Quantitative behavioural analysis of the developing nervous system in Caenorhabditis elegans: a new approach to investigating anaesthesia-induced neurobehavioral impairment

Abstract

Introduction: Vertebrate animal experiments focused on a limited subset of behaviour have suggested that anaesthesia cause neuroapoptosis and behavioural impairment in developing nervous systems (anaesthesia-induced neurotoxicity, AIN). There is no universally-accepted phenotype for AIN; hence a priori determination of relevant behaviour may not be an appropriate outcome measure. An alternative approach is to study behaviour in less constrained settings. A rich subset of Caenorhabditis elegans natural behaviour can be studied under experimental conditions; these are amenable to automated extraction and analysis, and can be aggregated to yield a phenotypic ‘signature’. Perturbation of the signature provides a tool that can be used to quantify the effects of anaesthetic regimes, and act as an outcome marker for drug screening and molecular target research. Methods: The behaviour of free-moving larval and adult C. elegans was first mapped to form a baseline quantitative description of developmental behaviour. L1s were then exposed to common anaesthetic agents: isoflurane, ketamine, morphine, and dexmedetomidine, in mono- and combination therapy. Behaviour was recorded, and videos analysed with automated algorithms to extract behavioural and morphological features. Results: Wild-type C. elegans displays significant developmental-stage specific behavioural variation in 51 features after correcting for multiple comparisons. Anaesthetic exposure at L1 leads to persisting morphological and behavioural feature variation (in total, 114 features/exposure combinations). Higher concentrations of isoflurane, and combinations of isoflurane with ketamine, lead to the most significant feature hits, and these are most evident at the L3 stage. Discussion: C. elegans undergoes quantifiable behavioural changes as it transitions through developmental stages. Anaesthesia exposure at L1 leads to persisting behavioural change into later stages. Patterns correlate with vertebrate experiments, in that impairment is dependent on agent, is concentration-specific, and is more likely with combination therapies. C. elegans may therefore be a useful model with which to pursue drug discovery and mechanistic research in AIN.