The Drosophila ortholog of vertebrate TRPA1 regulates thermotaxis - PubMed (original) (raw)

. 2005 Feb 15;19(4):419-24.

doi: 10.1101/gad.1278205. Epub 2005 Jan 28.

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The Drosophila ortholog of vertebrate TRPA1 regulates thermotaxis

Mark Rosenzweig et al. Genes Dev. 2005.

Abstract

Thermotaxis is important for animal survival, but the molecular identities of temperature sensors controlling this behavior have not been determined. We demonstrate dTRPA1, a heat-activated Transient Receptor Potential (TRP) family ion channel, is essential for thermotaxis in Drosophila. dTrpA1 knockdown eliminates avoidance of elevated temperatures along a thermal gradient. We observe dTRPA1 expression in cells without previously ascribed roles in thermosensation and implicate dTRPA1-expressing neurons in mediating thermotaxis. Our data suggest that thermotaxis relies upon neurons and molecules distinct from those required for high-temperature nociception. We propose dTRPA1 may control thermotaxis by sensing environmental temperature.

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Figures

Figure 1.

Figure 1.

Thermal preference assay. (A) Larvae are placed in the middle of the thermal gradient on an agarose-covered plate, within the release zone. Agar surface temperatures are as indicated. (B) Larval behavior on gradient quantified by an avoidance index (AI) (Liu et al. 2003) as indicated. Graph depicts behavior of uninjected wild-type (elav-Gal4;UAS-mCD8:GFP) late-first instar/early-second instar larvae (n = 9 assays). (C) Still images from thermal preference assay of uninjected wild-type larvae. Red dotted lines demarcate the release zone, with heated zone at left and unheated zone at right. In all figures, data are mean ± SEM. More than 39 larvae/assay.

Figure 2.

Figure 2.

dTrpA1 is required for thermosensory behavior. (A) RNAi strategy (methods). GFP-negative (asterisk), but not GFP-positive (circles), RNAi animals were retained for analysis. (B) Late-first/early-second instar behavior. Uninjected (n = 9 assays); TrpVs:dsRNA, injected with Gal4, Inactive, and Nanchung dsRNAs (Gal4 + Inactive + Nanchung) (n = 4); TrpM:dsRNA:Gal4 + CG30078 (n = 4); TrpAs:dsRNA:Gal4 + dTrpA1 + Painless + CG17142 + CG31284 (n = 6). (C) Preference assay (orientation as in Fig. 1C). (D) As in B except dTrpA1(RNAi)region 1: Gal4 + dTrpA1(region 1) (n = 7). dTrpA1(RNAi)region 2:Gal4 + dTrpA1(region 2) (n = 4); TrpAs-dTrpA1:dsRNA:Gal4 + Painless + CG17142 + CG31284 (n = 4); painless1 (n = 5); painless3 (n = 4); atonal(RNAi) (n = 4). (E) Third instar behavior. Uninjected (n = 9); _dTrpA1(RNAi)_region 1 (n = 13); painless1 (n = 7); painless3 (n = 4); md-Gal4:UAS-TeTxLC (n = 12). (**) p < 0.0001 (versus uninjected). Slight md-Gal4:UAS-TeTxLC effect was not statistically significant (p = 0.09, 2 min; p = 0.27, 5 min) and may reflect their noted uncoordination (Tracey et al. 2003). Nineteen to 75 larvae/assay.

Figure 3.

Figure 3.

dTrpA1(RNAi) animals respond to other stimuli. (A) n-Octyl acetate avoidance of late-first/early-second instar elav-Gal4;UAS-mCD8:GFP larvae, uninjected, or dTrpA1(RNAi) (region 1) (n = 6 assays for each). Odor avoidance = [(number of larvae in no odorant zone) – (number of larvae in odorant zone)]/[(number of larvae in no odorant zone) + (number of larvae in odorant zone)]. (B) Response to contact with ∼55°C probe of third instar uninjected and dTrpA1(RNAi) larvae. (C) Time between probe contact and initiation of curling. Uninjected (n = 41 larvae); _dTrpA1(RNAi)_region 1 (n = 24); md-Gal4 (n = 23); UAS-TeTxLC (n = 30); md-Gal4:UAS-TeTxLC (n = 23). Most larvae not responding within 3 sec never exhibited detectable curling.

Figure 4.

Figure 4.

dTRPA1 expression. (A) dTRPA1 expression in anterior (arrowhead) and posterior (bracket) groups of neurons within the brain and in neuroendocrine cells of corpus cardiacum (asterisk). (B) Posterior group of dTRPA1-expressing neurons. (C) dTrpA1(RNAi) animals lack dTRPA1 staining. Asterisk denotes corpus cardiacum. (D_–_F) dTrpA-Gal4:UAS-mCD8:GFP (green) expression. dTrpA1-Gal4 drives GFP expression in dTRPA1-expressing (purple) neurons and 100–150 additional cells within the brain. (E) GFP. (F) Anti-dTRPA1. (G) Thermal preference behavior. Wild type (n = 9 assays); UAS-TeTxLC (n = 8); dTrpA1-Gal4(n = 9); dTrpA1-Gal4:UAS-InactiveTeTxLC (n = 6); UAS-Hid (n = 6); dTrpA1-Gal4:UAS-TeTxLC (n = 10); dTrpA1-Gal4:UAS-Hid (n = 10). (*) p < 0.005. (H) Response to ∼55°C probe contact. dTrpA1-Gal4:UAS-TeTxLC (n = 19); dTrpA1-Gal4:UAS-Hid (n = 20).

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