No abstract

Each down stroke of an insect's wings accelerates axial airflow over the antennae. Modeling studies suggest that this can greatly enhance penetration of air and air-born odorants through the antennal sensilla thereby periodically increasing odorant-receptor interactions. Do these periodic changes result in entrainment of neural responses in the antenna and antennal lobe (AL)? Does this entrainment affect olfactory acuity? To address these questions, we monitored antennal and AL responses in the moth Manduca sexta while odorants were pulsed at frequencies from 10–72 Hz, encompassing the natural wingbeat frequency. Power spectral density (PSD) analysis was used to identify entrainment of neural activity. Statistical analysis of PSDs indicates that the antennal nerve tracked pulsed odor up to 30 Hz. Furthermore, at least 50% of AL local field potentials (LFPs) and between 7–25% of unitary spiking responses also tracked pulsed odor up to 30 Hz in a frequency-locked manner. Application of bicuculline (200 μM) abolished pulse tracking in both LFP and unitary responses suggesting that GABAA receptor activation is necessary for pulse tracking within the AL. Finally, psychophysical measures of odor detection establish that detection thresholds are lowered when odor is pulsed at 20 Hz. These results suggest that AL networks can respond to the oscillatory dynamics of stimuli such as those imposed by the wing beat in a manner analogous to mammalian sniffing.

Behavioral studies of olfactory discrimination and stimulus generalization in many species indicate that the molecular features of monomolecular odorants are important for odor discrimination. Here we evaluate how features, such as carbon chain length and functional group, are represented in the first level of synaptic processing. We recorded antennal lobe ensemble responses in the moth Manduca sexta to repeated 100-ms pulses of monomolecular alcohols and ketones. Most units exhibited a significant change in spike rate in response to most odorants that outlasted the duration of the stimulus. Peristimulus data were then sampled over 780 ms for each pulse of all odorants. Factor analysis was used to assess whether there were groups of units with common response patterns. We found that factors identified and represented activity for clusters of units with common temporal response characteristics. These temporally patterned responses typically spanned 780 ms and were often dependent on carbon chain length and functional group. Furthermore, cross-correlation analysis frequently indicated significant coincident spiking even during spontaneous activity. However, this synchrony occurred mainly between units recorded on the same tetrode. In a final analysis, the Euclidean distance between odor responses was calculated for each pair of odorants using factors as dimensions. The distance between responses for any two odorants was maximized by approximately 240 ms. This time course corresponded to the brief sequence of coordinated bursts across the recorded population. The distance during this period was also a function of systematic differences in molecular features. Results of this Euclidian analysis thus directly correlate to previous behavioral studies of stimulus generalization in M. sexta.

Recent evidence suggests that odor-driven responses in the insect antennal lobe (AL) can be modified by associative and nonassociative processes, as has been shown in the vertebrate olfactory bulb. However, the specific network changes that occur in response to olfactory learning remain unknown. To characterize changes in AL network activity during learning, we developed an in vivo protocol in Manduca sexta that allows continuous monitoring of neural ensembles and feeding behavior over the course of olfactory conditioning. Here, we show that Pavlovian conditioning produced a net recruitment of responsive neural units across the AL that persisted after conditioning. Recruitment only occurred when odor reliably predicted food. Conversely, when odor did not predict food, a net loss of responsive units occurred. Simultaneous measures of feeding responses indicated that the treatment-specific patterns of neural recruitment were positively correlated with changes in the insect's behavioral response to odor. In addition to recruitment, conditioning also produced consistent and profound shifts in the temporal responses of 16% of recorded units. These results show that odor representations in the AL are dynamic and related to olfactory memory consolidation. We furthermore provide evidence that the basis of the learningdependent changes in the AL is not simply an increase in activity in the neural network representing an odorant. Rather, learning produces a restructuring of spatial and temporal components of network responses to odor in the AL. In the olfactory systems of animals as diverse as insects (1, 2) and vertebrates (3, 4), a large number of odor stimuli can be represented combinatorially by relatively few glomeruli. These systems share a common organization whereby each glomerulus receives primary-afferent input from olfactory receptor cells expressing a specific receptor phenotype (5). A given odor compound activates a spatially distributed ensemble of glomerular projection neurons (PNs), and individual PNs participate in the representation of multiple odor compounds (1,3,(6)(7)(8). Within these ensembles, different features of an odor stimulus are also represented in the temporal structure of odor-driven responses among PNs (1,3,6,7,(9)(10)(11)(12).Like their vertebrate counterparts, foraging insects, such as honeybees, fruit flies, and moths, give modified behavioral responses to an odor stimulus associated with food (13-19). Evidence in vertebrates suggests that the representation of a given odor in the first synaptic relay, the olfactory bulb (OB, the analogue of the insect antennal lobe), can be modified as a result of experience with an odor (6,(20)(21)(22)(23)(24)(25)(26). Preliminary studies of learning-dependent changes in the insect antennal lobe (AL), using imaging techniques, also suggest that olfactory responses in this structure might change as a result of experience (27,28). With an appetitive conditioning paradigm, for example, increased odor-driven calcium signals were observed in honeybee ALs w...

A central problem facing studies of neural encoding in sensory systems is how to accurately quantify the extent of spatial and temporal responses. In this study, we take advantage of the relatively simple and stereotypic neural architecture found in invertebrates. We combine standard electrophysiological techniques, recently developed population analysis techniques, and novel anatomical methods to form an innovative 4-dimensional view of odor output representations in the antennal lobe of the moth Manduca sexta. This novel approach allows quantification of olfactory responses of characterized neurons with spike time resolution. Additionally, arbitrary integration windows can be used for comparisons with other methods such as imaging. By assigning statistical significance to changes in neuronal firing, this method can visualize activity across the entire antennal lobe. The resulting 4-dimensional representation of antennal lobe output complements imaging and multi-unit experiments yet provides a more comprehensive and accurate view of glomerular activation patterns in spike time resolution.