Recording central neurophysiological output from mosquito larvae for neuropharmacological and insecticide resistance studies

Authors: Norris, Edmund; BLOOMQUIST, JEFFREY - University Of Florida

Submitted to: Journal of Insect Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/2/2021
Publication Date: 10/7/2021
Citation: Norris, E.J., Bloomquist, J.R. 2021. Recording central neurophysiological output from mosquito larvae for neuropharmacological and insecticide resistance studies. Journal of Insect Physiology. 135:104319. https://doi.org/10.1016/j.jinsphys.2021.104319.
DOI: https://doi.org/10.1016/j.jinsphys.2021.104319

Interpretive Summary: Resistance to currently utilized chemical insecticidal agents represents a significant threat to public health and food security worldwide. Better understanding the physiological effects of market-available and candidate insecticidal molecules is valuable for characterizing the mechanisms of insecticide resistance, as well as for the design and study of novel control chemistries. As such, we sought to develop a method to record nerve firing from mosquito larvae, so that we could evaluate the effects of various neurotransmitters and insecticides directly on the mosquito central nervous system. In short, yellow fever mosquitoes (Aedes aegypti) mosquitoes were immobilized in a wax dish with small steel pins placed through the head and siphon. A longitudinal incision was made and the digestive tract of the mosquito larvae was removed. The nerve cord was then severed so that a recording electrode could record nerve firing in the ventral nerve cord of the mosquito larvae. A number of different excitatory/inhibitory neurotransmitters and insecticides were applied to the mosquito nervous system to better understand how these various agents affect mosquito neurophysiology. Furthermore, this method provides a new means to screen insecticide resistance directly on the mosquito nervous system. To validate this, the resistance of p,p’-dichlorodiphenyltrichlorethane (DDT) and its difluoro analog (DFDT) were characterized directly on the mosquito nervous system on the Puerto Rico strain of Aedes aegypti.

Technical Abstract: Resistance to currently utilized chemical insecticidal agents represents a significant threat to public health and food security worldwide. Better understanding the physiological effects of market-available and candidate insecticidal molecules is valuable for characterizing the mechanisms of insecticide resistance, as well as the design and study of novel control chemistries. In this paper, we describe a method of recording nerve firing from the central nervous system of Aedes aegypti fourth instar larvae. In short, mosquito larvae were immobilized by placing small pins through the head and siphon of the larvae in a wax dish. A single, longitudinal incision from the distal abdomen to the pronotum of the larva was made, the alimentary canal removed, and the ventral nerve cord severed between the second and third abdominal ganglia. A recording suction electrode was connected directly to axons within the severed end of the connective in a novel way to record nerve firing in the ventral nerve cord at high signal-to-noise ratio using conventional electrophysiological equipment. Using this novel method, we report the effects of four neuroactive compounds using this method: octopamine, pilocarpine, nicotine, and '-aminobutyric acid (GABA). The utility of this recording technique for elucidating target site mechanisms involved in insecticide resistance is demonstrated with p,p’-dichlorodiphenyltrichlorethane (DDT) and its difluoro analog (DFDT).