Behavioural responses of fifth instar H. angustipennis larvae were recorded using the impedance conversion technique in exactly the same experimental set-up as Heinis and Swain (1986). Prior to the measurements, larvae were placed individually in sealed nylon meshed tubes (7 cm length, 1.5 cm diameter) and a 24-hour acclimation period in Dutch Standard Water (DSW) was provided. DSW is a standardized synthetic analogue of common Dutch surface waters, containing 200 mg CaCl2 .2H2 0, 180 mg MgSO4.7H2 0, 100 mg NaHCO3 and 20 mg KHCO3 per L demineralized water (pH ~8.l, hardness 210 mg/L CaCO3 , alkalinity ~1.2 meq/L). During the acclimation period, Urtica powder was added as food. After the acclimation period, in which the larvae were able to construct their nets within the nylon tubes, they were divided over three different 5 L aquaria and exposed for 48-hour to the three different oxygen concentrations: 100%, 50% and 30% air saturation (ca. 9.2, 4.6 and 2.8 mg/L respectively). The oxygen concentration in the water in the aquarium was kept at a constant level by an oxystat that compared the actual oxygen concentration, measured by an oxygen electrode (Yellow Springs Instruments, Yellow Springs, OH, USA), with the preset concentration. When the measured oxygen concentration was too high, an electrical gas valve was automatically opened to pass nitrogen into the water. When the measured oxygen concentration was too low, air was passed into the water and when the measured oxygen concentration was the same as the preset concentration, both valves were closed. The aquarium was covered with a glass plate and nitrogen and air were brought into the water by means of air stones. During the exposure time, no food was present, a 16:8-hour light:dark regime was provided and the temperature was maintained at 20°C.
For each concentration, behavioural patterns of twenty fifth instar larvae were recorded. At the end of the 48-hour exposure period, activity patterns of all larvae were recorded during 1-hour by placing the nylon tubes with the larvae in a measuring chamber (7 × 3 × 2.5 cm) which received re-circulating water from the exposure aquarium at a flow rate of 5ml/min. After 5 minutes of acclimation, changes in the impedance of the system caused by the movement of the larvae were detected by two stainless steel electrodes placed at either side of the measuring chamber connected to the impedance converter and assimilated with 53 ms-seconds time intervals with the computer program Aqualand (Augustijn Onderzoek) on a MSDOS 486 micro-computer. From the activity signals, three different types of behaviour were defined according to the relative frequencies and amplitudes: undulatory movements or ventilation (mono-frequent with a relative high and constant amplitude), inactivity (signals below background noise) and other activity (multi-frequent with different amplitudes). Examples of the different types of behaviour as recorded with the impedance conversion technique are given in figure 1.