Olfactory Male Sensitivity and Its Variation in Response to Fluoranalogs of the Main Pheromone Component of Female Mamestra brassicae A. A. Nikonov, T. V. Tyazhelova, Ye. A. Nesterov, V. M. Rastegayeva, F. E. Ilyasov, P. V. Mashkin and B. G. Kovalyov Institute of Cell Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region, 142292, Russia Z. Naturforsch. 49c, 508-515 (1994); received September 14, 1993/March 14, 1994 Electroantennogramme, Chemical Communication. Fluoranalogs, Mamestra brassicae According to the EAG study and the field trapping tests, fluoranalogs of the main phero­ mone component of Mamestra brassicae females have been found to possess disguising properties for males when used together with the main pheromone component itself. In males, at low concentrations of the above substances the diminishing of attractivity of these blends correlates with the absence of EAG responses to them. An unequal action of these substances has been revealed in different concentration ranges of odours. The mechanisms of the action of these substances are under discussion. Introduction primary mechanisms of olfactory reception, in­ At present, an intensive search for effective bio­ cluding principles and mechanisms of odour infor­ logical ways of protecting plants from insect pests mation recognition; (ii) the changes in sensitivity is going on. One of the promising directions of this and those of distinguishing the odours by insects search is the use of natural pheromones as well as in natural conditions are weakly studied. their analogs, because their use in traps and other In this work we have studied electrical responses preparations has a weak toxicity and results in dis­ of antennae of Mamestra brassicae males to the turbances of sex communication of insects. So, it main pheromone component (Zll-16:OAc) and becomes possible to regulate the number of pests its fluoranalogs, as well as the attractivity of the (Campion, 1984). above substances for Mamestra brassicae males. On the other hand, some substances are known Materials and Methods to suppress the attraction of males of certain insect In the laboratory experiments the males of species to sex pheromones or their synthesized Mamestra brassicae were used for the study. analogs, effectively. These substances may be of Butterflies were bred out of larvae in the diapause the natural origin. Their presence in pheromone at 25 °C and at humidity of 70%. Mamestra bras­ blend appears to be necessary in the course of sicae with developed and intact antennae were interspecies communication of insects (Bogda­ kept at 10 °C after emergence and used 3-5 days nova, 1980; Christensen, 1990). They can be also old. synthesized de novo (Albans, 1984). The search for As pheromone we used a synthesized analog of these substances and their synthesis are rather ex­ the main component (Bestmann, 1978). Haloace- pensive. Investigations in this direction are per­ tate analogs of Mamestra brassicae pheromone formed for a long time (Roelofs, 1968). Here the were synthesized in the laboratory headed by possibility of a specific disturbance of the sexual Boris Kovalyov (Kovalev et al., 1979) (Research chain in some insect species without any marked Institute of Biological Methods of Plants Protec­ change in chemocommunication of the others is tion, Kishenev) as follows: 1) 16-fluor, (Z)-ll- very attractive. Although this problem seems to be hexadecenal- 1 -iltrifluoracetate easy, it has not been satisfactorily solved yet. The main reasons for this are; (i) little evidence on the (F 16,Z 11-16:OCOF3); 2) 16-fluor, (Z)-ll-hexa- decenal-l-ilacetate (F16,Z11 -16:OAc); 3) (Z)-l 1-hexadecenal-1 -iltrifluoracetate Reprint requests to Dr. A. A. Nikonov. (Z 11-16: OCOF 3 ). 0939-5075/94/0700-0508 $ 06.00 © 1994 Verlag der Zeitschrift für Naturforschung. All rights reserved. A. A. Nikonov et al. • Fluoranalogs of the Main Pheromone Component of Female Mamestra brassicae 509 For morphological investigations the antennae of butterflies caught was counted every 3-4 days were cut and fixed in 1.5% solution of glutar alde­ during 2 0 days in the summer of the second gener­ hyde at 4 °C for 2 h. Then after dehydration they ation of Mamestra brassicae (Buchko, 1985). The were embedded into resin “Spur” (Tiras and results obtained were statistically processed and Moshkov, 1978). represented as tables and diagrammes. Electrophysiological measurements were per­ formed at the Institute of Cell Biophysics (Push- Results chino) by using a standard electrophysiological de­ Electrophysiological study vice. According to Schneider’s method (Schneider, 1984), EAG was recorded from the ends of the The use for stimulation of antenna led to ap­ cut-off antenna by capillary electrodes filled with pearance of Z 11-16: OAc negative potential oscil­ an analog of hemolymph solution (Kaissling, lations (Fig. 1 A), the amplitude and the shape of 1986). Odour was pushed into the continuous flux which depended on concentration and duration of of purified air (v = 0.5 1/min) by using the system the stimulus. The first stimulation of the antenna allowing to avoid mechanical artefacts, as it had ( 1 0 “ 1 0 |!g per filter) always evoked a response of been described earlier (Minor and Vasilyeva, a greater amplitude than the subsequent ones used 1979). The exit end of the glass tube (4 mm in at the same or stronger concentrations. The con­ centration area of 1 0 “9 - 1 0 - 7 ^ig per filter was diameter) was at a distance of 1 0 mm off the an­ tenna plane. Paper filter bands (1 cm2) with 10 ^il characterized by small and stable amplitudes. A of the substance diluted in hexane served as a statistically significant increase of the amplitude of source of stimulating vapours. In different experi­ responses (1.05 ± 0.05 mV) was observed at ments the excitement duration was changed from higher concentrations (10 - 6 (ig per filter). A 0.1 to 0.5 sec. The time between excitements was further enhancement of the odour concentration 1-2 min. The parameters of stimuli (sequence fre­ resulted in a periodic increase of the amplitude of quency and push duration) were changed auto­ responses (Fig. 1 B). This dependence was not of a matically. The potentials were recorded and the typical linear character. The curve acquired an S- results obtained were statistically processed shaped form, on which a plateau in the region of (Zacks, 1971) under automatic conditions by using several concentration ranges as well as some de­ an extra developed device and software pro­ crease of the amplitude of EAG responses could grammes based on PC “Iskra-226”. be seen within the range between 1 0 _3 - 1 0 _ 1 ^ig The field trapping experiments were perform­ per filter. ed in 1988-1989 in the fields near Kishenev. The traps were hung about at a height of 1 . 2 m around vineyards and a pea field, placed in a line by blocks (5 traps in each line). The experiment was carried out in two variants: (i) the study of attractivity of used substances No. 1 (F 16,Z 11-16 :O CO F3), No. 2 (F16,Z 11-16: OAc) and No. 3 (Zll-16:OCOF3) and analog of main pheromone component (Z 1 1-16: OAc); (ii) the study of attractivity of used different mixture of the above-mentioned substances. For this purpose two capsules, the one with substance under study, the other with pheromone, were placed into each trap. Sometimes the capsule with substance under log CONCENTRATION ( |ig per filter ) study was placed near the trap filled with phero­ Fig. 1. A. Standard EAG response of Mamestra brassicae mone. The traps with uncharged capsules served males had been recorded after analog-digital converter. as a control. The distances between the traps were B. Dose-response curve for the main pheromone compo­ nent Z 11-16: OAc and fluoranalog of main pheromone as follows: 15 m in the block, 30 m between the component F 16,Z 11-16: OCOF 3 tested on males (n = blocks, 200 m between the variants. The number 10). Mean response ± SEM. 510 A. A. Nikonov et al. • Fluoranalogs of the Main Pheromone Component of Female Mamestra brassicae Using fluoranalog No. 1 for stimulation, we ob­ in the concentration ranges of 1 0 ~ - 1 0 ~ 4 (ig per served an EAG reaction of the antenna of Mame­ filter. In this case the amplitude increase within stra brassicae males similar to the reaction to the 10“3 -1 0 - 2 (.tg per filter observed (Fig. 3). It is main pheromone component. However, the con­ noteworthy that introduction of the main phero­ centration curve of the main component lay above mone component ( 1 0 ~ 7 (ig) into the continuous that of fluoranalog No. 1 (Fig. 1 B). In this case sig­ air flux blowing over the antenna resulted in the nificant differences between them ranged from same dependence of the amplitude of responses IO- 7 to 1 0 ” 3 (tg per filter while the beginning and on the concentration of fluoranalog No. 1 (Fig. 3). the end of the curves did not coincide with each Fluoranalog No. 2 at concentrations of 10“9- other. The activity of fluoranalogs No. 2 and No. 3 1 0 ~ 5 (ig did not make the response to the main showed a different picture. Substance No. 2 caused pheromone component disappear completely, yet two statistically distinguishable amplitudes of re­ the amplitude was small and constant (Fig. 4). sponses, relatively small in size, within concen­ tration ranges of 1 0 ~lo- 1 0 - 5 (tg and 1 0 _4 - 1 0 - 1 (ig per filter (Fig. 2). Fluoranalog No. 3 had one con­ centration region ( 1 0 - 9 (tg) where the antenno- gramme amplitude drastically rose.