Name: Author Spotlight: Advances in Chemoreception &#8211; From Insect Odor Receptors to Non-Coding RNAs
Uploaded: 2024-03-01T15:00:00
Description: Read this Scientific Article Protocol about Author Spotlight: Advances in Chemoreception at JoVE.com

Vi takker Zina Berman for støtte, Lisa Baik for kommentarer til manuskriptet og andre medlemmer af Carlson-laboratoriet for diskussion. Dette arbejde blev støttet NIH bevilling K01 DC020145 til H.K.M.D; og NIH giver R01 DC02174, R01 DC04729 og R01 DC011697 til JRC.

Følgende protokol overholder alle retningslinjer for dyrepleje fra Yale University.
1. Fluer
<ol><li>Anbring 10-15 nyfremkomne fluer i hætteglas med frisk standardkultur ved 25 °C og 60 % relativ luftfugtighed i en lys-mørk cyklus på 12:12 timer.</li>
	<li>Brug fluer, når de er 3-7 dage gamle.</li>
</ol>2. Kemosensoriske stimuli
<ol><li>Få kemosensoriske stimuli af den højeste tilgængelige renhed. Opbevar dem som anbefalet ...

A Novel Method for Enhanced Analysis of Insect Taste Neuron Activity

<ol>
	<li>Joseph, R. M., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+chemoreceptors:+a+molecular+interface+between+the+chemical+world+and+the+brain.">Drosophila chemoreceptors: a molecular interface between the chemical world and the brain.</a> Trends Genet. 31 (12), 683-695 (2015).</li><li>Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+sensory+receptors-a+set+of+molecular+Swiss+Army+Knives.">Drosophila sensory receptors-a set of molecular Swiss Army Knives.</a> Genetics. 217 (1), 1-34 (2021).</li><li>Dweck, H. K. M., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+logic+and+evolution+of+bitter+taste+in+Drosophila.">Molecular logic and evolution of bitter taste in Drosophila.</a> Curr Biol. 30 (1), 17-30 (2020).</li><li>Weiss, L. A., Dahanukar, A., Kwon, J. Y., Banerjee, D., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+molecular+and+cellular+basis+of+bitter+taste+in+Drosophila.">The molecular and cellular basis of bitter taste in Drosophila.</a> Neuron. 69 (2), 258-272 (2011).</li><li>He, Z., Luo, Y., Shang, X., Sun, J. S., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemosensory+sensilla+of+the+Drosophila+wing+express+a+candidate+ionotropic+pheromone+receptor.">Chemosensory sensilla of the Drosophila wing express a candidate ionotropic pheromone receptor.</a> PLoS Biol. 17 (5), e2006619 (2019).</li><li>Ling, F., Dahanukar, A., Weiss, L. A., Kwon, J. Y., Carlson, J. 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R., Park, S. K., Pikielny, C. W., Steinbrecht, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gustatory+organs+of+Drosophila+melanogaster:+fine+structure+and+expression+of+the+putative+odorant-binding+protein+PBPRP2.">Gustatory organs of Drosophila melanogaster: fine structure and expression of the putative odorant-binding protein PBPRP2.</a> Cell Tissue Res. 304 (3), 423-437 (2001).</li><li>Siddiqi, O., Rodrigues, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+analysis+of+a+complex+chemoreceptor.">Genetic analysis of a complex chemoreceptor.</a> Basic Life Sci. 16, 347-359 (1980).</li><li>Clyne, P. J., Warr, C. G., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Candidate+taste+receptors+in+Drosophila.">Candidate taste receptors in Drosophila.</a> Science. 287 (5459), 1830-1834 (2000).</li><li>Koh, T. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Drosophila+IR20a+clade+of+ionotropic+receptors+are+candidate+taste+and+pheromone+receptors.">The Drosophila IR20a clade of ionotropic receptors are candidate taste and pheromone receptors.</a> Neuron. 83 (4), 850-865 (2014).</li><li>S&#225;nchez-Alca&#241;iz, J. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+expression+atlas+of+variant+ionotropic+glutamate+receptors+identifies+a+molecular+basis+of+carbonation+sensing.">An expression atlas of variant ionotropic glutamate receptors identifies a molecular basis of carbonation sensing.</a> Nat Commun. 9 (1), 4252 (2018).</li><li>Dweck, H. K. M., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diverse+mechanisms+of+taste+coding+in+Drosophila.">Diverse mechanisms of taste coding in Drosophila.</a> Sci Adv. 9 (46), (2023).</li><li>Chyb, S., Dahanukar, A., Wickens, A., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+Gr5a+encodes+a+taste+receptor+tuned+to+trehalose.">Drosophila Gr5a encodes a taste receptor tuned to trehalose.</a> Proc Natl Acad Sci U S A. 100, 14526-14530 (2003).</li><li>Dahanukar, A., Lei, Y. T., Kwon, J. Y., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+Gr+genes+underlie+sugar+reception+in+Drosophila.">Two Gr genes underlie sugar reception in Drosophila.</a> Neuron. 56 (3), 503-516 (2007).</li><li>Delventhal, R., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bitter+taste+receptors+confer+diverse+functions+to+neurons.">Bitter taste receptors confer diverse functions to neurons.</a> Elife. 5, e11181 (2016).</li><li>Dweck, H. K. M., Talross, G. J. S., Luo, Y., Ebrahim, S. A. M., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ir56b+is+an+atypical+ionotropic+receptor+that+underlies+appetitive+salt+response+in+Drosophila.">Ir56b is an atypical ionotropic receptor that underlies appetitive salt response in Drosophila.</a> Curr Biol. 32 (8), 1776-1787 (2022).</li><li>Hiroi, M., Marion-Poll, F., Tanimura, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differentiated+response+to+sugars+among+labellar+chemosensilla+in+Drosophila.">Differentiated response to sugars among labellar chemosensilla in Drosophila.</a> Zoolog Sci. 19 (9), 1009-1018 (2002).</li><li>Jeong, Y. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+odorant-binding+protein+required+for+suppression+of+sweet+taste+by+bitter+chemicals.">An odorant-binding protein required for suppression of sweet taste by bitter chemicals.</a> Neuron. 79 (4), 725-737 (2013).</li><li>Jiao, Y., Moon, S. J., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Drosophila+gustatory+receptor+required+for+the+responses+to+sucrose,+glucose,+and+maltose+identified+by+mRNA+tagging.">A Drosophila gustatory receptor required for the responses to sucrose, glucose, and maltose identified by mRNA tagging.</a> Proc Natl Acad Sci U S A. 104 (35), 14110-14115 (2007).</li><li>Jiao, Y., Moon, S. J., Wang, X., Ren, Q., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gr64f+is+required+in+combination+with+other+gustatory+receptors+for+sugar+detection+in+Drosophila.">Gr64f is required in combination with other gustatory receptors for sugar detection in Drosophila.</a> Curr Biol. 18 (22), 1797-1801 (2008).</li><li>Kim, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+TRPA1+channel+mediates+chemical+avoidance+in+gustatory+receptor+neurons.">Drosophila TRPA1 channel mediates chemical avoidance in gustatory receptor neurons.</a> Proc Natl Acad Sci U S A. 107 (18), 8440-8445 (2010).</li><li>Lacaille, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+inhibitory+sex+pheromone+tastes+bitter+for+Drosophila+males.">An inhibitory sex pheromone tastes bitter for Drosophila males.</a> PLoS One. 2 (7), e661 (2007).</li><li>Lee, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gustatory+receptors+required+for+avoiding+the+insecticide+L-canavanine.">Gustatory receptors required for avoiding the insecticide L-canavanine.</a> J Neurosci. 32 (4), 1429-1435 (2012).</li><li>Lee, Y., Kim, S. H., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Avoiding+DEET+through+insect+gustatory+receptors.">Avoiding DEET through insect gustatory receptors.</a> Neuron. 67 (4), 555-561 (2010).</li><li>Lee, Y., Moon, S. J., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+gustatory+receptors+required+for+the+caffeine+response+in+Drosophila.">Multiple gustatory receptors required for the caffeine response in Drosophila.</a> Proc Natl Acad Sci U S A. 106 (11), 4495-4500 (2009).</li><li>Lee, Y., Moon, S. J., Wang, Y., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Drosophila+gustatory+receptor+required+for+strychnine+sensation.">A Drosophila gustatory receptor required for strychnine sensation.</a> Chem Senses. 40 (7), 525-533 (2015).</li><li>Meunier, N., Marion-Poll, F., Rospars, J. P., Tanimura, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peripheral+coding+of+bitter+taste+in+Drosophila.">Peripheral coding of bitter taste in Drosophila.</a> J Neurobiol. 56 (2), 139-152 (2003).</li><li>Moon, S. 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J., Lee, Y., Jiao, Y., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Drosophila+gustatory+receptor+essential+for+aversive+taste+and+inhibiting+male-to-male+courtship.">A Drosophila gustatory receptor essential for aversive taste and inhibiting male-to-male courtship.</a> Current Biology. 19, 1623-1627 (2009).</li><li>Rimal, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanism+of+acetic+acid+gustatory+repulsion+in+Drosophila.">Mechanism of acetic acid gustatory repulsion in Drosophila.</a> Cell Rep. 26 (6), 1432-1442 (2019).</li><li>Shim, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+full+repertoire+of+Drosophila+gustatory+receptors+for+detecting+an+aversive+compound.">The full repertoire of Drosophila gustatory receptors for detecting an aversive compound.</a> Nat Commun. 6, 8867 (2015).</li><li>Xiao, S., Baik, L. S., Shang, X., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Meeting+a+threat+of+the+Anthropocene:+Taste+avoidance+of+metal+ions+by+Drosophila.">Meeting a threat of the Anthropocene: Taste avoidance of metal ions by Drosophila.</a> Proc Natl Acad Sci U S A. 119 (25), e2204238119 (2022).</li><li>Zhang, Y. V., Ni, J., Montell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+molecular+basis+for+attractive+salt-taste+coding+in+Drosophila.">The molecular basis for attractive salt-taste coding in Drosophila.</a> Science. 340 (6138), 1334-1338 (2013).</li><li>Delventhal, R., Kiely, A., Carlson, J. 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(84), e51355 (2014).</li><li>Marella, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+taste+responses+in+the+fly+brain+reveals+a+functional+map+of+taste+category+and+behavior.">Imaging taste responses in the fly brain reveals a functional map of taste category and behavior.</a> Neuron. 49 (2), 285-295 (2006).</li><li>Thorne, N., Amrein, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atypical+expression+of+Drosophila+gustatory+receptor+genes+in+sensory+and+central+neurons.">Atypical expression of Drosophila gustatory receptor genes in sensory and central neurons.</a> J Comp Neurol. 506 (4), 548-568 (2008).</li><li>Wang, Z., Singhvi, A., Kong, P., Scott, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Taste+representations+in+the+Drosophila+brain.">Taste representations in the Drosophila brain.</a> Cell. 117 (7), 981-991 (2004).</li></ol>

I optagelser fra nogle typer sensila kan det være udfordrende at skelne piggene i forskellige neuroner. For eksempel producerer sukkerneuronerne og mekanosensoriske neuroner i S og I sensilla pigge med lignende amplituder, hvilket gør det vanskeligt at skelne dem 4,14. Vi finder, at brugen af en meget skarp wolframoptagelseselektrode reducerer affyringen af det mekanosensoriske neuron, ligesom den velovervejede placering af optageelektroden. Indsættelse af opt...

Insekter, herunder drosophilidfluer, er udstyret med et sofistikeret smagssystem, der gør det muligt for dem at udtrække komplekse kemiske oplysninger fra deres omgivelser. Dette system giver dem mulighed for at skelne den kemiske sammensætning af forskellige stoffer og skelne mellem dem, der er nærende og dem, der er skadelige 1,2.
Kernen i dette system er specialiserede strukturer kendt som smagshår eller sensilla, strategisk placeret på forskellige kropsdele. Hos drosophilidfluer er disse sensillaer placeret på etiketten, som er fluehovedets vigtigste smagsorgan 1,2,3,4, samt på ben og vinger 1,2,5,6. Labellum er placeret på spidsen af proboscis og indeholder to lapper 4,7,8. Hver lap er dækket med 31 smagssensiler kategoriseret som kort, lang og mellemliggende 4,7,8. Disse sensilla hvert hus 2-4 smag neuroner 1,2,9,10. Disse smagsneuroner udtrykker medlemmer af mindst fire forskellige genfamilier, nemlig gustatorisk receptor (Gr), ionotropisk receptor (Ir), lommetyv (ppk) og forbigående receptorpotentiale (Trp) gener 1,2,11,12,13 . Denne mangfoldighed af receptorer og kanaler udstyrer insekter med evnen til at genkende en bred vifte af kemiske forbindelser, herunder både ikke-flygtige og flygtige signaler 1,2,14.
I over 50 år har forskere kvantificeret responsen af smagsneuroner og deres receptorer ved hjælp af en teknik kaldet tip recording 3,4,6,8,13,15,16,17,18,19,20,21,22,23,24 ,25,26,27,28, 29,30,31,32,33,34,35. Denne metode har dog store begrænsninger. For det første kan neural aktivitet kun måles under kontakt med stimulus og ikke før eller efter kontakt. Denne begrænsning udelukker måling af spontan spidsaktivitet og forhindrer måling af OFF-responser. For det andet kan kun smagsstoffer, der er opløselige i vandige opløsninger, testes.
Disse begrænsninger kan overvindes ved en sjældent anvendt alternativ elektrofysiologisk teknik kaldet "baseoptagelse". Her beskriver vi denne teknik, som vi har tilpasset fra en metode, der anvendes af Marion-Poll og kolleger24, og viser de afgørende smagskodningsfunktioner, som den nu bekvemt kan måle14.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Microscope</td>
			<td>Olympus</td>
			<td>BX51WI</td>
			<td>equipped with a 50X objective (LMPLFLN 50X, Olympus) and 10X eyepieces.&#160;</td>
		</tr>
		<tr>
			<td>Antivibration Table</td>
			<td>TMC</td>
			<td>63-7590E</td>
			<td></td>
		</tr>
		<tr>
			<td>motorized Micromanipulators</td>
			<td>Harvard Apparatus and M&#228;rzh&#228;user Micromanipulators</td>
			<td>Micromanipulator PM 10 Piezo Micromanipulator</td>
			<td></td>
		</tr>
		<tr>
			<td>manual Micromanipulators</td>
			<td>M&#228;rzh&#228;user Micromanipulators</td>
			<td>MM33 Micromanipulator</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic stands</td>
			<td>ENCO</td>
			<td>Model #625-0930</td>
			<td></td>
		</tr>
		<tr>
			<td>Reference&#160; and recording Electrode Holder</td>
			<td>Ockenfels Syntech GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Stimulus glass capillary Holder</td>
			<td>Ockenfels Syntech GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Universal Single Ended Probe</td>
			<td>Ockenfels Syntech GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>4-CHANNEL USB ACQUISITION CONTROLLER , IDAC-4</td>
			<td>Ockenfels Syntech GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Stimulus Controllers</td>
			<td>Ockenfels Syntech GmbH</td>
			<td>Stimulus Controller CS 55</td>
			<td></td>
		</tr>
		<tr>
			<td>Personal Computer</td>
			<td>Dell</td>
			<td>Vostro</td>
			<td>Check for compatibility with digital acquisition system and software</td>
		</tr>
		<tr>
			<td>Tungsten Rod</td>
			<td>A-M Systems</td>
			<td>Cat#716000</td>
			<td></td>
		</tr>
		<tr>
			<td>Aluminum Foil and/or Faraday Cage</td>
			<td></td>
			<td></td>
			<td>Electromagnetic noise shielding</td>
		</tr>
		<tr>
			<td>Borosilicate Glass Capillaries</td>
			<td>World Precision Instruments</td>
			<td>1B100F-4</td>
			<td></td>
		</tr>
		<tr>
			<td>Pipette Puller</td>
			<td>Sutter Instrument Company</td>
			<td>Model P-97 Flaming/Brown Micropipette Puller</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereomicroscope</td>
			<td>Olympus</td>
			<td>VMZ 1x-4x</td>
			<td>For fly preparation</td>
		</tr>
		<tr>
			<td>p200 Pipette Tips</td>
			<td></td>
			<td></td>
			<td>Generic</td>
		</tr>
		<tr>
			<td>Microloader tips&#160;</td>
			<td>Eppendorf</td>
			<td>E5242956003</td>
			<td></td>
		</tr>
		<tr>
			<td>1 ml Syringe</td>
			<td></td>
			<td></td>
			<td>Generic</td>
		</tr>
		<tr>
			<td>Crocodile clips</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Power Transformers</td>
			<td>STACO ENERGY PRODUCTS</td>
			<td>STACO 3PN221B</td>
			<td>Assembled from P1000 pipette tips, flexible plastic tubing, and mesh</td>
		</tr>
		<tr>
			<td>Modeling Clay</td>
			<td></td>
			<td></td>
			<td>Generic</td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td></td>
			<td></td>
			<td>Generic</td>
		</tr>
		<tr>
			<td>Plastic Tubing</td>
			<td>Saint Gobain</td>
			<td>Tygon S3&#8482; E-3603</td>
			<td></td>
		</tr>
		<tr>
			<td>Standard culture vials</td>
			<td>Archon Scientific</td>
			<td></td>
			<td>Narrow 1-oz polystyrene vails, each with 10 mL of glucose medium, preloaded with cellulose acetate plugs</td>
		</tr>
		<tr>
			<td>Berberine chloride (BER)</td>
			<td>Sigma-Aldrich</td>
			<td>Cat# Y0001149</td>
			<td></td>
		</tr>
		<tr>
			<td>Denatonium benzoate (DEN)</td>
			<td>Sigma-Aldrich</td>
			<td>Cat# D5765</td>
			<td></td>
		</tr>
		<tr>
			<td>N,N-Diethyl-m- toluamide (DEET)</td>
			<td>Sigma-Aldrich</td>
			<td>Cat# 36542</td>
			<td></td>
		</tr>
	</tbody>
</table>

base recording: a technique for analyzing responses of taste neurons in drosophila

Insekter smager den ydre verden gennem smagshår eller sensilla, der har porer i spidserne. Når en sensillum kommer i kontakt med en potentiel fødekilde, kommer forbindelser fra fødekilden ind gennem porerne og aktiverer neuroner indeni. I over 50 år er disse reaktioner blevet registreret ved hjælp af en teknik kaldet tipoptagelse. Denne metode har imidlertid store begrænsninger, herunder manglende evne til at måle neural aktivitet før eller efter stimuluskontakt og kravet om, at smagsstoffer skal være opløselige i vandige opløsninger. Vi beskriver her en teknik, som vi kalder baseoptagelse, som overvinder disse begrænsninger. Baseoptagelse tillader måling af smagsneuronaktivitet før, under og efter stimulus. Det tillader således omfattende analyse af OFF-reaktioner, der opstår efter en smagsstimulus. Det kan bruges til at studere hydrofobe forbindelser såsom langkædede feromoner, der har meget lav opløselighed i vand. Sammenfattende tilbyder baseoptagelse fordelene ved single-sensillum elektrofysiologi som et middel til måling af neuronal aktivitet - høj rumlig og tidsmæssig opløsning uden behov for genetiske værktøjer - og overvinder vigtige begrænsninger i den traditionelle spidsoptagelsesteknik.

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

Figur 4A viser spontane pigge, der opstår fra et sensillum. De falder i to klasser baseret på amplitude, hvor de større pigge stammer fra neuronen, der er følsom over for bitre forbindelser, og de mindre pigge fra neuronen, der reagerer på sukkerarter. Forholdet mellem spikeamplitude og funktionel specificitet er blevet bekræftet af genetiske eksperimenter 4,14,37,38,39 .<sup class=...

Read this Scientific Article Protocol about Author Spotlight: Advances in Chemoreception at JoVE.com

En sjældent anvendt metode til elektrofysiologisk optagelse, baseoptagelse, tillader analyse af funktioner i smagskodning, der ikke kan undersøges ved konventionelle registreringsmetoder. Baseoptagelse tillader også analyse af smagsresponser på hydrofobe stimuli, der ikke kan studeres ved hjælp af traditionelle elektrofysiologiske metoder.

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 ...

Vores laboratorium undersøger, hvordan dyr opdager og reagerer på lugt, smag og feromon. Vi har gjort betydelige fremskridt med at forstå det molekylære grundlag for kemomodtagelse. Vi har gjort flere fremskridt på området, herunder opdagelsen af de første insektlugtreceptorer, opdagelsen af de første insektsmagsreceptorer og belysning af grundlæggende principper for logikken i lugt- og smagskodning.Denne teknik tillader måling af testneuronaktivitet før, under og efter stimulus. Det tillader således omfattende analyse af OFF-reaktioner, der opstår efter en smagsstimulus. Det tillader også måling af smagsneuronaktivitet til floden af flygtige forbindelser.Den klassiske teknik til smagselektrofysiologi var ikke i stand til at måle smagsresponser på hydrofobe forbindelser såsom langkædede feromoner, der har meget lav opløselighed i vand. Ved hjælp af denne teknik kan vi imidlertid overvinde denne begrænsning ved at adskille optageelektroden fra glasstimuluskapillæren. Vores forskning undersøger funktionen af de kemosensoriske systemer hos dyr med fokus på deres rolle i fodrings- og parringsvalg.Vi undersøger i øjeblikket ikke-kodende RNA'er og mikropeptider i bananfluers olfaktoriske systemer, og hvordan myg og tsetsefluer bruger disse systemer til at finde menneskelige værter.