34Inhibition of acetylcholinesterase by either organophosphates or carbamates causes anti-35 cholinesterase poisoning. This arises through a wide range of neurotoxic effects triggered by the 36 overstimulation of the cholinergic receptors at synapses and neuromuscular junctions. Without 37 intervention, this poisoning can lead to profound toxic effects, including death, and the incomplete 38 efficacy of the current treatments, particularly for oxime-insensitive agents, provokes the need to find 39 better antidotes. Here we show how the non-parasitic nematode Caenorhabditis elegans offers an 40 excellent tool for investigating the acetylcholinesterase intoxication. The C. elegans neuromuscular 41 junctions show a high degree of molecular and functional conservation with the cholinergic 42 transmission that operates in the autonomic, central and neuromuscular synapses in mammals. In fact, 43 the anti-cholinesterase intoxication of the worm's body wall neuromuscular junction has been 44 unprecedented in understanding molecular determinants of cholinergic function in nematodes and 45 other organisms. We extend the use of the model organism's feeding behaviour as a tool to investigate 46 carbamate and organophosphate mode of action. We show that inhibition of the cholinergic-47 dependent rhythmic pumping of the pharyngeal muscle correlates with the inhibition of the 48 acetylcholinesterase activity caused by aldicarb, paraoxons and DFP exposure. Further, this bio-assay 49 allows one to address oxime dependent reversal of cholinesterase inhibition in the context of whole 50 organism recovery. Interestingly, the recovery of the pharyngeal function after such anti-51 cholinesterase poisoning represents a sensitive and easily quantifiable phenotype that is indicative of 52 the spontaneous recovery or irreversible modification of the worm acetylcholinesterase after 53 inhibition. These observations highlight the pharynx of C. elegans as a new tractable approach to 54 explore anti-cholinesterase intoxication and recovery with the potential to resolve critical genetic 55

Highlights C. elegans pharyngeal pumping inhibition by organophosphates correlates with worm acetylcholinesterase inhibition by the anticholinesterases.  The recovery of the pharyngeal function in C. elegans in the presence of obidoxime is due to the recovery of the acetylcholinesterase function after anti-cholinesterase intoxication.  The pharyngeal neuromuscular function represents a quantitative bioassay for investigation of anti-cholinesterase toxicity and recovery with excellent 3Rs potential.

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Synapses are specialized contact sites that mediate information flow between neurons and their targets. Important physical interactions across the synapse are mediated by synaptic adhesion molecules. These adhesions regulate formation of synapses during development and play a role during mature synaptic function. Importantly, genes regulating synaptogenesis and axon regeneration are conserved across the animal phyla. Genetic screens in the nematode Caenorhabditis elegans have identified a number of molecules required for synapse patterning and assembly. C. elegans is able to survive even with its neuronal function severely compromised. This is in comparison with Drosophila and mice where increased complexity makes them less tolerant to impaired function. Although this fact may reflect differences in the function of the homologous proteins in the synapses between these organisms, the most likely interpretation is that many of these components are equally important, but not absolutely essential, for synaptic transmission to support the relatively undemanding life style of laboratory maintained C. elegans. Here, we review research on the major group of synaptic proteins, involved in the presynaptic machinery in C. elegans, showing a strong conservation between higher organisms and highlight how C. elegans can be used as an informative tool for dissecting synaptic components, based on a simple nervous system organization.

Neuroligins are neuronal and neuromuscular transmembrane proteins that have been implicated in autism spectrum disorder and other cognitive diseases. The nlg-1 gene from Caenorhabditis elegans is orthologous to human neuroligin genes. In the nematode, the locomotory rate is mediated by dopaminergic and serotonergic pathways, which result in two different behavioral responses known as basal slowing response (BSR) and enhanced slowing response (ESR), respectively. We report that nlg-1-deficient mutants are defective in both the BSR and ESR behaviors. In addition, we demonstrate that methylphenidate (a dopamine reuptake inhibitor) and fluoxetine (a serotonin reuptake inhibitor), two drugs widely used for the treatment of behavioral disorders in humans, are able to restore the BSR and ESR wild type phenotypes, respectively, in nlg-1 defective mutant nematodes. The abnormal locomotory behavior patterns were rescued in nlg-1-deficient mutant by expressing a cDNA from the human NLGN1 gene under the C. elegans nlg-1 promoter. However, human NLGN1 (R453C) and NLGN1 (D432X) mutant alleles did not rescue any of the two mutant phenotypes. The results indicate that neuroligin is involved in modulating the action of dopamine and serotonin in the nematode and suggest that the functional mechanism underpinning both methylphenidate and fluoxetine in C. elegans might be comparable to that in humans. The neuroligin-deficient mutants may undergo inefficient synaptic transmissions which could affect different traits in the nervous system. In particular, neuroligin might be required for normal neurotransmitters release. The understanding of the mechanisms by which methylphenidate and fluoxetine are able to restore the behavior of these mutants could help to explain the etiology of some human neurological diseases.