Sensory electrophysiological recording techniques such as EAG (electroantennogram), GCEAD (coupled gas chromatograph-electroantennogram detection) and SSR (single sensillum recording) have been useful in the chemical ecology studies of insects and plants. Numerous pheromones and other semiochemicals have been identified through GCEAD analysis, and the response profiles of antennae and individual olfactory receptor neurons have been characterized by using EAG and SSR techniques. In this talk, the practical aspects of these techniques are presented in detail. Standard setup and procedure of each electrophysiological recording technique, and important parameters and proper data analysis methods will be introduced as well as its applications. Common mistakes and limitations of these techniques will also be discussed.

Phytophagous insects detect volatile compounds produced by host and non-host plants, using species-specific sets of olfactory receptor neurons (ORNs). To investigate the relationship between the range of host plants and the profile of ORNs, single cell recordings were carried out to identify ORNs and corresponding active compounds in female Uraba lugens (Lepidoptera: Nolidae), an oligophagous eucalypt feeder. Based on the response profiles to 39 plant volatile compounds, 13 classes of sensilla containing 40 classes of ORNs were identified in female U. lugens. More than 95% (163 out of 171) of these sensilla contained 16 classes of ORNs with narrow response spectra, and 62.6% (107 out of 171) 18 classes of ORNs with broad response spectra. Among the specialized ORNs, seven classes of ORNs exhibited high specificity to 1,8-cineole, (±)-citronellal, myrcene, (±)-linalool and (E)-β-caryophyllene, major volatiles produced by eucalypts, while nine other classes of ORNs showed highly specialized responses to green leaf volatiles, germacrene D, (E)-β)-farnesene and geranyl acetate that are not produced by most eucalypts. We hypothesize that female U. lugens can recognize their host plants by detecting key host volatile compounds, using a set of ORNs tuned to host volatiles, and discriminate them from non-host plants using another set of ORNs specialized for non-host volatiles. The ORNs with broad response spectra may enhance the discrimination between host and non-host plants by adding moderately selective sensitivity. Based on our finding, it is suggested that phytophagous insects use the combinational input from both host-specific and non-host specific ORNs for locating their host plants, and the electrophysiological characterization of ORN profiles can be used for predicting the range of host plants in phytophagous insects.

The effect of CO2 on the opening of stomata in the intact leaf of Commelina communis has been investigated. Full opening of stomatal apertures(around 18 μm) was achieved in the intact leaf by addition of CO2(900 μmol mol-1). At 90 minutes, the stomatal apertures of leaves treated with CO2 free air were reduced. In contrast, stomata opened most widely with the treatment of CO2 air at 90 minutes. The effects of light, CO2 air and CO2 free air on the change of membrane potential difference(PD) were measured. Fast hyperpolarization of guard cell membrane PD was recorded reaching up to -12 mV in response to light. If CO2 free air was given firstly, there was no response. When light was given after CO2 free air, the light effect was very clear. At the onset of CO2 air, the PD showed a dramatic hyperpolarization to about -25 mV. Changes in the pH of apoplast in intact leaves in response to CO2 air were observed. CO2 air caused a change of 0.4 pH unit. Therefore, it can be hypothesized that CO2 flowing could stimulate proton efflux which is a necessary precursor of stomatal opening.