Published: Vol 7, Iss 14, Jul 20, 2017 DOI: 10.21769/BioProtoc.2400 Views: 9115
Reviewed by: Neelanjan BoseLU HANManish Chamoli
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Abstract
Aldicarb treatment causes an accumulation of acetylcholine in the synaptic cleft of the neuromuscular junction, resulting in sustained muscle activation and eventually paralysis. Aldicarb-induced paralysis assay is an easy and fast method to determine whether synaptic transmission of a C. elegans mutant of interest is altered. This assay is based on the correlation of the rate of neurotransmitter release with the rate of paralysis. In this protocol, we describe a method for simultaneously assessing the aldicarb sensitivity of animals with different genotypes.
Keywords: AldicarbBackground
Synaptic transmission is initiated by arrival of action potential at presynaptic terminals, which in turn results in the release of neurotransmitters. The released neurotransmitters bind to and activate postsynaptic receptors (Sudhof, 2013). C. elegans locomotion is controlled by acetylcholine-releasing excitatory motor neurons and GABA (γ-aminobutyric acid)-releasing-inhibitory motor neurons (Richmond and Jorgensen, 1999; Zhen and Samuel, 2015). Acetylcholine released from cholinergic motor neurons activates acetylcholine receptors on the muscle membrane, leading to muscle excitation and contraction. Acetylcholinesterase breaks down acetylcholine in the synaptic cleft and thus terminates neurotransmission. Aldicarb is an acetylcholinesterase inhibitor. In the presence of aldicarb, acetylcholine continues to accumulate, causing persistent muscle contraction and eventual paralysis. Mutant animals with decreased levels of synaptic transmission are resistant to the paralyzing effect of aldicarb because acetylcholine more slowly accumulates in the synaptic cleft of these animals than that of wild type animals. Conversely, mutant animals with increased levels of synaptic transmission, and as a result, a faster accumulation of acetylcholine, are more sensitive to the paralyzing effect of aldicarb than wild-type animals (Rand, 2007). Thus, by comparing the time-course of aldicarb-induced paralysis, it is possible to infer the relative efficiency of synaptic transmission. However, it is necessary to note that this assay does not necessarily demonstrate that abnormal aldicarb sensitivity is a result of a presynaptic defect. For example, a defect in post-synaptic acetylcholine receptor function may also result in resistance to aldicarb (Loria et al., 2004). Thus, the aldicarb-induced paralysis assay should be considered as the first step in investigating the involvement of synaptic transmission, and further corroborated by other means, such as electrophysiological analysis (Richmond, 2006).
Materials and Reagents
Equipment
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Data analysis
Notes
Recipes
Note: We have used plates stored at 4 °C for up to 2 weeks without any issue.
Acknowledgments
The protocol has been adapted from Oh et al. (2017), eLife 6: e24733. This work was supported, in part, by a grant from the National Institute of Health.
References
Article Information
Copyright
Oh and Kim. This article is distributed under the terms of the Creative Commons Attribution License (CC BY 4.0).
How to cite
Readers should cite both the Bio-protocol article and the original research article where this protocol was used:
Category
Neuroscience > Behavioral neuroscience > Animal model
Neuroscience > Cellular mechanisms > Intracellular signalling
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