Single-Sensillum Taste Recordings in Mosquitoes.

In insects, gustatory neurons sense chemicals upon contact and directly inform many behaviors critical for survival and reproduction, including biting, feeding, mating, and egg laying. However, the taste sensory system is underexplored in many anthropophilic disease vectors such as mosquitoes, which acquire and transmit human pathogens during blood feeding from human hosts. This results in a big gap in vector biology-the study of organisms that spread disease by transmitting pathogens-because insect vectors closely interact with humans while selecting suitable individuals and appropriate bite sites for blood meals. Human sweat and skin-associated chemistries are rich in nonvolatile compounds that can be sensed by the mosquito's taste system when she lands on the skin. Taste sensory units, called sensilla, are distributed in many organs across the mosquito body, including the mouthparts, legs, and ovipositors (female-specific structures used to lay eggs). Each sensillum is innervated by as many as five taste neurons, which allow detection and discrimination between various tastants such as water, sugars, salts, amino acids, and plant-derived compounds that taste bitter to humans. Single-sensillum recordings provide a robust way to survey taste responsiveness of individual sensilla to various diagnostic and ecologically relevant chemicals. Such analyses are of immense value for understanding links between mosquito taste responses and behaviors to specific chemical cues and can provide insights into why mosquitoes prefer certain hosts. The results can also aid development of strategies to disrupt close-range mosquito-human interactions to control disease transmission. Here we describe a protocol that is curated for electrophysiological recordings from taste sensilla in mosquitoes and sure to yield exciting results for the field.

Taste Sensory Responses in Mosquitoes.

Analysis of taste sensory responses has been a powerful approach for understanding principles of taste detection and coding. The shared architecture of external taste sensing units, called sensilla, in insects opened up the study of tastant-evoked responses in any model of choice using a single-sensillum tip recording method that was developed in the mid-1900s. Early studies in blowflies were instrumental for identifying distinct taste neurons based on their responses to specific categories of chemicals. Broader system-wide analyses of whole organs have since been performed in the genetic model insect Drosophila melanogaster, revealing principles of stereotypical organization and function that appear to be evolutionarily conserved. Although limited in scope, investigations of taste sensory responses in mosquitoes showcase conservation in sensillar organization, as well as in groupings of functionally distinct taste neurons in each sensillum. The field is now poised for more thorough dissections of mosquito taste function, which should be of immense value in understanding close-range chemosensory interactions of mosquitoes with their hosts and environment. Here, we provide an introduction to the basic structure of a taste sensillum and functional analysis of the chemosensory neurons within it.

Control of sugar and amino acid feeding via pharyngeal taste neurons.

Insect gustatory systems comprise multiple taste organs for detecting chemicals that signal palatable or noxious quality. Although much is known about how taste neurons sense various chemicals, many questions remain about how individual taste neurons in each taste organ control feeding. Here, we use the Drosophila pharynx as a model to investigate how taste information is encoded at the cellular level to regulate consumption of sugars and amino acids. We first generate taste-blind animals and establish a critical role for pharyngeal input in food selection. We then investigate feeding behavior of both male and female flies in which only selected classes of pharyngeal neurons are restored via binary choice feeding preference assays as well as Fly Liquid-Food Interaction Counter (FLIC) assays. We find instances of integration as well as redundancy in how pharyngeal neurons control behavioral responses to sugars and amino acids. Additionally, we find that pharyngeal neurons drive sugar feeding preference based on sweet taste but not on nutritional value. Finally, we demonstrate functional specialization of pharyngeal and external neurons using optogenetic activation. Overall, our genetic taste neuron protection system in a taste-blind background provides a powerful approach to elucidate principles of pharyngeal taste coding and demonstrates functional overlap and subdivision among taste neurons.SIGNIFICANCE STATEMENTDietary intake of nutritious chemicals such as sugars and amino acids is essential for an animal's survival. In insects, distinct classes of taste neurons control acceptance or rejection of food sources. Here we develop a genetic system to investigate how individual taste neurons in the Drosophila pharynx encode specific tastants, focusing on sugars and amino acids. By examining flies in which only a single class of taste neurons is active, we find evidence for functional overlap as well as redundancy in responses to sugars and amino acids. We also uncover functional subdivision between pharyngeal and external neurons in driving feeding responses. Overall, we find that different pharyngeal neurons act together to control intake of the two categories of appetitive tastants.

Combinatorial Pharyngeal Taste Coding for Feeding Avoidance in Adult Drosophila.

Taste drives appropriate food preference and intake. In Drosophila, taste neurons are housed in both external and internal organs, but the latter have been relatively underexplored. Here, we report that Poxn mutants with a minimal taste system of pharyngeal neurons can avoid many aversive tastants, including bitter compounds, acid, and salt, suggesting that pharyngeal taste is sufficient for rejecting intake of aversive compounds. Optogenetic activation of selected pharyngeal bitter neurons during feeding events elicits changes in feeding parameters that can suppress intake. Functional dissection experiments indicate that multiple classes of pharyngeal neurons are involved in achieving behavioral avoidance, by virtue of being inhibited or activated by aversive tastants. Tracing second-order pharyngeal circuits reveals two main relay centers for processing pharyngeal taste inputs. Together, our results suggest that the pharynx can control the ingestion of harmful compounds by integrating taste input from different classes of pharyngeal neurons.