Labeled lines. Drosophila avoid ingesting alkaloids. Bitter modulated sugar detection in Drosophila melanogaster

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1 2/2/213 Drosophila avoid ingesting alkaloids Bitter modulated sugar detection in Drosophila melanogaster nl/fly/h Females Males Frédéric MARION POLL AgroParisTech, Paris INRA UMR 1272 PISC multicafe 1 Concentrations (+ 35 mm Fru): /.1 /.1 /.1 / 1 / 1 mm Sellier et al. (211) Chem. Senses 2 Drosophila «bitter» cells respond to alkaloids Labeled lines Strychnine I9 i type sensillum.1 mm 1 mm 1 mm Tip recording 1 ms 2 mv Meunier et al. (23) 3 4 1

2 2/2/213 Sugar + quinine Quinine inhibitssugardetection Consumption / fly / h (nl) MultiCAFE Quinine (mm) + 35 mm Fructose N spikes Tip- recording Quinine (mm) + 35 mm Fructose Sellier et al. (211) Chem. Senses Sellier et al. (211) Chem. Senses 5 6 Sugar sensing inhibition Many Lepidoptera larvae ( or inositol) Coleoptera, Diptera, Honeybees Humans & Vertebrates (TRPM5) + quinine + quinine Electrophysiological recordings from labellar sensilla of Calliphora vomitoria. (A and C) 6 mm ; (B) suc mm quinine HCI; (D) suc + 6 mm quinine HCI From Morita, H., J. Cell. Comp. Physiol., 54, 189, Sensilla Proboscis 8 2

3 2/2/213 Peripheral interactions? Hypotheses: Lateral interaction Mixture suppression Use DTI toxin to selectively kill Gr66a cells Expected: Electrophy Behavior No more inhibition Inhibition 9 1 Responses to sugar + strychnine Responses to sugar + L canavanine 6 Consumption / fly / hour (nl) MANOVA, Profile analysis, p <.5,1 1 1,1 1 1,1 1 1 Gr66a Gal4 UAS DTI Gr66a DTI Consumption / fly / hour (nl) Gr66a Gal4 UAS DTI Gr66a DTI 1 1 Strychnine (mm) + 1 mm Sucrose.1 M with L canavanine (mm) Sellier et al. (211) in prep. Sellier et al. (211) in prep

4 2/2/213 Sugar responses are inhibited by strychnine Partial conclusions 75 All bitter compounds are not inhibiting sugarsensitive cells 5 spikes/s 25 S6 (UAS DTI) S6 (Gr66a DTI) Parental line Strychnine can be detected behaviorally without a bitter cell and it inhibits sugar detection.1m.1m 1M.1mM.1mM 1mM.1mM.1mM 1mM Sucrose Strychnine Mixture Sellier et al. (211) in prep. L canavanine cannot be detected behaviorally without a bitter cell and it does not inhibit sugar detection Proboscis extension/retraction Strychnine EXPERIMENT Extension Touch leg and monitor proboscis during5 s no Ext Retraction Sucrose + strychnine 1 mm strychnine Mixed (+) Dissociated (//).1 M Sugar sensing AND bitter sensing cells are exposed to the mixture Sugar sensing cells are stimulated by Bitter sensing cells are stimulated by strychnine p <.5 p <.1 p <.1 Red Pearson s Chi2 test Black McNemar Chi2 with continuity correction + : mixed // : dissociated Sellier et al. (211) in prep

5 2/2/213 Strychnine L canavanine + : mixed // : dissociated + : mixed // : dissociated p <.5 p <.1 p <.1 Red Pearson s Chi2 test Black McNemar Chi2 with continuity correction Sellier et al. (211) in prep. p <.5 p <.1 p <.1 Red Pearson s Chi2 test Black McNemar Chi2 with continuity correction Sellier et al. (211) in prep L canavanine Conclusions (2) + : mixed // : dissociated Sugar sensing inhibition plays a major role in preventing proboscis EXTENSION Bitter sensing plays a major role in proboscis RETRACTION p <.5 p <.1 p <.1 Red Black Pearson s Chi2 test McNemar Chi2 with continuity correction Sellier et al. (211) in prep

6 2/2/213 Acknowledgements Speculations/questions: Molecular mechanisms? Role of bitter sensing cells? Ancestral mechanism? Co workers: Marie Jeanne Sellier (PhD) Alexandra Guigue (PhD) Moutaz Ali Agha (M2R/PhD) Marie Ange Chabaud (Postdoc) Nicolas Meunier (PhD) Makoto Hiroi (PhD) Alice French (PhD) Philippe Touton (INRA) Support: ANR INSAVEL, ADAPTANTHROP MNESER, DGER Institutions: AgroParisTech, INRA, UPMC Collaboration: Yves Grau, CNRS, Montpellier Pablo Reeb, Argentina Fly strains: Kristin Scott, UC Berkeley John Carlson, Yale Craig Montell, Baltimore Teiichi Tanimura, Kyushu University Taste neurons co express specific receptors Bitter cells = potential target / insects ppk28 ppk11, ppk19 ppk23 (pheromones) Gr5a Gr61a, Gr64a,,f Salt Water Sugar Bitter Gr66a Gr33a, Gr93a Gr8a Gr22b Gr22e Gr22f Gr28b Gr28be Gr32a Gr32b Gr47a Gr59a Gr59b Gr93a DmX painless Postingestive mechanisms Detoxification Elimination = P45, etc Pre-ingestive mechanisms Avoidance = Bitter taste

7 2/2/213 Avoidance vs toxicity? 1959 Antifeedancy + _ Compound EC5 (mm) Strychnine.5 Lobeline.11 Berberine.6 Theophylline.4 Quinine.5 Caffeine 1.1 Papaverine 3 Nicotine 4 Total consumption / fly / hour (nl) a a a, b a, b a, b a, b b, c No compensative feeding b, c c Bonferroni, p < Taste in Drosophila Each sensilla is different Adult: 66 taste neurons Legs, mouthparts, wings, ovipositor CNS =1 «taste brain» / segment 1 sensillum = 4 neurons (+ 1) S: sugars L2: bitter L1: salt W: water mm -> 2 spk/s quininetype berberine -type in L2 cell 5b/4s 5s Berberine no.5 Quinine.1 no Denatonium.5.5 Strychnine 3 3 Caffeine no 13 Aristolochic acid no no Salicin no no From Hiroi et al., Zool J

8 2/2/213 Taste receptors Co receptors? Gr66a, Gr33a, Gr93a 6 putative taste receptors 68 proteins Large divergence (about 8% homology) Structurally close from Ors van Naters & Carlson (26) Nature Clyne, P. J., C. G. Warr, et al. (2) "Candidate taste receptors in Drosophila." Science 287: Lee et al. (29) PNAS Moon et al. (29) Curr. Biol Individual components mixture Use of UAS/Gal4 to express DTI in Gr66a (bitter) cells.1 M Sucrose Gr66a Gal4 UAS DTI 1 mm Strychnine.1 M Sucrose + 1 mm Strychnine Gr66a Gal4 Diphtheria toxin = cell death Moutaz ALI AGHA (21) unpubl. Wang et al., Cell, 24 Brand and Perrimon, Development, 1993 F1 UAS GAL4 DTI

9 2/2/213 Where do we go from there? Sugar responses are inhibited by strychnine Expand observations on other bitter substances Improve stimulus/recording methods Find molecular mechanisms involved S6 (Gr66a Gal4) S6 (UAS DTI) S6 (Gr66a DTI).1M.1M 1M.1mM.1mM 1mM.1mM.1mM 1mM spikes/s Parental lines Sucrose Strychnine Mixture Sellier et al. (211) in prep Sugar responses are not inhibited by L canavanine L5 S spikes/s KCl L cana Suc Suc + L cana KCl L cana Suc Suc + L cana NS NS NS Schoonhoven, L. M., W. M. Blaney, et al. (1992). Sensory coding of feeding deterrents in phytophagous insects. Insect Plant Interactions: volume IV. E. A. Bernays. Boca Raton, USA, CRC Press Inc.: Sellier et al. (211) in prep