Circuits for integrating learned and innate valences in the insect brain.

Animal behavior is shaped both by evolution and by individual experience. Parallel brain pathways encode innate and learned valences of cues, but the way in which they are integrated during action-selection is not well understood. We used electron microscopy to comprehensively map with synaptic resolution all neurons downstream of all mushroom body (MB) output neurons (encoding learned valences) and characterized their patterns of interaction with lateral horn (LH) neurons (encoding innate valences) in Drosophila larva. The connectome revealed multiple convergence neuron types that receive convergent MB and LH inputs. A subset of these receives excitatory input from positive-valence MB and LH pathways and inhibitory input from negative-valence MB pathways. We confirmed functional connectivity from LH and MB pathways and behavioral roles of two of these neurons. These neurons encode integrated odor value and bidirectionally regulate turning. Based on this, we speculate that learning could potentially skew the balance of excitation and inhibition onto these neurons and thereby modulate turning. Together, our study provides insights into the circuits that integrate learned and innate valences to modify behavior.

Escape band in Escherichia coli chemotaxis in opposing attractant and nutrient gradients.

It is commonly believed that bacterial chemotaxis helps cells find food. However, not all attractants are nutrients, and not all nutrients are strong attractants. Here, by using microfluidic experiments, we studied Escherichia coli chemotaxis behavior in the presence of a strong chemoattractant (e.g., aspartate or methylaspartate) gradient and an opposing gradient of diluted tryptone broth (TB) growth medium. Our experiments showed that cells initially accumulate near the strong attractant source. However, after the peak cell density (h) reaches a critical value [Formula: see text], the cells form a "escape band" (EB) that moves toward the chemotactically weaker but metabolically richer nutrient source. By using various mutant strains and varying experimental conditions, we showed that the competition between Tap and Tar receptors is the key molecular mechanism underlying the formation of the escape band. A mathematical model combining chemotaxis signaling and cell growth was developed to explain the experiments quantitatively. The model also predicted that the width w and the peak position [Formula: see text] of EB satisfy two scaling relations: [Formula: see text] and [Formula: see text], where l is the channel length. Both scaling relations were verified by experiments. Our study shows that the combination of nutrient consumption, population growth, and chemotaxis with multiple receptors allows cells to search for optimal growth condition in complex environments with conflicting sources.

Structured Odorant Response Patterns across a Complete Olfactory Receptor Neuron Population.

Odor perception allows animals to distinguish odors, recognize the same odor across concentrations, and determine concentration changes. How the activity patterns of primary olfactory receptor neurons (ORNs), at the individual and population levels, facilitate distinguishing these functions remains poorly understood. Here, we interrogate the complete ORN population of the Drosophila larva across a broadly sampled panel of odorants at varying concentrations. We find that the activity of each ORN scales with the concentration of any odorant via a fixed dose-response function with a variable sensitivity. Sensitivities across odorants and ORNs follow a power-law distribution. Much of receptor sensitivity to odorants is accounted for by a single geometrical property of molecular structure. Similarity in the shape of temporal response filters across odorants and ORNs extend these relationships to fluctuating environments. These results uncover shared individual- and population-level patterns that together lend structure to support odor perceptions.

Mechanical regulation of stem-cell differentiation by the stretch-activated Piezo channel.

Somatic stem cells constantly adjust their self-renewal and lineage commitment by integrating various environmental cues to maintain tissue homeostasis. Although numerous chemical and biological signals have been identified that regulate stem-cell behaviour, whether stem cells can directly sense mechanical signals in vivo remains unclear. Here we show that mechanical stress regulates stem-cell differentiation in the adult Drosophila midgut through the stretch-activated ion channel Piezo. We find that Piezo is specifically expressed in previously unidentified enteroendocrine precursor cells, which have reduced proliferation ability and are destined to become enteroendocrine cells. Loss of Piezo activity reduces the generation of enteroendocrine cells in the adult midgut. In addition, ectopic expression of Piezo in all stem cells triggers both cell proliferation and enteroendocrine cell differentiation. Both the Piezo mutant and overexpression phenotypes can be rescued by manipulation of cytosolic Ca2+ levels, and increases in cytosolic Ca2+ resemble the Piezo overexpression phenotype, suggesting that Piezo functions through Ca2+ signalling. Further studies suggest that Ca2+ signalling promotes stem-cell proliferation and differentiation through separate pathways. Finally, Piezo is required for both mechanical activation of stem cells in a gut expansion assay and the increase of cytosolic Ca2+ in response to direct mechanical stimulus in a gut compression assay. Thus, our study demonstrates the existence of a specific group of stem cells in the fly midgut that can directly sense mechanical signals through Piezo.

Barrier Crossing in Escherichia coli Chemotaxis.

We study cell navigation in spatiotemporally complex environments by developing a microfluidic racetrack device that creates a traveling wave with multiple peaks and a tunable wave speed. We find that while the population-averaged chemotaxis drift speed increases with wave speed for low wave speed, it decreases sharply for high wave speed. This reversed dependence of population-averaged chemotaxis drift speed on wave speed is caused by a "barrier-crossing" phenomenon, where a cell hops backwards from one peak attractant location to the peak behind by crossing an unfavorable (barrier) region with low attractant concentrations. By using a coarse-grained model of chemotaxis, we map bacterial motility in an attractant field to the random motion of an overdamped particle in an effective potential. The observed barrier-crossing phenomenon of living cells and its dependence on the spatiotemporal profile of attractant concentration are explained quantitatively by Kramers reaction rate theory.

The wiring diagram of a glomerular olfactory system.

The sense of smell enables animals to react to long-distance cues according to learned and innate valences. Here, we have mapped with electron microscopy the complete wiring diagram of the Drosophila larval antennal lobe, an olfactory neuropil similar to the vertebrate olfactory bulb. We found a canonical circuit with uniglomerular projection neurons (uPNs) relaying gain-controlled ORN activity to the mushroom body and the lateral horn. A second, parallel circuit with multiglomerular projection neurons (mPNs) and hierarchically connected local neurons (LNs) selectively integrates multiple ORN signals already at the first synapse. LN-LN synaptic connections putatively implement a bistable gain control mechanism that either computes odor saliency through panglomerular inhibition, or allows some glomeruli to respond to faint aversive odors in the presence of strong appetitive odors. This complete wiring diagram will support experimental and theoretical studies towards bridging the gap between circuits and behavior.

Reverse-correlation analysis of navigation dynamics in Drosophila larva using optogenetics.

Neural circuits for behavior transform sensory inputs into motor outputs in patterns with strategic value. Determining how neurons along a sensorimotor circuit contribute to this transformation is central to understanding behavior. To do this, a quantitative framework to describe behavioral dynamics is needed. In this study, we built a high-throughput optogenetic system for Drosophila larva to quantify the sensorimotor transformations underlying navigational behavior. We express CsChrimson, a red-shifted variant of channelrhodopsin, in specific chemosensory neurons and expose large numbers of freely moving animals to random optogenetic activation patterns. We quantify their behavioral responses and use reverse-correlation analysis to uncover the linear and static nonlinear components of navigation dynamics as functions of optogenetic activation patterns of specific sensory neurons. We find that linear-nonlinear models accurately predict navigational decision-making for different optogenetic activation waveforms. We use our method to establish the valence and dynamics of navigation driven by optogenetic activation of different combinations of bitter-sensing gustatory neurons. Our method captures the dynamics of optogenetically induced behavior in compact, quantitative transformations that can be used to characterize circuits for sensorimotor processing and their contribution to navigational decision making.

Discovery of novel chemoeffectors and rational design of Escherichia coli chemoreceptor specificity.

Bacterial chemoreceptors mediate chemotactic responses to diverse stimuli. Here, by using an integrated in silico, in vitro, and in vivo approach, we screened a large compound library and found eight novel chemoeffectors for the Escherichia coli chemoreceptor Tar. Six of the eight new Tar binding compounds induce attractant responses, and two of them function as antagonists that can bind Tar without inducing downstream signaling. Comparison between the antagonist and attractant binding patterns suggests that the key interactions for chemotaxis signaling are mediated by the hydrogen bonds formed between a donor group in the attractant and the main-chain carbonyls (Y149 and/or Q152) on the α4 helix of Tar. This molecular insight for signaling is verified by converting an antagonist to an attractant when introducing an N-H group into the antagonist to restore the hydrogen bond. Similar signal triggering effect by an O-H group is also confirmed. Our study suggests that the Tar chemoeffector binding pocket may be separated into two functional regions: region I mainly contributes to binding and region II contributes to both binding and signaling. This scenario of binding and signaling suggests that Tar may be rationally designed to respond to a nonnative ligand by altering key residues in region I to strengthen binding with the novel ligand while maintaining the key interactions in region II for signaling. Following this strategy, we have successfully redesigned Tar to respond to l-arginine, a basic amino acid that does not have chemotactic effect for WT Tar, by two site-specific mutations (R69'E and R73'E).

Pathway-based mean-field model for Escherichia coli chemotaxis.

We develop a mean-field theory for Escherichia coli chemotaxis based on the coupled spatiotemporal dynamics of the cell population and the mean receptor methylation level field. This multiscale model connects the cells' population level motility behavior with the molecular level pathway dynamics. It reveals a simple scaling dependence of the chemotaxis velocity on the adaptation rate in exponential gradients. It explains the molecular origin of a maximum chemotaxis velocity. Simulations of our model in various spatiotemporal stimuli profiles show quantitative agreements with experiments. Moreover, it predicts a surprising reversal of chemotaxis group velocity in traveling wave environments. Our approach may be used to bridge molecular level pathway dynamics with cellular behavior in other biological systems.

Frequency-dependent Escherichia coli chemotaxis behavior.

We study Escherichia coli chemotaxis behavior in environments with spatially and temporally varying attractant sources by developing a unique microfluidic system. Our measurements reveal a frequency-dependent chemotaxis behavior. At low frequency, the E. coli population oscillates in synchrony with the attractant. In contrast, in fast-changing environments, the population response becomes smaller and out of phase with the attractant waveform. These observations are inconsistent with the well-known Keller-Segel chemotaxis equation. A new continuum model is proposed to describe the population level behavior of E. coli chemotaxis based on the underlying pathway dynamics. With the inclusion of a finite adaptation time and an attractant consumption rate, our model successfully explains the microfluidic experiments at different stimulus frequencies.

A parallel diffusion-based microfluidic device for bacterial chemotaxis analysis.

We developed a multiple-channel microfluidic device for bacterial chemotaxis detection. Some characteristics such as easy operation, parallel sample adding design and fast result readout make this device convenient for most biology labs. The characteristic feature of the design is the agarose gel channels, which serve as a semi-permeable membrane. They can stop the fluid flow and prevent bacteria getting across, but permit the diffusion of small molecules. In the device fabrication process a novel thermal-based method was used to control the shape of agarose gel in the microfluidic channel. The chemical gradient is established by diffusion which can be precisely controlled and measured. Combined with an 8-channel pipette, different attractants, repellent chemicals or different bacteria were analyzed by a two step operation with a readout time of one hour. This device may be useful in the high throughput detection of chemotaxis related molecules and genes.