An analysis of ethanol-induced behavioural plasticity in Caenorhabditis elegans

Ethanol is one of the most widely used and socially acceptable drugs in the world. However its chronic use can lead to serious problems including the development of dependence. Alcohol dependence is a chronic, relapsing disorder characterised by tolerance, withdrawal, preoccupation with obtaining alcohol, loss of control over its consumption and impairment in social and occupational functioning. In humans this develops over years, primarily driven by adaptations in many distinct signalling pathways and neural circuits as a result of continued heavy drinking. Whilst alcohol dependence has been extensively studied our understanding of how its distinct targets integrate to produce various behavioural responses remains far from clear. The nematode worm Caenorhabditis elegans is a model genetic organism with a simple nervous system and well-defined behaviour. These nematodes can display plasticity in the form of tolerance to, and withdrawal from, 5-HT or nicotine. They are thus a genetically tractable system in which to investigate the neural substrates of adaptive responses to ethanol. In this simple system the impact of changes at the molecular level on signalling in defined neural circuits and the resultant animal behaviour can be investigated. The aims of this thesis were to establish a C. elegans paradigm for alcohol dependence and to use this to define the genetic basis of the ethanol-dependent behaviours of intoxication, tolerance and withdrawal. Evidence was provided that ethanol equilibrates rapidly across the worm cuticle indicating that the internal concentration closely approximates to the external concentration in which the animal is placed. Ethanol-dependent behaviours were carefully characterised using a variety of behavioural assays. C. elegans exhibit distinct behavioural states, corresponding to intoxication and withdrawal, which impair the ability to navigate towards food. Visual and automated analysis defined a sub-behaviour, an increased tendency to form spontaneous deep body bends, which was specifically associated with withdrawal. This was ameliorated by a low dose of alcohol supporting the contention that it arises from ethanol-induced neuroadaptation. A series of loss of function mutants, were analysed for alterations in ethanoldependent behaviour. The absence of withdrawal in a strain of worms depleted in neuropeptides (egl-3) demonstrated that peptidergic signalling is key to the chronic adaption to, but not to the acute effects of, ethanol. However the neuropeptide receptor NPR-1, previously shown to impact on ethanol responses in C. elegans, had no effect on withdrawal behaviour in these assays. Alterations in intoxication and withdrawal behaviour in strains of worms depleted in 5-HT (tph-1) and dopamine (cat-2) indicated that serotonergic and dopaminergic signalling may also be involved in the ethanol response in C. elegans. This study has therefore provided a quantitative analysis of distinct ethanol-induced behavioural states and highlighted a role for neuropeptides and major classes of neuromodulatory transmitters. In particular this data is consistent with the emerging role of neuropeptides in ethanol withdrawal.