The glomeruli of the antennal lobes in insects reflect the organization of the olfactory system, which is important for species-specific behaviors in response to insect- and plant-produced odorants. We studied the antennal lobes of the polyphagous moth Helicoverpa armigera and the oligophagous H. assulta (Heliothinae; Lepidoptera; Noctuidae) in order to see whether there are any anatomical differences that might elucidate how information about odorants is analyzed. Three-dimensional models of the antennal lobes were made, based on synaptic antibody staining combined with confocal laser scanning microscopy. These showed 65 glomeruli in each sex of H. armigera and 66 glomeruli in females of H. assulta. Sixty-two of the glomeruli were identified in both sexes and species and were given the same numbers. The sex-specific glomeruli included three macroglomerular units in H. armigera males, as well as three and four female-specific glomeruli in H. armigera and H. assulta, respectively. The species specificity of H. assulta females also appeared by the particular large size of two ordinary glomeruli. The accumulating knowledge on how biologically relevant information is encoded in receptor and antennal lobe neurons in heliothines makes these moths particularly interesting for studying the functional organization of the glomeruli. The anatomical atlases of the antennal lobes, as presented here, are prerequisites for identifying glomeruli ascribed to particular functions across sexes and species.

The importance of olfactory learning in host plant selection is well demonstrated in insects, including the heliothine moths. In the present study olfactory conditioning of the proboscis extension response was performed to determine the moths' ability to learn and discriminate three plant odorants: β-ocimene and β-myrcene (activating the same receptor neurone type), and racemic linalool (activating two different types). The conditioned stimulus (CS) was an air puff with each odorant blown into a constant air stream and over the antennae, and the unconditioned stimulus (US) was sucrose solution applied first to the antennal taste sensilla, then to the proboscis. Conditioning with increasing odorant concentrations induced increased learning performance. The concentration threshold for learning was 100 times lower for racemic linalool than for the two other odorants, a fact that can be correlated with a higher sensitivity of the moths' antennae to racemic linalool as shown in electroantennogram recordings. After correcting for the different odour sensitivities, the moths' ability to discriminate the odorants was studied. Differential conditioning experiments were carried out, in which moths had to distinguish between a rewarded (CS+) odorant and an explicitly unrewarded odorant (CS-), choosing odour concentrations giving the same learning rate in previous experiments. The best discrimination was found with β-myrcene as the rewarded odorant and racemic linalool as the unrewarded. The opposite combination gave lower discrimination, indicating a higher salience for β-myrcene than for racemic linalool. The moths could also discriminate between β-ocimene and β-myrcene, which was surprising, since they activate the same receptor neurone type. No difference in salience was found between these two odorants.

The primary olfactory centre, the antennal lobe of Heliothis virescens moths, contains 62 glomeruli which process plant odour information and four male-specific glomeruli which form the macroglomerular complex, involved in processing information about pheromone and interspecific signals. Using calcium imaging, we recorded the spatio-temporal activity pattern of the glomeruli in the anterior antennal lobe during stimulation with odorants produced by plants or insects. Each odorant elicited specific excitatory responses in one or a few glomeruli: the major pheromone component did so exclusively in the large glomerulus of the macroglomerular complex and the plant odours exclusively in the ordinary glomeruli. Eight glomeruli, with corresponding plant odour responses and positions, were identified within each sex. Glomeruli responded specifically to linalool, beta-ocimene/beta-myrcene or germacrene D/alpha-farnesene. Responses to two essential plant oils covered the response areas of their major constituents, as well as activating additional glomeruli. Stronger activation in the AL due to increased odour concentration was expressed as increased response strength within the odorant-specific glomeruli as well as recruitment of less sensitive glomeruli.