Novel natural ligands for Drosophila olfactory receptor neurones
Marcus C. Stensmyr1,
Elena Giordano2,
Annalisa Balloi2,
Anna-Maria Angioy2 and
Bill S. Hansson1,*
1 Division of Chemical Ecology, Department of Crop Science, Swedish
University of Agricultural Sciences, PO Box 44, SE-23053 Alnarp,
Sweden
2 Department of Experimental Biology, University of Cagliari, S.S. 554 Km,
4500 Monserrato, Italy
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Fig. 1. Simultaneously recorded activity of two co-located olfactory receptor
neurones. Differences in action potential (spike) amplitude allow for
separation of activity. Stimulation with a blank results in no increased
activity from any of the neurones. Stimulation with ethyl 3-hydroxybutyrate
elicits a strong response from the smaller spiking neurone, indicated by `B'
(neurone later classified as S2B), whereas the larger spiking neurone,
indicated by `A' is unaffected by the stimulus. Boxed area shows an expanded
part of the recording. Horizontal bar indicates stimulus duration, 0.5 s.
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Fig. 2. A linked gas chromatography—single cell recording from an antennal
sensillum (S5) containing two olfactory receptor neurones (ORNs). (A) Gas
chromatogram (GC) of a papaya head-space sample and the corresponding
single-cell recording (SC), presented in histogram form (spikes
s-1; vertical bar, 25 spikes s-1; horizontal bar, 1
min). The two ORNs present are sensitive to different components in the papaya
volatile collection. The S5A neurone responds primarily to butyl butyrate (5),
but also to 1-hexanol (2) and isovaleric acid (4). The S5B neurone responds
strongly to isoamyl acetate (3) and moderately to butyl acetate (1). (B)
Action potential amplitudes recorded during the above experiment. The
distribution of amplitudes from the recording shows the clear physiological
separation of the two ORNs (S5A and S5B).
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Fig. 3. Physiological sensillum types were identified by the response spectra of
their olfactory receptor neurones (ORNs) when challenged with volatiles
collected from different fruits. (A) An S3A ORN (number 52) stimulated with a
volatile collection from passion fruit. A strong response is elicited by ethyl
hexanoate [peak 4 in the gas chromatogram (GC)], while smaller responses are
elicited by ethyl butyrate (1), ethyl 3-hydroxybutyrate (2), ethyl
3-hydroxyhexanoate (5) and an unidentified compound (3). The physiological
response is represented in the histogram displaying the single cell (SC)
response over time (spikes s-1; vertical bar, 25 spikes
s-1; horizontal bar, 1 min). (B) The same neurone challenged with a
volatile collection from banana reveals three smaller responses to isoamyl
acetate (6), methyl hexanoate (7) and butyl butyrate (8). (C) Two S3A ORNs
(numbers 19 and 31) show the repeatability of the responses of a certain ORN
type to a particular extract, here a pineapple volatile collection, where
ethyl hexanoate (4) is again the major stimulus, while methyl hexanoate (7)
elicits a weaker response.
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Fig. 4. Identified classes of antennal sensilla based on physiological
characteristics of olfactory receptor neurones (ORNs) present. Each sensillum
type (S1—S8) is presented together with the response spectra of the
respective ORNs (A, B, C and D) housed within each sensillum type. Beneath the
sensillum type the number (N) encountered is indicated. Beneath the
molecular configuration of each ligand type the maximum response is indicated
(spikes s-1). NR, no response to any tested stimulus.
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Fig. 5. Antennal distribution of the sensillum types identified. (A) Frontal view
of a Drosophila head. The olfactory organs, antennae (above) and
maxillary palps (below), are marked in green. (B) Schematic drawing of the
Drosophila third antennal segment's posterior and anterior face. LB,
large s. basiconica region; A, arista; S, sacculus entrance; ST, s. trichodea
region. The locations of the different sensillum types investigated are
indicated by coloured circles (corresponding to the colours in
Fig. 4). The total number of
each sensillum type encountered is indicated in parentheses. Circles with a
black dot in the centre represent investigation with GC—SC methodology,
while open circles were characterised by stimulation with synthetic
compounds.
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Fig. 6. Dose—response relationships for five of the identified olfactory
receptor neurone (ORN) classes. Odorants were presented in increasing dosages
from 0.1 ng to 10 µg (x-axis). ORN response to a stimulus is given
as the net number of action potentials (number of spikes during 1 s after
stimulation minus the number of spikes during 1 s prior to stimulation) minus
the net blank response (y-axis). (A) Dose—response curves for
the S3A neurone type, (B) S2B type, (C) S1B type, (D) S5A type and (E) S8B
type. For details on each ORN type, see
Fig. 4. Vertical bars indicate
the S.E.M.
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Fig. 7. Behavioural responses to a set of the identified ligands. Fly behaviour was
tested in a T-maze setup and odorants were presented over a wide concentration
range. The behavioural response functions are based on 392 experiments with a
total of 7120 flies. A response index (RI) of 1.0 equals full
attraction, whereas an RI of -1.0 equals full avoidance. Indifference
to the odour is indicated by an RI of 0. (A) RI curve for
acetoin (N=89, number of experiments used to create the RI curve),
(B) butyl butyrate (N=43), (C) ethyl hexanoate (N=57), (D)
ethyl 3-hydroxybutyrate (N=52), (E) 1-hexanol (N=51), (F)
phenylacetonitrile (N=46), (G) ethyl acetate (N=48).
Asterisks indicate RI values significantly different from zero
(P<0.05,  2 test). Vertical bars indicate the
S.E.M.
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© The Company of Biologists Ltd 2003
