preparing the recording electrodes and drosophila for analyzing responses of taste neurons

A Novel Method for Enhanced Analysis of Insect Taste Neuron Activity

Insects, including drosophilid flies, are endowed with a sophisticated taste system that enables them to extract complex chemical information from their surroundings. This system allows them to discern the chemical composition of various substances, distinguishing between those that are nutritious and those that are harmful1,2.
At the core of this system are specialized structures known as taste hairs or sensilla, strategically located on various body parts. In drosophilid flies, these sensilla are located on the labellum, which is the major taste organ of the fly head1,2,3,4, as well as on the legs and wings1,2,5,6. The labellum is located at the tip of the proboscis and contains two lobes4,7,8. Each lobe is covered with 31 taste sensilla categorized as short, long, and intermediate4,7,8. These sensilla each house 2-4 taste neurons1,2,9,10. These taste neurons express members of at least four different gene families, namely the Gustatory receptor (Gr), Ionotropic receptor (Ir), Pickpocket (Ppk), and Transient receptor potential (Trp) genes1,2,11,12,13. This diversity of receptors and channels equips insects with the ability to recognize a wide array of chemical compounds, including both nonvolatile and volatile cues1,2,14.
For over 50 years, scientists have quantified the response of taste neurons and their receptors using a technique called tip recording3,4,6,8,13,15,16,17,18,19,20,21,22,23,24,25,26,27,28, 
29,30,31,32,33,34,35. However, this method has major limitations. First, neural activity can be measured only during contact with the stimulus, and not before or after contact. This limitation precludes the measurement of spontaneous spiking activity and prevents the measurement of OFF responses. Second, only tastants that are soluble in aqueous solutions can be tested.
These limitations can be overcome by a rarely used alternative electrophysiological technique called &#34;base recording.&#34; Here we describe this technique, which we have adapted from a method used by Marion-Poll and colleagues24, and show the crucial taste coding features that it can now conveniently measure14.

Author Spotlight: Advances in Chemoreception &#8211; From Insect Odor Receptors to Non-Coding RNAs

Baseoptagelse: En teknik til analyse af reaktioner fra smagsneuroner i Drosophila

Forberedelse af optageelektroderne og drosophila til analyse af responser af smagsneuroner

Begynd med at placere 10 til 15 nyopståede Drosophila-fluer i friske standardkulturhætteglas. Oprethold disse hætteglas ved 25 grader Celsius og 60% relativ luftfugtighed i en 12-timers lys og mørk cyklus. For at forberede en glaskapillær til at holde stimulus skal du vælge en borosilikatglaskapillær og bruge et pipetteudtrækkerinstrument til at trække glaskapillæren.Tag derefter wolframstænger til både reference- og optagelseselektroder. Dyp disse elektroder gentagne gange i flere sekunder i enten en 10% kaliumnitratopløsning eller en kaliumhydroxidopløsning for at skærpe dem til en diameter på en mikrometer spids. For at trække en enkelt flue fra hætteglasset til en aspirator skal du forsigtigt trække aspiratoren ud og placere en finger over enden af aspiratoren for at fange fluen.Hold nu enden af aspiratoren i pipettespidsen på 200 mikroliter, og brug enden til at skubbe fluen fremad med hovedet først mod den smalle ende af pipettespidsen. Brug et barberblad til at trimme begge ender af pipettespidsen, forreste og bageste til fluen. Ved hjælp af ler eller et lille stykke bomuld skubbes fluen yderligere fremad, indtil halvdelen af hovedet stikker ud fra enden af den trimmede pipettespids.Brug pincet til forsigtigt at udsætte labellum foran på hovedet. Under et stereomikroskop skal du placere etiketten sideværts på en dækseddel, og sørg for, at en lap med dens 31 smagssans eksponeres.

Research

JoVE Journal

Neuroscience

Preparing the Recording Electrodes and Drosophila for Analyzing Responses of Taste Neurons

Base Recording: A Technique for Analyzing Responses of Taste Neurons in Drosophila

Insects taste the external world through taste hairs, or sensilla, that have pores at their tips. When a sensillum comes into contact with a potential ...

Base Recording from Taste Sensilla of Drosophila for Analyzing Responses of Taste Neurons

Baseoptagelse fra smagssensilla af Drosophila til analyse af responser fra smagsneuroner