Chemical Senses Vol. 30 No. suppl 1 © Oxford University
Press 2005; all rights reserved
Innate and Changed Responses to Plant Odours in Moths and Weevils
Department of Biology, Neuroscience Unit, Norwegian University of Science and Technology, Trondheim, Norway
Correspondence to be sent to: Hanna Mustaparta, e-mail: hanna.mustaparta{at}bio.ntnu.no
Key words: antennal lobe projection neurons, olfactory coding, olfactory receptor neurons, suboesophageal ganglion, taste receptor neuron projections
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Introduction |
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 Top Introduction Identification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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The challenge for the olfactory system in animals is to detect the large diversity of molecules released by food or host plants and to discriminate these odours from irrelevant ones. Molecular biological studies of vertebrates and insects have shown that the olfactory information is not handled by a few, but by a large and species-specific number of receptor proteins (Vosshall, 2003
;
Mombarts, 2004). Furthermore, the
olfactory receptor neurons (ORNs) are divided into subpopulations, each expressing only
one type of receptor protein and converging in one or two specific glomeruli in the
primary olfactory centres, i.e. the antennal lobe in insects and olfactory bulb in
vertebrates. Questions of interest in our studies of insects are how receptor and central
interneurons encode plant odour information leading to behavioural responses as well as
the neuronal mechanisms underlying olfactory learning. By comparing related species of
moths (subfamily Heliothinae) with distantly related moths and weevils, our intention is
to identify similarities and differences of the olfactory coding mechanisms across
species. Recognition of odours requires learning and memory of the odour, as shown in the
honeybee (Menzel, 2001). Heliothine
moths also can learn odours, which seems to be important in the selection of host plants
for nectar feeding and egg-laying (Cunningham
et al., 1998;
Hartlieb et al., 1999; H.T.
Skiri et al., unpublished data). |
Identification of biologically relevant plant odorants |
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TopIntroductionIdentification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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The first question to be asked is which compounds of the complex mixtures of volatiles released by host and non-host plants are detected by the ORNs. Naturally produced volatiles are trapped by adsorbents during aeration of intact plants or cut plant materials (‘head-space’ technique) (Røstelien et al., 2000a
). The compounds are
then eluted by solvents and used as test samples. Gas chromatography with two parallel
columns linked to electrophysiological recordings from single neurons is employed to test
the stimulation of the ORNs by the separated compounds. The use of one chiral column for
separating optical isomers has enabled tests of pure enantiomers on single ORNs
(Stranden et al., 2002,
2003a). The effective compounds are then identified by GC-MS and NMR, followed by
retesting authentic materials on the ORNs. This provides information not only on which
odorants are detected, but also about the molecular receptive ranges of the ORNs. |
The molecular receptive range and enantioselectivity of the olfactory receptor neurons in closely and distantly related species |
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TopIntroductionIdentification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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The results have shown that the ORNs in these insect species primarily respond by excitation to the odorants and fall into distinct types according to their molecular receptive ranges (Røstelien et al., 2000a
,b;
Stranden et al., 2002,
2003a,b). They show strong responses to one or two primary odorants and weak responses to
a few chemically related compounds (secondary odorants), demonstrating a narrow tuning.
These results correlate well with the principle that one type of receptor protein is
expressed in each ORN. The molecular receptive ranges show no or minimal overlap for
neurons responding to the same chemical group. For instance, the monoterpene alcohol,
(+)-linalool, being the primary odorant for one ORN type, is a secondary odorant for
the ORNs primarily responding to another monoterpene alcohol, geraniol. In three related
species of heliothine moths the same functional types of ORNs have been found. One
example is the germacrene D type, for which (–)-germacrene D has a 10-fold stronger
effect than (+)-germacrene D, which differs in the direction of the isopropyl group
in relation to the 10-carbon ring (Stranden
et al., 2003a,b). The much weaker effect of three other
sesquiterpene molecules seems to be due to their less flexible ring systems. These
properties are similar for all germacrene D types of ORNs in three heliothine species. In
addition, the strawberry weevil (Anthonomus rubi) has one ORN type tuned to
(–)-germacrene D, responding with lower sensitivity to (+)-germacrene D
(Bichao et al., unpublished data). However, in this species the secondary
odorants are different molecules to those in the heliothine moths, suggesting that the
germacrene D ORNs in the two insect groups have evolved independently in the adaptation
to their host plants. Species differences are also shown in respect to the presence of
different ORN types tuned to the opposite enantiomers of linalool (Røstelien et al., 2000a,b). Altogether, the
results obtained in these moth and weevil species show a narrow tuning and a minimal
overlap of the molecular receptive ranges of the ORNs. These data on insects are in
contrast to the results of broadly tuned ORNs reported in vertebrates, and may reflect
the low homology found between the receptor proteins in insects and vertebrates
(Breer, 2003). |
The antennal lobe: optical imaging and intracellular recordings from projection neurons |
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TopIntroductionIdentification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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Models of the antennal lobes of three heliothine species have shown the presence of 62–64 glomeruli involved in plant odour information (Berg et al., 2001
; H.T. Skiri et al.,
unpublished data). In one study, optical recordings of the glomerular activity elicited
by some of the primary and secondary odorants have been studied (H.T. Skiri et
al., unpublished data). In these recordings, covering ~20 glomeruli of the anterior
antennal lobe (AL), each primary odorant elicited activity in specific glomeruli. In
addition, three glomeruli were found that responded to two or three primary odorants,
which is in contrast to the principle of one ORN type converging in each glomerulus. The
results from these optical recordings, indicating the input to the glomeruli, are
compared with the responses of projection neurons (PNs) in the antennal lobe,
representing the glomerular output. Responses of PNs to primary and secondary odorants
are studied by the use of intracellular recordings combined with injection of fluorescent
dyes for visualization of the morphology of the neurons by confocal laser scanning
microscopy (CLSM). This is followed by reconstruction of the neurons and the innervated
brain structures using the AMIRA software. Different types of PNs have been demonstrated.
They are characterized by physiological responses to different odorants, dendrite
arborizations in one or more glomeruli, axons in different antenno-cerebral tracts and
projection patterns in the calyces of the mushroom bodies and lateral protocerebrum
(Müller et al., 2002).
The responses of the PNs are compared with the molecular receptive ranges of the
functionally characterized ORNs, and this indicates that there is excitatory input from
one ORN type and inhibition mediated via local interneurons from another ORN type.
Recordings from a multiglomerular PN with scarce innervation in each of many glomeruli
indicated that specific blends of odorants might be required for spike firing. In further
studies we hope to determine the overall representation of the odour quality of primary
and secondary odorants in the glomeruli of the antennal lobe and find out whether the
different antenno-cerebral tracts in these moth species convey different biologically
relevant information. |
Projections of gustatory receptor neurons in the brain and the possible neuronal connection with the olfactory pathway involved in associative learning |
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TopIntroductionIdentification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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Associative learning is studied by the use of the proboscis extension response (PER) (Menzel, 2001
). By pairing an odour
stimulus (conditioned stimulus), followed by sucrose stimulation to the gustatory
sensilla (unconditioned stimulus), the insects learn to associate the odour with the
sucrose reward. In order to trace the neuronal connections between the pathways mediating
the unconditioned and the conditioned information in heliothine moths, the projections of
the gustatory receptor neurons have been traced. Two populations of gustatory receptor
neurons are present, one located in the contact chemosensilla on the antennae (sensilla
chaeticae) and another in sensilla styloconicae on the proboscis. By the use of
fluorescent dyes for visualization in CLSM and reconstruction in AMIRA, the projections
are found in two closely located fingerlike areas in the SOG (Jørgensen et al., 2002;
Kvello et al., 2002). One
neuron has been found that might form the connection between the taste and the olfactory
pathway (Müller et al.,
2002). The morphology of this neuron resembles the VUMmx1 involved in
associative learning in the honeybee (Hammer,
1993). Thus, it may mediate the connection between the US and CS in the
heliothine moths. In behavioural experiments we have started to study the importance of olfactory learning in moths and weevils. In moths, conditioning of the PER is used to determine the ability to learn the identified odorants, including concentration dependency and salience (H.T. Skiri et al., unpublished data). By differential conditioning, the ability to discriminate between primary and secondary odorants are revealed, e.g. whether the moths may more easily distinguish different primary odorants than primary and secondary odorants activating the same ORNs, as indicated in the study by Skiri et al. The perception and discrimination of single odorants and mixtures are investigated in further experiments on PER conditioning. In another learning study of the pine weevil (Hylobius abietis), multiple-choice experiments are used to investigate how previous experience with food materials influences the preference for an odour. The weevils were found to change their preference for two attractive enantiomers, depending on the experience with different food materials (O. Roten et al., personal communication). The enantiomeric content in the food plants correlated with the preferred enantiomer. Thus, olfactory learning in addition to the innate responses to odorants may play a significant role for host attraction in the pine weevil as well.
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References |
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TopIntroductionIdentification of biologically...The molecular receptive range...The antennal lobe: optical...Projections of gustatory...References |
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