Automated screening ofC. elegansneurodegeneration mutants enabled by microfluidics and image analysis algorithms

Abstract

Spinal muscular atrophy (SMA) is a degenerative disorder that selectively deteriorates motor neurons due to a deficiency of survival motor neuron protein (SMN). The illness is the leading genetic cause of death in infants and is difficult to study in complex biological systems such as humans. A simpler model system, such as the nematode C. elegans, can be used to study potential mechanisms underlying this disease; C. elegans expresses the smn-1 gene, a homologue of SMN; powerful genetic tools in C. elegans research can be used to discover novel genes whose effect on SMN remains unknown or uncharacterized. Currently, conventional screening methods are time-consuming and laborious, as well as being subjective and mostly qualitative. To address these issues, we engineer an automated system capable of performing genetic suppressor screens on C. elegans using microfluidics in combination with custom image analysis software. We demonstrate the utility of this system by isolating 21 alleles that significantly suppress motor neuron degeneration at a screening rate of approximately 300 worms per hour. Many of these mutants also have improved motor function. These isolated alleles can potentially be further studied to understand mechanisms of protection against neurodegeneration. Our system is easily adaptable, providing a means to saturate screens not only implicated in the smn-1 pathway, but also for genes involved in other neurodegenerative phenotypes.

Quantitative Behavioural Phenotyping to Investigate Anaesthesia-Induced Neurobehavioural Impairment

“…These hits can then be further validated in vertebrate models or extended screens. Developments in robotics, microfluidics and image-analysis now allow for true high-throughput screening of nematodes 23,24,48 . C. elegans therefore is an excellent model with which to pursue drug discovery and mechanistic research.…”

“…For these reasons, until there is better mechanistic understanding of AIN, a high-throughput phenotypic screening method may be an important parallel line of investigation. This approach is difficult in mammals, but has been successfully demonstrated with other whole-animal models, in particularly the nematode worm Caenorhabditis elegans 23,24 . Because of its small size, short life cycle (Fig 1a), and ease of maintenance and propagation, C. elegans can be raised in large numbers in the lab.…”

Understanding the Effects of Deep Space Radiation on Nervous System: The Role of Genetically Tractable Experimental Models

“…Moreover, the degeneration of specific neuronal populations, can be easily analyzed in living animals [93], thanks to their transparency and the expression of fluorescent proteins. The choice of C. elegans is specifically justified by the following elements: a high resistance to extreme conditions (also thanks to a cuticle); the possibility of hibernation (i.e., as dauer larvae, a resistant larval stage) and of freezing; the self-fertilizing hermaphroditism that avoid the need for crossings; a large progeny (300 eggs per each animal); and the possibility to culture them in microfluidic devices [94].…”

Drug Screening and Drug Repositioning as Promising Therapeutic Approaches for Spinal Muscular Atrophy Treatment

“…Its small size, short life cycle, body transparency, ease to generate transgenic animals, and low maintenance costs contribute to its large use in experimental studies. Moreover its nervous and locomotor systems are well known, since its 302 neurons and 95 body wall muscles are all identified (C. elegans Sequencing Consortium, 1998; Dimitriadi and Hart, 2010;Wolozin et al, 2011;Cáceres et al, 2012;Lee et al, 2013;de Carlos Cáceres et al, 2018). To date, several C. elegans Smn mutants have been developed, such as smn-1(ok355) (null-mutant form) and smn-1(cb131).…”

“…These evaluations can be also correlated to MN degeneration analysis. To this aim, the most recent automated system has been developed by de Carlos Cáceres group: it employs microfluidic and image analysis that assess worm phenotypes analyzing D-type ventral MN degeneration through a quick genetic screening technique (De Carlos Cáceres et al, 2018).…”