Electrogenetic signaling and information propagation for controlling microbial consortia via programmed lysis.

To probe signal propagation and genetic actuation in microbial consortia, we have coopted the components of both redox and quorum sensing (QS) signaling into a communication network for guiding composition by "programming" cell lysis. Here, we use an electrode to generate hydrogen peroxide as a redox cue that determines consortia composition. The oxidative stress regulon of Escherichia coli, OxyR, is employed to receive and transform this signal into a QS signal that coordinates the lysis of a subpopulation of cells. We examine a suite of information transfer modalities including "monoculture" and "transmitter-receiver" models, as well as a series of genetic circuits that introduce time-delays for altering information relay, thereby expanding design space. A simple mathematical model aids in developing communication schemes that accommodate the transient nature of redox signals and the "collective" attributes of QS signals. We suggest this platform methodology will be useful in understanding and controlling synthetic microbial consortia for a variety of applications, including biomanufacturing and biocontainment.

MEGF10 and MEGF11 mediate homotypic interactions required for mosaic spacing of retinal neurons.

In many parts of the nervous system, neuronal somata display orderly spatial arrangements. In the retina, neurons of numerous individual subtypes form regular arrays called mosaics: they are less likely to be near neighbours of the same subtype than would occur by chance, resulting in 'exclusion zones' that separate them. Mosaic arrangements provide a mechanism to distribute each cell type evenly across the retina, ensuring that all parts of the visual field have access to a full set of processing elements. Remarkably, mosaics are independent of each other: although a neuron of one subtype is unlikely to be adjacent to another of the same subtype, there is no restriction on its spatial relationship to neighbouring neurons of other subtypes. This independence has led to the hypothesis that molecular cues expressed by specific subtypes pattern mosaics by mediating homotypic (within-subtype) short-range repulsive interactions. So far, however, no molecules have been identified that show such activity, so this hypothesis remains untested. Here we demonstrate in mouse that two related transmembrane proteins, MEGF10 and MEGF11, have critical roles in the formation of mosaics by two retinal interneuron subtypes, starburst amacrine cells and horizontal cells. MEGF10 and 11 and their invertebrate relatives Caenorhabditis elegans CED-1 and Drosophila Draper have hitherto been studied primarily as receptors necessary for engulfment of debris following apoptosis or axonal injury. Our results demonstrate that members of this gene family can also serve as subtype-specific ligands that pattern neuronal arrays.

XPORT ENTRAP: A droplet microfluidic platform for enhanced DNA transfer between microbial species.

A significant hurdle for the widespread implementation and use of synthetic biology is the challenge of highly efficient introduction of DNA into microorganisms. This is especially a barrier for the utilization of non-model organisms and/or novel chassis species for a variety of applications, ranging from molecular biology to biotechnology and biomanufacturing applications. Common approaches to episomal and chromosomal gene editing, which employ techniques such as chemical competence and electroporation, are typically only amenable to a small subset of microbial species while leaving the vast majority of microorganisms in nature genetically inaccessible. To address this challenge, we have employed the previously described B. subtilis broad-host conjugation strain, XPORT, which was modularly designed for loading DNA cargo and conjugating such DNA into recalcitrant microbes. In this current work, we have leveraged and adapted the XPORT strain for use in a droplet microfluidic platform to enable increased efficiency of conjugation-based DNA transfer. The system named DNA ENTRAP (DNA ENhanced TRAnsfer Platform) utilizes cell-encapsulated water-in-oil emulsion droplets as pico-liter-volume bioreactors that allows controlled contacts between the donor and receiver cells within the emulsion bioreactor. This allowed enhanced XPORT-mediated genetic transfer over the current benchtop XPORT process, demonstrated using two different Bacillus subtilis strains (donor and receiver), as well as increased throughput (e.g., number of successfully conjugated cells) due to the automated assay steps inherent to microfluidic lab-on-a-chip systems. DNA ENTRAP paves the way for a streamlined automation of culturing and XPORT-mediated genetic transfer processes as well as future high-throughput cell engineering and screening applications.

Retinal ganglion cells with distinct directional preferences differ in molecular identity, structure, and central projections.

The retina contains ganglion cells (RGCs) that respond selectively to objects moving in particular directions. Individual members of a group of ON-OFF direction-selective RGCs (ooDSGCs) detect stimuli moving in one of four directions: ventral, dorsal, nasal, or temporal. Despite this physiological diversity, little is known about subtype-specific differences in structure, molecular identity, and projections. To seek such differences, we characterized mouse transgenic lines that selectively mark ooDSGCs preferring ventral or nasal motion as well as a line that marks both ventral- and dorsal-preferring subsets. We then used the lines to identify cell surface molecules, including Cadherin 6, CollagenXXVα1, and Matrix metalloprotease 17, that are selectively expressed by distinct subsets of ooDSGCs. We also identify a neuropeptide, CART (cocaine- and amphetamine-regulated transcript), that distinguishes all ooDSGCs from other RGCs. Together, this panel of endogenous and transgenic markers distinguishes the four ooDSGC subsets. Patterns of molecular diversification occur before eye opening and are therefore experience independent. They may help to explain how the four subsets obtain distinct inputs. We also demonstrate differences among subsets in their dendritic patterns within the retina and their axonal projections to the brain. Differences in projections indicate that information about motion in different directions is sent to different destinations.

Social functioning and brain imaging in individuals at clinical high-risk for psychosis: A systematic review.

Impairments in social functioning are a core impairment in psychosis and are associated with poor outcomes. These deficits are found in those at clinical high-risk (CHR) for psychosis, and can persist even in the absence of transition. However, the neurobiological underpinnings of social functioning remain unclear, therefore we conducted a systematic review of brain metrics that have been associated with social functioning in youth at CHR for psychosis. Five databases (MEDLINE, CINAHL, EBM reviews, Embase, and PsycINFO) were searched from inception to May 5, 2020. Studies were selected if they examined brain imaging, and social functioning in youth at CHR for psychosis. Of the 9629 citations found through online database searching, 12 studies with 696 CHR participants met inclusion criteria. Too few studies were focused on the same brain region using the same methodology to perform a meta-analysis, however, loci within the prefrontal cortex were most often associated with social functioning. Few studies have linked social functioning to brain imaging metrics, suggesting that future work should focus on this relationship.

Adult-born neurons facilitate olfactory bulb pattern separation during task engagement.

The rodent olfactory bulb incorporates thousands of newly generated inhibitory neurons daily throughout adulthood, but the role of adult neurogenesis in olfactory processing is not fully understood. Here we adopted a genetic method to inducibly suppress adult neurogenesis and investigated its effect on behavior and bulbar activity. Mice without young adult-born neurons (ABNs) showed normal ability in discriminating very different odorants but were impaired in fine discrimination. Furthermore, two-photon calcium imaging of mitral cells (MCs) revealed that the ensemble odor representations of similar odorants were more ambiguous in the ablation animals. This increased ambiguity was primarily due to a decrease in MC suppressive responses. Intriguingly, these deficits in MC encoding were only observed during task engagement but not passive exposure. Our results indicate that young olfactory ABNs are essential for the enhancement of MC pattern separation in a task engagement-dependent manner, potentially functioning as a gateway for top-down modulation.

Lack of Pattern Separation in Sensory Inputs to the Olfactory Bulb during Perceptual Learning.

Recent studies revealed changes in odor representations in the olfactory bulb during active olfactory learning (Chu et al., 2016; Yamada et al., 2017). Specifically, mitral cell ensemble responses to very similar odorant mixtures sparsened and became more distinguishable as mice learned to discriminate the odorants over days (Chu et al., 2016). In this study, we explored whether changes in the sensory inputs to the bulb underlie the observed changes in mitral cell responses. Using two-photon calcium imaging to monitor the odor responses of the olfactory sensory neuron (OSN) axon terminals in the glomeruli of the olfactory bulb during a discrimination task, we found that OSN inputs to the bulb are stable during discrimination learning. During one week of training to discriminate between very similar odorant mixtures in a Go/No-go task, OSN responses did not show significant sparsening, and the responses to the trained similar odorants did not diverge throughout training. These results suggest that the adaptive changes of mitral cell responses during perceptual learning are ensured by mechanisms downstream of OSN input, possibly in local circuits within olfactory bulb.

Balancing the Robustness and Efficiency of Odor Representations during Learning.

For reliable stimulus identification, sensory codes have to be robust by including redundancy to combat noise, but redundancy sacrifices coding efficiency. To address how experience affects the balance between the robustness and efficiency of sensory codes, we probed odor representations in the mouse olfactory bulb during learning over a week, using longitudinal two-photon calcium imaging. When mice learned to discriminate between two dissimilar odorants, responses of mitral cell ensembles to the two odorants gradually became less discrete, increasing the efficiency. In contrast, when mice learned to discriminate between two very similar odorants, the initially overlapping representations of the two odorants became progressively decorrelated, enhancing the robustness. Qualitatively similar changes were observed when the same odorants were experienced passively, a condition that would induce implicit perceptual learning. These results suggest that experience adjusts odor representations to balance the robustness and efficiency depending on the similarity of the experienced odorants.

Dynamic sensory representations in the olfactory bulb: modulation by wakefulness and experience.

How are sensory representations in the brain influenced by the state of an animal? Here we use chronic two-photon calcium imaging to explore how wakefulness and experience shape odor representations in the mouse olfactory bulb. Comparing the awake and anesthetized state, we show that wakefulness greatly enhances the activity of inhibitory granule cells and makes principal mitral cell odor responses more sparse and temporally dynamic. In awake mice, brief repeated odor experience leads to a gradual and long-lasting (months) weakening of mitral cell odor representations. This mitral cell plasticity is odor specific, recovers gradually over months, and can be repeated with different odors. Furthermore, the expression of this experience-dependent plasticity is prevented by anesthesia. Together, our results demonstrate the dynamic nature of mitral cell odor representations in awake animals, which is constantly shaped by recent odor experience.

Neurod6 expression defines new retinal amacrine cell subtypes and regulates their fate.

Most regions of the CNS contain many subtypes of inhibitory interneurons with specialized roles in circuit function. In the mammalian retina, the ∼30 subtypes of inhibitory interneurons called amacrine cells (ACs) are generally divided into two groups: wide/medium-field GABAergic ACs and narrow-field glycinergic ACs, which mediate lateral and vertical interactions, respectively, within the inner plexiform layer. We used expression profiling and mouse transgenic lines to identify and characterize two closely related narrow-field AC subtypes. Both arise postnatally and one is neither glycinergic nor GABAergic (nGnG). Two transcription factors selectively expressed by these subtypes, Neurod6 and special AT-rich-sequence-binding protein 2 (Satb2), regulate a postmitotic cell fate choice between these subtypes. Satb2 induces Neurod6, which persists in nGnG ACs and promotes their fate but is downregulated in the related glycinergic AC subtype. Our results support the view that cell fate decisions made in progenitors and their progeny act together to diversify ACs.