-molecule compounds in chodl chodl−/− mutants and found four that strongly rescued motor axon length. Aberrant presynaptic neuromuscular synapse morphology was also corrected. The most-effective compound, the adenosine uptake inhibitor drug dipyridamole, also rescued axon growth defects in the UBA1-dependent zebrafish model of SMA. Hence, we describe an automated screening pipeline that can detect compounds with relevance to SMA. This versatile platform can be used for drug and genetic screens, with wider relevance to synapse formation and stabilisation.

Bioluminescent Zebrafish Transplantation Model for Drug Discovery

“…Usually, a trade-off average concentration has to be selected within the activity range of all small-molecule compounds used. This value is generally within the range of 1–15 µM in most zebrafish in vivo pharmaceutical screens ( Bowman and Zon, 2010 ; Colanesi et al, 2012 ; Precazzini et al, 2020 ; Haney et al, 2021 ; Oprisoreanu et al, 2021 ). Therefore, we decided to use 10 µM as our selected in vivo screening concentration.…”

Bioluminescent zebrafish transplantation model for drug discovery

“…Usually, a trade-off average concentration has to be selected within the activity range of all small-molecule compounds used. This value is generally within the range of 1 – 15 μM in most zebrafish in vivo pharmaceutical screens (Bowman and Zon, 2010; Colanesi et al, 2012; Haney et al, 2021; Oprisoreanu et al, 2021; Precazzini et al, 2020). Therefore, we decided to use 10 μM as our selected in vivo screening concentration.…”

A ligand-receptor interactome atlas of the zebrafish

“…The presence of several conserved ligand-receptor pairs in the zebrafish synapses further stresses the importance of zebrafish in understanding the role of ligand-receptor in synapse biology and in drugs targeting synapses (Hoffman et al, 2016;Hutson and Chien, 2002;Oprişoreanu et al, 2021). Although our synaptosome and PSD ligandreceptor pairs lack spatiotemporal accuracy, DanioTalk can be further combined with zebrafish single-cell brain transcriptome records to identify synaptosome-PSD ligand-receptor interactions for neuronal cell types of interest (Farnsworth et al, 2020;Raj et al, 2020).…”