Phys~ologzcalEntomology (1985) 10, 257-265

A pulsed cloud of sex elicits upwind flight in male

T C BAKER, M A WILLIS, K F HAYNES and P L PHELAN Division of Toxicology and Physiology, Department of Entomology, University of California, Riverside, California

ABSTRACT Male oriental fruit moths do not fly upwind in a continuous uniform cloud of pheromone, but readily do so when the cloud is pulsed at 1 or 0 51s or when a plume from a point source of pheromone is placed within the continuous cloud It is suggested that males of species that require such fluctuating pheromone stimula- tion for upwind flight will normally receive it from a filamentous, point-source-produced plume However, we hypothesize that upwind progress may cease close to the source due to excessively high emission rates or inappropriate blend ratios, when fluctuating sensory output becomes attenuated, despite higher actual molecular concentration fluctuations

Key words. Sex pheromone, Graphohta molesta, Lepidopter a, Tortrici- dae, orientation, upwind flight, intermittent stimulation, odour percep- tion

Introduction Willis & Baker, 1984) Rather, experiments have strongly suggested (Kennedy et a1 , 1980, Hypotheses about how the sex pheromone 1981; Willis & Baker, 1984) that upwind flight blend ratios and emission rates of is elicited by significant amplitudes and moths attract males have centred around the frequencies of peak-to-trough fluctuations concept of a behaviourally 'active space' (Bos- (Kennedy, 1983) in pheromone concentra- sert & Wilson, 1963), in which a time-averaged tion created by a plume's filamentous structure concentration of pheromone is above the re- (Wright, 1958) < sponse threshold for upwind flight Recently We demonstrate here, more directly than in there has been increasing evidence, however, earlier studies, that fluctuating, discontinuous, that for some species the continuous presence pheromone stimulation produces upwind flight of pheromone, regardless of average concen- in male moths In addition, we suggest that tration, does not result in such behaviour such flight, having begun far downwind under (Kennedy et al, 1980, 1981; Kennedy, 1982; intermittent conditions characteristic of phero- mone plumes, may cease close to the source Corres~ondence: Baker, Division of when, as a result of an excessively high con- Toxicology and Physiology, Department of En- tomology, University of California, Riverside, CA centration 01 incorrect the fluc- 92521, U S A tuating signal becomes too uniform 257 258 T C Baker tt

Materials and Methods on average A point-source plume of phero- mone was created using a rubber septum Male oriental fruit moths, Grapholzta molesta impregnated with 30pg of the same blend used (Busck), were released three at a time from a for clouds The septum was suspended with 6 5 x8cm (height diameter) screen cone, nylon filament 20cm above the floor, 10cm 30cm from the downwind end of a previously from the tunnel's upwind end described 1 8x0 6x0 6 m wind tunnel (Willis Our ability to produce a continuous uniform & Baker, 1984) Wind velocity was 0 5 mls, cloud of pheromone or a 'pulsed' cloud inter- created by means of a 115 h p rotary-blade rupted by periods of cleaner air was verified blower (Dayton Coip ) All air in the tunnel visually using TiC14 smoke emanating from was exhausted continuously via a 30cm dia- dispensers in the pulse-generating device, by meter duct connecting a fume hood to a 0 6m infra-red photocell recordings of the smoke long sheet-metal extension which sealed the pattern (Fig lA), and by recording electro- downwind end of the tunnel's plexiglas work- antennogram (EAG) responses to pulsed and ing section Air pressure was equalized by continuous pheromone clouds (Fig 1A) using means of a valve in the exhaust connection so the same concentrations as in the behavioural that when moths were introduced via a side experiments To the eye, the continuous cloud . door in the sheet-metal section, no perceptible of smoke from the pulse-generator appeared as disruption of the continuous or pulsed clouds a uniformly grey fog, and the pulses of smoke occurred, as visualized by smoke during intermittent clouds appeared as more or , The fine-mesh brass screening and metal less rectangular swaths of c 0 4m length . strips used previously for uniform cloud forma- travelling down the tunnel, sandwiched be- tion (Willis & Baker, 1984) were left in place, tween swaths of nearly-clean air but instead of rows of pheromone-impregnated The photometer was placed on a circular rubber septa placed downwind of the strips, metal ring (12 cm diameter) opposite an infra- pheromone was introduced into the blower via red light-emitting diode The ring was posi- an airstream blowing over septa housed in a tioned perpendicularly to the wind, at the 27 x 3 5 cm (length x diameter) glass tube Each same position and height as the males' take-off septum was impregnated with lOOOpg of the point Smoke interrupting the light beam was optimum G molesta three-component phero- registered as a decrease in light intensity, and mone blend: 6% (E)-8-dodecenyl acetate and this fluctuation was amplified and recorded on 4% (2)-8-dodecenyl alcohol (Card6 et al, a Gould Brush 220 strip chart recorder EAG 1979) in (Z)-8-dodecenyl acetate (Roelofs et preparations were made using intact males al, 1969) The septa were placed on a rack (Kuenen & Baker, 1981) and were recorded at 8cm from the mouth of the tube and arranged the position and height of the males' take-off so that they formed a disc 3 cm thick across the point The same pheromone septa and concen- tube, whose mouth was 6cm from the blower trations used during behavioural recordings intake A vacuum was drawn continuously were used for EAGs Responses were re- from the base of the tube to prevent phero- corded on a Hewlett-Packard 3390A integrator mone from being emitted Either intermittent used as a strip-chart recorder '. or continuous clouds were generated by open- All experiments were conducted 0-2h be- ing a valve which released pressurized air into fore lights-off; the optimal time of sexual the base of the tube The vacuum was thus activity for this species (Baker & Cardk, 1979), overcome, creating an airstream blowing at 880 lux fluorescent and incandescent lighting rapidly over the septa, thereby injecting intensity Several seconds after the pheromone pheromone-laden air into the blower Only regime had begun, the three males were clean air entered the tunnel between emis- placed, point of cone down, in a ring-stand sions The tempo of pulsing, either 1 or 0 5/s, (8 4cm diameter) located 20cm above the was performed by manually opening and clos- floor in the centre of the tunnel, 30cm from ing the pressurized air valve in time to an the downwind end Distance of up- or down- electronic tone beeping at 11s The duration of tunnel flight was scored as the first point at each burst of air through the tube was kept as which males touched the tunnel's walls, where- short as possible with this mechanism, 0 25 s upon observations were terminated Flight Pulsed sex pheromone cloud evokes upwind flight 259

tracks of these males were also video-recorded pheromone loss (Marsh et a1 , 1978; Kuenen & and analysed using video-recording equipment Baker, 1983), but these counterturns were and computer analyses of digitized tracks de- performed as symmetrically across the wind- scribed previously (Kuenen & Baker, 1982; line as the other males' upwind zigzags, despite Willis & Baker, 1984) The video-camera's their larger magnitude (Fig ID) In contrast, field of view encompassed the entire width of the few males that took flight when released the tunnel and from 30 cm downwind to 70 cm into clean air displaced predominantly down- upwind of the release cone wind with very few across-wind counterturns These were performed significantly less sym- metrically than those observed in response to Results any of the pheromone treatments (Fig ID) The flight tracks of males flying upwind in A significantly greater percentage of males the pulsed clouds did not appear to be as took flight in the point-source plume (88%l:', regular in zigzagging tempo or width as those n=60) and in the clouds of pheromone pulsed of males flying in the plume (Fig 1B) Tracks at 0 51s ('slow') n=59) and 11s ('fast') of males flying at least 25cm upwind in the * T (92%ab, n=60) than when the cloud was con- pulsed clouds exhibited zigzags approximately tinuously presented to males (67%', n=60) or twice as wide, on average, as those of males in clean air (45%*, n=60) (percentages having flying upwind in the plume (10 4k6 Ocm SD, L no letters in common are significantly different n=28 males, fast-pulsed; 12 0k8 8cm SD, 4 according to the Ryan (1960) test; P<0 05) n=29, slow-pulsed; 5 3k1 4cm SD, n=25, Additionally, a greater percentage of the males plume) This difference may have been due, in that took flight flew upwind in the pulsed part, to our inability to generate concentration clouds (56%a, fast; 63%a slow; n=55 and 59, fluctuations in our pulsed clouds that were as respectively) than in the continuous one rapid or as sharply rising and falling as those in (lo%", n=40), and their distances of upwind the plume, as indicated by smoke and EAG progress were significantly greater than those (Fig 1A) Previous work (Kuenen & Baker, of males in the continuous cloud The latter 1982) established that plumes from Ipg males did not fly significantly farther upwind sources elicited zigzags in G molesta males than those flying in the absence of pheromone approximately twice the width of those per- ('clean air') (Fig IE) formed in plumes from sources loaded with 10 Analysis of video-recorded flight tracks re- or 30% Thus our pulsed clouds may have vealed that the males flying in the pulsed been mimicking the types of fluctuating sti- clouds counterturned across the windline with mulation found in weaker point-source inter-reversal track angles (Marsh et a1 , 1978; plumes Alternatively, the more shallowly and Kuenen & Baker, 1983) and across-wind coun- less sharply fluctuating stimulation might be * terturn symmetries not significantly different similar to that found at greater distances from males flying to the point-source plume (10-15 m) downwind from a stronger point- (Figs 1C and ID, respectively) The latter source (Murlis & Jones, 1981) The tracks of values were measured by taking successive x moths flying at great distances from a phero- * pairs of inter-reversal track angles from each mone source have never been analysed in flight track and calculating for each pair the detail, although some published tracks of male deviation of the second angle from a perfectly gypsy moths (David et al, 1983) suggest a symmetrically performed track angle; one that wider variety of zigzagging widths under such matches the first but on the other side of the conditions than in a wind tunnel (Card6 & windline We view the symmetry of successive Hagaman, 1979), perhaps similar to the flight across-wind inter-reversal track angles as re- variation seen in our pulsed clouds flecting the degree of integration of the self- Male G rnolesta's lack of upwind progress in steered counterturning programme with the the continuous cloud of pheromone was not polarizing effect of optomotor anemotaxis due to an excessively high concentration that On the other hand, males flying in the was then diluted by pulsing because, in a continuous cloud exhibited wide, 90' cross- second experiment, a significantly greater per- wind reversals characteristic of casting after centage of males that took flight progressed 260 T C Baker et a1

4ova CLEAN AIR

0 40 80 120 160 DEGREES FIG. 1. (A) Measurements of stimulus patterns at point of take-off using recordings of TiC14 smoke made with a photocell ('smoke'), and of pheromone using an EAG preparation ('EAG'). Vertical and horizontal bars in slow-pulsed figure denote for all figures 1mV EAG response and 2s respectively. Smoke and pheromone used in creating continuous cloud for illustration were turned off after 10 s. (B) Representative flight tracks of males video-recorded from above flying in response to different pheromone stimulation regimes. Wind was from the top in each figure, and open circles at beginning of each track denote the take-off point, which has been shifted to various locations in the figure only for the purpose of illustrating several tracks at once. Dots along tracks denote 1130s intervals. (C) Distributions and grand means of mean inter-reversal track angles of all males that took flight to different pheromone regimes and completed at least one turn (n (males) and SD, from top, are 36,16.3; 44,20.7; 45.23.6; 33,25.5; 17.40.3). (D) Distributions and grand means of across-wind counterturn symmetries of all males flying in different pheromone regimes that completed at least two turns (n (males) and SD, from top, are 36,32.1; 44,23.6; 45,16.7; 30, 18.1; 14,27.9). (E) Mean upwind flight distances LSD of all males that took flight in response to different regimes. In (C), (D) and (E), means in each column having no letters in common are significantly different according to Duncan's multiple range test (P<0.05), 262 T C Baker el a1

upwind when a plume from a 30pg source was moth species known to rhythmically extrude placed within the cloud (42%, n=48; 50%, their pheromone glands (Card6 & Roelofs, n=48 for plume in clean air) than when this 1973; Card6 et a1 , 1984; Conner et a1 , 1980) plume was absent (17%, n=42; P0 05), and significantly already provide sufficiently intermittent sti- farther than in the cloud alone mulation to elicit upwind flight A further (-14 5k15 8cm SD, n=42; P<0 05, Duncan's enhancement in perceived fluctuation and in Multiple Range Test (Steel & Torrie, 1960)) attraction may be possible due to the pulsing of glands, but this has not yet been demons- trated behaviourally Should an increase in Discussion attraction eventually be demonstrated in such species, we think it would not likely be due to There are three species of moths for which it is a coded, species-specific pulsing pattern, but known that continuous stimulation with uni- rather to an enhancement of intermittencv form pheromone clouds does not evoke sus- There is recent evidence that a major reason tained upwind flight: Adoxophyes orana (Ken- for gland pulsation in arctiids may be to nedy et a1 , 1980, 1981), G molesta (Willis & increase the emission rate of pheromone from Baker, 1984; this paper), and Lymantria dcspar within the invaginated tubes of these complex (C T David, personal communication) In glands (Schal & Card6, 1985) both A orana and G molesta the onset of a Upwind flight in a plume can cease befoie cloud elicits a brief upwind surge, but the the source is reached, becoming instead in- continuous stimulation subsequently fails to flight station-keeping (arrestment) if too much sustain it Kennedy et a1 (1980, 1981) and pheromone is emitted or if the blend ratio is Kennedy (1983) hypothesized that it is the ups slightly wrong (Baker & Roelofs, 1981; Baker and downs in concentration from a plume's et al, 1981; Linn & Roelofs, 1983) We filaments that cause sustained upwind flight, suggest that the neuronal cause of arrestment and showed that a plume placed in a uniform in such cases may be a relative fusion of the cloud readily evoked such flight when the 'flickering' receptor output generated by the cloud had failed to do so Willis & Bake1 (1984) plume's filaments When arrestment occurs showed that G molesta males flew upwind close to an excessively emitting source, the along either vertical or horizontal edges of a receptors may be unable to continue to reflect clean-air-pheromone cloud interface, but not the actual peak filament concentrations as more than briefly after being engulfed by a they increase past the receptors' capacities cloud They hypothesized that such progress (Baker, 1985) but, also, the surplus molecules was due to the intermittencv of stimulation will take longer to locate unoccupied receptor that males received by their movement in and sites and be cleared by enzymatic degradation out of the cloud, plus a slight inhomogeneity of (Kaissling, 1971) The receptors, therefore, concentration at the edge due to turbulent might fire more continuously at significant eddies Now we have shown more directly than rates even during intervals when the moth is in previous studies that it is the fluctuations actually in a pocket of clean air or 'outside' the alone, apart from any other possible spatial plume during a reversal The moth, earlier gradient cues, that sustain upwind flight The kept in the immediate vicinity of the plume by same cloud that previously failed to cause its programme of narrow counterturns to signi- upwind flight now, when pulsed, did so ficantly flickering stimulation farther down- Despite the fact that we have demonstrated wind, now reaches a zone where the in- directly that pulsed pheromone concentration creasingly narrow counterturns trap it even is necessary to sustain upwind flight in moths, more and prevent excursions into clean air we do not propose that of the several from lasting long enough to reduce fusion of Pulsed sex pheromone cloud evokes upwind flight 263 the neuronal signal Wrong blend ratios at for describing such stimulation foi many moths normal emmission rates may also result in (Kennedy, 1983) However, for some species relatively fused signals In Argyrotaenza veluti- such as G molesta which are arrested by high nana, the slower time course required for concentrations, there may be upper limits to clearing of one component, (£)-11-14:A abilities to process ever higher and sharper (Roelofs & Comeau, 1971; Baker & Roelofs, rises and falls, with the possibility that at some 1976), suggests that too much of this compo- point before the source is reached the neuronal nent would be more likely to 'fill in', with fluctuations become too attenuated to sustain neuronal firing, the troughs between peak upwind flight Consequently, the neuronal stimulation by pheromone filaments Lower thresholds of flickering and fusion may both frequencies of source location are observed in play a significant part in defining the upwind response to such high (E): (Z) ratios flight active space for a particular blend and Recently, interneurons that respond phasi- emission rate cally to changes in pheromone concentration In light of these considerations, the plume's have been discovered in physical structure as created by the wind speed (Olberg, 1983) The interneurons exhibit two and large- and small-scale turbulence, among distinct firing frequencies, 'high' and 'low', and other factors, would all play a large role in the neurons alternate between these states defining the active space (Murlis & Jones, upon repeated presentations of pheromone 1981) However, measuring molecular concen- Usually increases in concentration, but some- tration changes using a perfect detector that times also decreases, trigger changes in state responds virtually instantaneously to flux If such interneurons were present in G molesta changes, and registers the actual peaks and they would require fluctuating pheromone sti- troughs in pheromone concentration, will not mulation to change states, and the frequency give the entire picture of the active space of state changes may be what modulates the What will be important, as it is to a responding counterturning programme (Willis & Baker , male in nature, are the fluctuations as reg- 1984) istered by neuronal elements Moth olfactory Such 'flip-flopping' interneurons (Olberg, neurons thus far do not seem to be perfect 1983) will not change states if the receptors detectors Their limitations to registering modulating them are not themselves providing actual fluctuations of concentiation would significantly fluctuating output In our study, most likely start at the receptor level, including EAGs in the three pheromone regimes that receptor capacities and lag times in processing elicited upwind flight (the plume and slow- and and clearing pheromone molecules (Kaissling, fast-pulsed clouds) clearly changed amplitude 1971) The properties of these imperfect detec- In contrast, EAGs did not fluctuate noticeably tors will determine the behaviourally relevant under constant stimulation by the continuous amount of pheromone fluctuation and help cloud, in which upwind flight was not determine the active space for upwind flight observed, although they did remain constantly Apart from the practical implications of high, indicating that the receptors were not these findings and those of Kennedy et a1 adapted This lends support to the idea that it (1980, 1981) for disruption strategies is not the overall magnitude of receptor output, that would employ a diffuse cloud of pher- but the magnitude of receptor output fluctua- omone instead of multiple discrete point- tions which is important for evoking upwind source plumes, they raise questions concerning flight odour perception in all animals For instance, For moths requiring fluctuating pheromone 'sniffing' short bursts of odour, instead of stimulation, the traditional view of above- inhaling a continuous stream, has been sug- threshold pheromone stimulation as a time- gested as a mechanism in humans and other averaged concentration (Bossert & Wilson, mammals for preventing and a 1963) needs revision to take into account more rapid waning of sensitivity (Stoddart, 1976) than the average concentration or even the 'Flicking' of the olfactory organs by the spiny peak pheromone flux (Elkinton & Card6, lobster enhances the detection of gradual con- 1984) Frequency and amplitude of peak-to- centration changes by receptors (Schmitt & trough concentration changes may be sufficient Ache, 1979) Insects have no known mechan- 264 T C Baker et a1

ism for sniffing odou~,and hence the intermit- Carde, R T & Hagaman, T E (1979) Behavioral tent stimulation provided by odour plumes, responses of the gypsy moth in a wind tunnel to air-borne enantiomers of disparlure Environ- perhaps enhanced by the insects' zigzagging mental Entomology, 8, 475-484 flight paths, may result in the same effect Cardk, R T & Roelofs, W L (1973) Temperature modification of male sex pheromone r esponse and factors affecting female calling in Holomelina immaculate: (Lepidoptera Arctiidae) Canadian Acknowledgments Entomologist, 105, 1505-1512 Conner , W E , Eisner , T , Vander Meer , R K , Guerrero, A , Ghirmgelli, D & Meinwald, J This research was supported by NSF grant (1980) Sex attractant of an arctiid moth (Utethe- ENS 8310980 (to T C B ) We thank J S sia ornatrix) a pulsed chemical signal Behavioral Kennedy and C T David for helpful reviews Ecology and Sociobiology, 7, 55-63 of earlier drafts of this manuscript, J S David, C T , Kennedy, J S & Ludlow, A R (1983) Kennedy foi suggesting the use of the word Finding of a sex pheromone source by gypsy moths released in the field Nature, 303,804-806 'symmetry' for inter-reversal track angles per- Elkinton, I S & Card&, R T (1984) Odor disper- formed across the windline, C T David for sion of Insects (ed by W J permission to cite his unpublished results, C Bell and R T Cardk), pp 73-91 Chapman & Schal and R T Card6 for permission to cite Hall, London 1 their manuscript in press, and R S Vetter for Kaissling, K E (1971) Insect olfaction Handbook of Sensory Physiology (ed by L Beidler), Vol 4, assistance in insect rearing and preparing pp 351-431 Springer, New York I Fig 1 Kennedy, J S (1982) Mechanisms of moth sex t attraction a modified view based on wind-tunnel experiments with flying male Adoxophyes Les Colloques de l'INRA, 7, 189-192 References Kennedy, J S (1983) Zigzagging and casting as a programmed response to wind-borne odour a Baker, T C (1985) Chemical control of behavior review Physiological Entomology, 8, 109-120 Comprehensive Insect Physiology, Biochemistry, Kennedy, J S , Ludlow, A R & Sanders, C J and Pharmacology (ed by G A. Kerkut and (1980) Guidance system used in moth sex attrac- L I Gilbert), Vol 9, pp 621-672 Pergamon tion Nature, 288, 475-477 Press, Oxford Kennedy, J S , Ludlow, A R & Sanders, C J Baker, T C & Cardk, R T (1979) Endogenous and (1981) Guidance of flying male moths by wind- exogenous factors affecting periodicities of borne sex pheromone Physiological Ento- female calling and male sex pheromone response mology, 6, 395-412 in Grapholitha molesta (Busck) Journal of Insect Kuenen, L P S & Baker, T C (1981) Habituation Physiology, 25, 943-950 versus sensory adaptation as the cause of reduced Baker, T C & Roelofs, W L (1976) Electro- attraction following pulsed and constant sex antennogram responses of the male moth, Argyr- pheromone pie-exposure in Trichoplusia ni otaenia velutinana to mixtures of sex pheromone Journal of Insect Physiology, 27, 721-726 components of the female Journal of Insect Kuenen, L P S & Baker, T C (1982) The effects of pheromone concentration on the flight behavioui t Physiology, 22, 1357-1364 f Baker, T C & Roelofs, W L (1981) Initiation and of the oriental fruit moth, Grapholitha molesta termination of oriental fruit moth male response Physiological Entomology, 7, 423-434 to pheromone concentrations in the field En- Kuenen, L P S & Baker, T C (1983) A non- vironmental Entomology, 10, 211-218 anemotactic mechanism used in pheromone b Baker, T C , Meyei, W & Roelofs, W L (1981) souice location by flying moths Physiological Sex pheromone dosage and blend specificity of Entomology, 8, 277-289 response by oriental fruit moth males Entomolo- Linn, C E , Jr & Roelofs, W L (1983) The effect of gia Experimentalis et Applicata, 30, 269-279 varying proportions of the alcohol component on Bossert, W H & Wilson, E 0 (1963) The analysis sex pheromone blend discrimination in male of olfactory communication in animals Journal oriental fruit moths Physiological Entomology, of Theoretical Biology, 5, 443-469 8, 291-306 Caide, A M , Baker, T C & Cardk, R T (1979) Maish, D , Kennedy, J S & Ludlow, A R (1978) Identification of a four-component sex phei- An analysis of anemotactic zigzagging flight in omone of the female oriental fruit moth, male moths stimulated by pheromone Physiolo- Graphohtha molesta (Lepidopter a: Toi tiicidae) gical Entomology, 3, 221-240 Journal of Chemical Ecology, 5, 423-427 Murlis, J & Jones, C D (1981) Fine-scale structure Card6, R T , Dindonis, L L , Agar, B & Foss, J of odour plumes in relation to distant pheromone (1984) Apparency of pulsed and continuous pher- and other attractant sources Physiological En- omone to male gypsy moths Journal of Chemical tomology, 6, 71-86 Ecology, 10, 335-348 Olberg, R M (1983) Pheromone-triggered flip- Pulsed sex pheromone cloud evokes upwind flight 265

flopping interneurons in the ventral nerve cord of responses of a decapod crustacean are enhanced the silkworm moth, Bombyx mori Journal of by flicking Science, 205, 204-206 Comparative Physiology, 152, 297-307 Steel, R G D & Torrie, J H (1960) Principles and Roelofs, W L & Comeau, A (1971) Sex pher- Procedures of Statistics McGraw-Hill, New omone perception electroantennogram re- York sponses of the redbanded leafroller moth Jour- Stoddart, D M (1976) Mammalian Odours and nal of Insect Physiology, 17, 1967-1982 Edward Arnold, London. Roelofs, W L , Comeau, A & Selle, R (1969) Sex Willis, M A & Baker, T C (1984) Effects of pheromone of the onental fruit moth Nature, intermittent and continuous pheromone stimula- 224, 723 tion on the flight behaviour of the oriental fruit Ryan, T A (1960) Significance tests for multiple moth, Grapholita molesta Physiological En- comparison of proportions, variances and other tomology, 9, 341-358 statistics Psychological Bulletin, 57, 318-328 Wright, R H (1958) The olfactory guidance of flying Schal, C & Cardi, R T (1985) Rhythmic extrusion insects Canadian Entomologist, 90, 81-89 of pheromone gland elevates pheromone release rate Expenentia (in press) Schmitt, B C & Ache, B W (1979) Olfaction. Accepted 1 May 1985