The molecular basis of CO2 reception in Drosophila

Abstract

CO(2) elicits a response from many insects, including mosquito vectors of diseases such as malaria and yellow fever, but the molecular basis of CO(2) detection is unknown in insects or other higher eukaryotes. Here we show that Gr21a and Gr63a, members of a large family of Drosophila seven-transmembrane-domain chemoreceptor genes, are coexpressed in chemosensory neurons of both the larva and the adult. The two genes confer CO(2) response when coexpressed in an in vivo expression system, the "empty neuron system." The response is highly specific for CO(2) and dependent on CO(2) concentration. The response shows an equivalent dependence on the dose of Gr21a and Gr63a. None of 39 other chemosensory receptors confers a comparable response to CO(2). The identification of these receptors may now allow the identification of agents that block or activate them. Such agents could affect the responses of insect pests to the humans they seek.

Fig. 1.

Gr21a and Gr63a are coexpressed in larval and adult Drosophila sensory neurons. (A–C) Expression patterns driven by Gr21a-GAL4 (A), Gr63a-GAL4 (B), and Gr21a-GAL4+Gr63a-GAL4 (C) in larvae were visualized by using a UAS-GFP reporter. Pictures show only one side of the larval head region, and a single labeled neuron is visible in each case. (D) The ab1 sensillum, which houses four sensory neurons as indicated (A, B, C, and D). In the adult fly, Gr21a (E) and Gr63a (F) promoters drive expression of the GFP reporter in neurons that project to the V glomerulus (green) of the antennal lobe. Brain neuropil is labeled with antibody nc82 (magenta). One antennal lobe is shown in each case.

Fig. 2.

Gr21a and Gr63a receptors together confer an electrophysiological response to CO₂ in the mutant ab3A neuron. (A) Schematic illustrating the empty neuron system used to test electrophysiological responses of receptor combinations in the mutant ab3A neuron. (B) Representative traces showing the electrophysiological responses of ab3 ORNs. Flies contain Or22a promoter-GAL4 and UAS-Gr transgenes encoding the indicated receptors. Spikes of two amplitudes can be distinguished; the large spikes represent the activity of the ab3A neuron and are counted; the small spikes represent the activity of the neighboring ab3B neuron (16). Bars indicate a 0.5-sec stimulus period of 100% CO₂. (C) Mean responses to 100% CO₂ of the mutant ab3A neurons expressing the indicated receptors. Error bars indicate SEM; n = 8–20.

Fig. 3.

CO₂ response of Gr21a and Gr63a is dose-dependent. Error bars indicate SEM and are too small to be seen in some cases; n = 8–9.

Fig. 4.

Gr21a and Gr63a response is narrowly tuned to CO₂. (A) Responses of the empty neuron expressing either Gr21a+Gr63a+Gr63a (Left), Gr21a+Gr21a+Gr63a (center), and of the ab1C neuron, which was analyzed in Or83b² flies (right). 100% CO₂ was used to test the responses of Gr21a and Gr63a combinations in the empty neuron system (Left and Center), and 1% was used to test responses of the ab1C neuron. Error bars indicate SEM; n = 8–10. (B and C) CO₂ responses of Gr21a+Gr21a+Gr63a and of 39 odorant receptors (refs. and ; S. A. Kreher and J.R.C., data not shown) in the mutant ab3A neuron (B). (C) Responses after subtraction from values in (B) of control responses of the 39 odorant receptors to paraffin oil or water. All odorant receptors were stimulated with a 500-ms puff of 100% CO₂ as described (17); Gr21a+Gr21a+Gr63a was stimulated for 500 ms with a stream of 100% CO₂ as described in Materials and Methods. Some receptors were tested with both delivery methods, which gave identical results. All receptors are arranged along the abscissa according to the strengths of their responses. Receptors with the highest responses are placed near the center of the distribution; those that have the lowest responses are placed near the edges. The order of receptors is thus different for B and C and is available in the supporting information (SI) Text.