Sensory processing in the Drosophila antennal lobe increases reliability and separability of ensemble odor representations

Intravital microscopy is a key means of monitoring cellular function in live organisms, but surgical preparation of a live animal for microscopy often is time-consuming, requires considerable skill, and limits experimental throughput. Here we introduce a spatially precise (<1-μm edge precision), high-speed (<1 s), largely automated, and economical protocol for microsurgical preparation of live animals for optical imaging. Using a 193-nm pulsed excimer laser and the fruit fly as a model, we created observation windows (12-to 350-μm diameters) in the exoskeleton. Through these windows we used two-photon microscopy to image odor-evoked Ca 2+ signaling in projection neuron dendrites of the antennal lobe and Kenyon cells of the mushroom body. The impact of a laser-cut window on fly health appears to be substantially less than that of conventional manual dissection, for our imaging durations of up to 18 h were ∼5-20 times longer than prior in vivo microscopy studies of hand-dissected flies. This improvement will facilitate studies of numerous questions in neuroscience, such as those regarding neuronal plasticity or learning and memory. As a control, we used phototaxis as an exemplary complex behavior in flies and found that laser microsurgery is sufficiently gentle to leave it intact. To demonstrate that our techniques are applicable to other species, we created microsurgical openings in nematodes, ants, and the mouse cranium. In conjunction with emerging robotic methods for handling and mounting flies or other small organisms, our rapid, precisely controllable, and highly repeatable microsurgical techniques should enable automated, high-throughput preparation of live animals for optical experimentation.laser surgery | calcium imaging | Drosophila melanogaster I ntravital imaging of small model organisms is a staple technique in many fields of biomedicine. Gaining optical access to the body's interior usually requires removal or thinning of a highly scattering, protective exterior. Unfortunately, surgical creation of an optical window is generally time-consuming, requires considerable skill and dexterity, and often represents an unwanted source of experimental variability. To address these challenges, we created laser surgical means to open optical windows in the bodies of live animals.Our approach is adaptable to many species and applications, but we mainly focused on fluorescence brain imaging in alert fruit flies, a pursuit of growing importance for the study of neural systems (1, 2). Most prior studies that have monitored flies' neural activity have relied on immobilized fly preparations suitable for optical microscopy (3, 4) or electrophysiology (5, 6). Recent improvements in genetically encoded Ca 2+ indicators and imaging methods now allow in vivo fluorescence imaging of neural activity in behaving flies (2). The fly brain's small size, <250 μm deep in the adult, is conducive to two-photon imaging of a variety of neural populations after opening of the cuticle (2, 4).Multiple laboratories have developed protoco...

Section: Two-photon Ca 2+ Imaging Of Neural Dynamics Over Extended Timesupporting

confidence: 86%

High-speed laser microsurgery of alert fruit flies for fluorescence imaging of neural activity

Sinha¹,

Liang²,

et al. 2013

“…1A) and then determining the corresponding PN responses on the basis of an experimental characterization of the transformations occurring within the antennal lobe (see below) (21,22). Finally, we use the computed PN responses to drive model LHNs and KCs.…”

Section: Resultsmentioning

confidence: 99%

Generating sparse and selective third-order responses in the olfactory system of the fly

Luo¹,

Axel

Abbott

2010

134

137

In the antennal lobe of Drosophila, information about odors is transferred from olfactory receptor neurons (ORNs) to projection neurons (PNs), which then send axons to neurons in the lateral horn of the protocerebrum (LHNs) and to Kenyon cells (KCs) in the mushroom body. The transformation from ORN to PN responses can be described by a normalization model similar to what has been used in modeling visually responsive neurons. We study the implications of this transformation for the generation of LHN and KC responses under the hypothesis that LHN responses are highly selective and therefore suitable for driving innate behaviors, whereas KCs provide a more general sparse representation of odors suitable for forming learned behavioral associations. Our results indicate that the transformation from ORN to PN firing rates in the antennal lobe equalizes the magnitudes of and decorrelates responses to different odors through feedforward nonlinearities and lateral suppression within the circuitry of the antennal lobe, and we study how these two components affect LHN and KC responses.antennal lobe | decorrelation | lateral horn | normalization | olfaction I n the olfactory system, as in other sensory systems, signals from primary receptors are processed and transformed before being relayed to higher brain areas. In Drosophila melanogaster, olfactory receptor neurons (ORNs) located in the antennae and maxillary palps synapse onto projection neurons (PNs) within the glomeruli of the antennal lobes (Fig. S1). ORNs expressing a given receptor converge on an anatomically invariant glomerulus. PNs innervate a single glomerulus and thus receive their primary input from ORNs expressing the same olfactory receptor (1), although crossglomerular interactions mediated by interneurons are also present (2-5). The PNs send their axons to two distinct regions of the fly brain, the lateral horn and the mushroom body. By constructing models of neurons in these regions, we study the effects of the transformations arising from circuitry within the antennal lobe on the capacity of neurons in the lateral horn and the mushroom body to represent and discriminate odors.Any functional interpretation of the transformation from ORN to PN responses depends on the nature of the processing being performed by the third-order neurons that receive PN input. The lateral horn and mushroom body appear to be involved in different forms of sensory processing. The lateral horn is believed to be important in generating innate behaviors (6), including those elicited by pheromones (7). PN projections to lateral horn neurons (LHNs) are spatially stereotyped (8, 9) and clustered (10), suggesting that circuits in this region may be "hardwired" for the detection of specific odors that elicit innate behavioral responses. The mushroom body is implicated in decision making (11) and in the formation of associative memories (12, 13). Both calcium imaging (14) and electrophysiology (15) indicate that the responses of Kenyon cell (KCs) in the mushroom body are sparse, with e...

“…This information-theoretic approach may also be extended to analyzing the dynamic role of PNs and local interneurons in processing and transmitting information to higher-order structures. Functional imaging and electrophysiological studies have provided useful insights into information processing in the antennal lobe and mushroom body (39)(40)(41)42). Although local interneurons modulate PNs activity, ORNs are the primary drivers of this activity (43).…”

Section: Discussionmentioning

confidence: 99%

Post-eclosion odor experience modifies olfactory receptor neuron coding in Drosophila

Iyengar

Chakraborty

Goswami

et al. 2010

Olfactory responses of Drosophila undergo pronounced changes after eclosion. The flies develop attraction to odors to which they are exposed and aversion to other odors. Behavioral adaptation is correlated with changes in the firing pattern of olfactory receptor neurons (ORNs). In this article, we present an information-theoretic analysis of the firing pattern of ORNs. Flies reared in a synthetic odorless medium were transferred after eclosion to three different media: (i) a synthetic medium relatively devoid of odor cues, (ii) synthetic medium infused with a single odorant, and (iii) complex cornmeal medium rich in odors. Recordings were made from an identified sensillum (type II), and the Jensen-Shannon divergence (D JS ) was used to assess quantitatively the differences between ensemble spike responses to different odors. Analysis shows that prolonged exposure to ethyl acetate and several related esters increases sensitivity to these esters but does not improve the ability of the fly to distinguish between them. Flies exposed to cornmeal display varied sensitivity to these odorants and at the same time develop greater capacity to distinguish between odors. Deprivation of odor experience on an odorless synthetic medium leads to a loss of both sensitivity and acuity. Rich olfactory experience thus helps to shape the ORNs response and enhances its discriminative power. The experiments presented here demonstrate an experience-dependent adaptation at the level of the receptor neuron.imaginal conditioning | sensory adaptation | odor imprinting | JensenShannon divergence | chemo receptor tuning O lfaction in the fruit fly, Drosophila melanogaster, is crucial for a variety of behaviors, including associative learning (1, 2), courtship (3), foraging (4), and flight (5, 6). Odorants are detected by approximately 1,300 olfactory receptor neurons (ORNs), which are housed in sensilla on the third antennal segment and individually express one of approximately 50 functional odor receptors in adults (7-9). All ORNs expressing the same olfactory receptor project to one of approximately 50 glomeruli in the antennal lobe, where they synapse with a set of projection neurons (PNs) (10, 11). The activity of ORNs, either excitation or inhibition, provides behaviorally relevant information about odorants such as their identity, concentration, and source. The information transduced by an ORN is processed in the antennal lobe (12, 13) and sent via PNs to the mushroom bodies, which are believed to be centers for olfactory learning and memory (14, 15).Experience-dependent modifiability (i.e., plasticity) of olfactory representation at the level of the central nervous system is well known (2,16,17). Relatively little attention has been paid to long-term changes in sensory neuron activity resulting from odor experiences. It was previously shown that exposure of newly born imago to a particular set of odorants alters their responses to these odors (18,19). The flies develop an attraction to the chemicals to which they are exposed and an av...