A Molecular Code for Identity in the Vomeronasal System.

In social interactions among mammals, individuals are recognized by olfactory cues, but identifying the key signals among thousands of compounds remains a major challenge. To address this need, we developed a new technique, component-activity matching (CAM), to select candidate ligands that "explain" patterns of bioactivity across diverse complex mixtures. Using mouse urine from eight different sexes and strains, we identified 23 components to explain firing rates in seven of eight functional classes of vomeronasal sensory neurons. Focusing on a class of neurons selective for females, we identified a novel family of vomeronasal ligands, steroid carboxylic acids. These ligands accounted for much of the neuronal activity of urine from some female strains, were necessary for normal levels of male investigatory behavior of female scents, and were sufficient to trigger mounting behavior. CAM represents the first step toward an exhaustive characterization of the molecular cues for natural behavior in a mammalian olfactory system.

The Accessory Olfactory System: Innately Specialized or Microcosm of Mammalian Circuitry?

In mammals, the accessory olfactory system is a distinct circuit that has received attention for its role in detecting and responding to pheromones. While the neuroscientific investigation of this system is comparatively new, recent advances and its compact size have made it an attractive model for developing an end-to-end understanding of such questions as regulation of essential behaviors, plasticity, and individual recognition. Recent discoveries have indicated a need to reevaluate our conception of this system, suggesting that ( a) physical principles-rather than biological necessity-play an underappreciated role in its raison d'être and that ( b) the anatomy of downstream projections is not dominated by unique specializations but instead consists of an abbreviated cortical/basal ganglia motif reminiscent of other sensorimotor systems. These observations suggest that the accessory olfactory system distinguishes itself primarily by the physicochemical properties of its ligands, but its architecture is otherwise a microcosm of mammalian neurocircuitry.

Julia for biologists.

Major computational challenges exist in relation to the collection, curation, processing and analysis of large genomic and imaging datasets, as well as the simulation of larger and more realistic models in systems biology. Here we discuss how a relative newcomer among programming languages-Julia-is poised to meet the current and emerging demands in the computational biosciences and beyond. Speed, flexibility, a thriving package ecosystem and readability are major factors that make high-performance computing and data analysis available to an unprecedented degree. We highlight how Julia's design is already enabling new ways of analyzing biological data and systems, and we provide a list of resources that can facilitate the transition into Julian computing.

Sensory experience remodels genome architecture in neural circuit to drive motor learning.

Neuronal-activity-dependent transcription couples sensory experience to adaptive responses of the brain including learning and memory. Mechanisms of activity-dependent gene expression including alterations of the epigenome have been characterized1-8. However, the fundamental question of whether sensory experience remodels chromatin architecture in the adult brain in vivo to induce neural code transformations and learning and memory remains to be addressed. Here we use in vivo calcium imaging, optogenetics and pharmacological approaches to show that granule neuron activation in the anterior dorsal cerebellar vermis has a crucial role in a delay tactile startle learning paradigm in mice. Of note, using large-scale transcriptome and chromatin profiling, we show that activation of the motor-learning-linked granule neuron circuit reorganizes neuronal chromatin including through long-distance enhancer-promoter and transcriptionally active compartment interactions to orchestrate distinct granule neuron gene expression modules. Conditional CRISPR knockout of the chromatin architecture regulator cohesin in anterior dorsal cerebellar vermis granule neurons in adult mice disrupts enhancer-promoter interactions, activity-dependent transcription and motor learning. These findings define how sensory experience patterns chromatin architecture and neural circuit coding in the brain to drive motor learning.

Publisher Correction: Sensory experience remodels genome architecture in neural circuit to drive motor learning.

In this Letter, '≥' should be '≤' in the sentence: "Intra-chromosomal reads were further split into short-range reads (≥1 kb) and long-range reads (>1 kb)". This error has been corrected online.An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Circadian pacemaker neurons display cophasic rhythms in basal calcium level and in fast calcium fluctuations.

Circadian pacemaker neurons in the Drosophila brain display daily rhythms in the levels of intracellular calcium. These calcium rhythms are driven by molecular clocks and are required for normal circadian behavior. To study their biological basis, we employed genetic manipulations in conjunction with improved methods of in vivo light-sheet microscopy to measure calcium dynamics in individual pacemaker neurons over complete 24-h durations at sampling frequencies as high as 5 Hz. This technological advance unexpectedly revealed cophasic daily rhythms in basal calcium levels and in high-frequency calcium fluctuations. Further, we found that the rhythms of basal calcium levels and of fast calcium fluctuations reflect the activities of two proteins that mediate distinct forms of calcium fluxes. One is the inositol trisphosphate receptor (ITPR), a channel that mediates calcium fluxes from internal endoplasmic reticulum calcium stores, and the other is a T-type voltage-gated calcium channel, which mediates extracellular calcium influx. These results suggest that Drosophila molecular clocks regulate ITPR and T-type channels to generate two distinct but coupled rhythms in basal calcium and in fast calcium fluctuations. We propose that both internal and external calcium fluxes are essential for circadian pacemaker neurons to provide rhythmic outputs and thereby, regulate the activities of downstream brain centers.

Sensorimotor Coding of Vermal Granule Neurons in the Developing Mammalian Cerebellum.

The vermal cerebellum is a hub of sensorimotor integration critical for postural control and locomotion, but the nature and developmental organization of afferent information to this region have remained poorly understood in vivo Here, we use in vivo two-photon calcium imaging of the vermal cerebellum in awake behaving male and female mice to record granule neuron responses to diverse sensorimotor cues targeting visual, auditory, somatosensory, and motor domains. Use of an activity-independent marker revealed that approximately half (54%) of vermal granule neurons were activated during these recordings. A multikernel linear model distinguished the relative influences of external stimuli and co-occurring movements on neural responses, indicating that, among the subset of activated granule neurons, locomotion (44%-56%) and facial air puffs (50%) were more commonly and reliably encoded than visual (31%-32%) and auditory (19%-28%) stimuli. Strikingly, we also uncover populations of granule neurons that respond differentially to voluntary and forced locomotion, whereas other granule neurons in the same region respond similarly to locomotion in both conditions. Finally, by combining two-photon calcium imaging with birth date labeling of granule neurons via in vivo electroporation, we find that early- and late-born granule neurons convey similarly diverse sensorimotor information to spatially distinct regions of the molecular layer. Collectively, our findings elucidate the nature and developmental organization of sensorimotor information in vermal granule neurons of the developing mammalian brain.SIGNIFICANCE STATEMENT Cerebellar granule neurons comprise over half the neurons in the brain, and their coding properties have been the subject of theoretical and experimental interest for over a half-century. In this study, we directly test long-held theories about encoding of sensorimotor stimuli in the cerebellum and compare the in vivo coding properties of early- and late-born granule neurons. Strikingly, we identify populations of granule neurons that differentially encode voluntary and forced locomotion and find that, although the birth order of granule neurons specifies the positioning of their parallel fiber axons, both early- and late-born granule neurons convey a functionally diverse sensorimotor code. These findings constitute important conceptual advances in understanding the principles underlying cerebellar circuit development and function.

Vasoactive intestinal polypeptide mediates circadian rhythms in mammalian olfactory bulb and olfaction.

Accumulating evidence suggests that the olfactory bulbs (OBs) function as an independent circadian system regulating daily rhythms in olfactory performance. However, the cells and signals in the olfactory system that generate and coordinate these circadian rhythms are unknown. Using real-time imaging of gene expression, we found that the isolated olfactory epithelium and OB, but not the piriform cortex, express similar, sustained circadian rhythms in PERIOD2 (PER2). In vivo, PER2 expression in the OB of mice is circadian, approximately doubling with a peak around subjective dusk. Furthermore, mice exhibit circadian rhythms in odor detection performance with a peak at approximately subjective dusk. We also found that circadian rhythms in gene expression and odor detection performance require vasoactive intestinal polypeptide (VIP) or its receptor VPAC2R. VIP is expressed, in a circadian manner, in interneurons in the external plexiform and periglomerular layers, whereas VPAC2R is expressed in mitral and external tufted cells in the OB. Together, these results indicate that VIP signaling modulates the output from the OB to maintain circadian rhythms in the mammalian olfactory system.

Robust encoding of stimulus identity and concentration in the accessory olfactory system.

Sensory systems represent stimulus identity and intensity, but in the neural periphery these two variables are typically intertwined. Moreover, stable detection may be complicated by environmental uncertainty; stimulus properties can differ over time and circumstance in ways that are not necessarily biologically relevant. We explored these issues in the context of the mouse accessory olfactory system, which specializes in detection of chemical social cues and infers myriad aspects of the identity and physiological state of conspecifics from complex mixtures, such as urine. Using mixtures of sulfated steroids, key constituents of urine, we found that spiking responses of individual vomeronasal sensory neurons encode both individual compounds and mixtures in a manner consistent with a simple model of receptor-ligand interactions. Although typical neurons did not accurately encode concentration over a large dynamic range, from population activity it was possible to reliably estimate the log-concentration of pure compounds over several orders of magnitude. For binary mixtures, simple models failed to accurately segment the individual components, largely because of the prevalence of neurons responsive to both components. By accounting for such overlaps during model tuning, we show that, from neuronal firing, one can accurately estimate log-concentration of both components, even when tested across widely varying concentrations. With this foundation, the difference of logarithms, log A - log B = log A/B, provides a natural mechanism to accurately estimate concentration ratios. Thus, we show that a biophysically plausible circuit model can reconstruct concentration ratios from observed neuronal firing, representing a powerful mechanism to separate stimulus identity from absolute concentration.

Reliable sex and strain discrimination in the mouse vomeronasal organ and accessory olfactory bulb.

Animals modulate their courtship and territorial behaviors in response to olfactory cues produced by other animals. In rodents, detecting these cues is the primary role of the accessory olfactory system (AOS). We sought to systematically investigate the natural stimulus coding logic and robustness in neurons of the first two stages of accessory olfactory processing, the vomeronasal organ (VNO) and accessory olfactory bulb (AOB). We show that firing rate responses of just a few well-chosen mouse VNO or AOB neurons can be used to reliably encode both sex and strain of other mice from cues contained in urine. Additionally, we show that this population code can generalize to new concentrations of stimuli and appears to represent stimulus identity in terms of diverging paths in coding space. Together, the results indicate that firing rate code on the temporal order of seconds is sufficient for accurate classification of pheromonal patterns at different concentrations and may be used by AOS neural circuitry to discriminate among naturally occurring urine stimuli.

STARDUST: a pipeline for the unbiased analysis of astrocyte regional calcium dynamics.

Calcium imaging has become a popular way to probe astrocyte activity, but few analysis methods holistically capture discrete calcium signals that occur across the astrocyte domain. Here, we introduce STARDUST, a pipeline for the Spatio-Temporal Analysis of Regional Dynamics & Unbiased Sorting of Transients from fluorescence recordings of astrocytes, and provide step-by-step guidelines. STARDUST yields fluorescence time-series from data-defined regions of activity and performs systematic signal detection and feature extraction, enabling the in-depth and unbiased study of astrocyte calcium signals.

The Representation of Decision Variables in Orbitofrontal Cortex is Longitudinally Stable.

The computation and comparison of subjective values underlying economic choices rely on the orbitofrontal cortex (OFC). In this area, distinct groups of neurons encode the value of individual options, the binary choice outcome, and the chosen value. These variables capture both the input and the output of the choice process, suggesting that the cell groups found in OFC constitute the building blocks of a decision circuit. Here we show that this neural circuit is longitudinally stable. Using two-photon calcium imaging, we recorded from mice choosing between different juice flavors. Recordings of individual cells continued for up to 20 weeks. For each cell and each pair of sessions, we compared the activity profiles using cosine similarity, and we assessed whether the cell encoded the same variable in both sessions. These analyses revealed a high degree of stability and a modest representational drift. A quantitative estimate indicated this drift would not randomize the circuit within the animal's lifetime.

Organization of vomeronasal sensory coding revealed by fast volumetric calcium imaging.

A long-standing goal in neuroscience is to perform exhaustive recording of each neuron in a functional local circuit. To achieve this goal, one promising approach is optical imaging of fluorescent calcium indicators, but typically the tens or hundreds of cells imaged simultaneously comprise only a tiny percentage of the neurons in an intact circuit. Here, we show that a recent innovation, objective-coupled planar illumination (OCPI) microscopy, permits simultaneous recording from three-dimensional volumes containing many thousand neurons. We used OCPI microscopy to record chemosensory responses in the mouse vomeronasal epithelium, for which expression of hundreds of receptor types implies high functional diversity. The implications of this diversity for sensory coding were examined using several classes of previously reported vomeronasal ligands, including sulfated steroids. A collection of just 12 sulfated steroids activated more than a quarter of the neurons in the apical vomeronasal epithelium; unexpectedly, responses were functionally organized into a modest number of classes with characteristic spatial distribution. Recording from a whole sensory system thus revealed new organizational principles.

Mapping a complete neural population in the retina.

Recording simultaneously from essentially all of the relevant neurons in a local circuit is crucial to understand how they collectively represent information. Here we show that the combination of a large, dense multielectrode array and a novel, mostly automated spike-sorting algorithm allowed us to record simultaneously from a highly overlapping population of >200 ganglion cells in the salamander retina. By combining these methods with labeling and imaging, we showed that up to 95% of the ganglion cells over the area of the array were recorded. By measuring the coverage of visual space by the receptive fields of the recorded cells, we concluded that our technique captured a neural population that forms an essentially complete representation of a region of visual space. This completeness allowed us to determine the spatial layout of different cell types as well as identify a novel group of ganglion cells that responded reliably to a set of naturalistic and artificial stimuli but had no measurable receptive field. Thus, our method allows unprecedented access to the complete neural representation of visual information, a crucial step for the understanding of population coding in sensory systems.

Polyphasic circadian neural circuits drive differential activities in multiple downstream rhythmic centers.

Circadian clocks align various behaviors such as locomotor activity, sleep/wake, feeding, and mating to times of day that are most adaptive. How rhythmic information in pacemaker circuits is translated to neuronal outputs is not well understood. Here, we used brain-wide, 24-h in vivo calcium imaging in the Drosophila brain and searched for circadian rhythmic activity among identified clusters of dopaminergic (DA) and peptidergic neurosecretory (NS) neurons. Such rhythms were widespread and imposed by the PERIOD-dependent clock activity within the ∼150-cell circadian pacemaker network. The rhythms displayed either a morning (M), evening (E), or mid-day (MD) phase. Different subgroups of circadian pacemakers imposed neural activity rhythms onto different downstream non-clock neurons. Outputs from the canonical M and E pacemakers converged to regulate DA-PPM3 and DA-PAL neurons. E pacemakers regulate the evening-active DA-PPL1 neurons. In addition to these canonical M and E oscillators, we present evidence for a third dedicated phase occurring at mid-day: the l-LNv pacemakers present the MD activity peak, and they regulate the MD-active DA-PPM1/2 neurons and three distinct NS cell types. Thus, the Drosophila circadian pacemaker network is a polyphasic rhythm generator. It presents dedicated M, E, and MD phases that are functionally transduced as neuronal outputs to organize diverse daily activity patterns in downstream circuits.

A public confession: the retina trumpets its failed predictions.

To compensate for delays of phototransduction, the retina anticipates the future by extrapolating the position of a moving object. But what if the object's motion changes, and the extrapolation is wrong? In this issue of Neuron, Schwartz and colleagues show that these prediction failures trigger a large burst of firing that helps to rapidly correct the neural representation of the object's new position.

Chemosensory burst coding by mouse vomeronasal sensory neurons.

The capabilities of any sensory system are ultimately constrained by the properties of the sensory neurons: the ability to detect and represent stimuli is limited by noise due to spontaneous activity, and optimal decoding in downstream circuitry must be matched to the nature of the encoding performed at the input. Here, we investigated the firing properties of sensory neurons in the accessory olfactory system, a distinct sensory system specialized for detection of socially relevant odors. Using multielectrode array recording, we observed that sensory neurons are spontaneously active and highly variable across time and trials and that this spontaneous activity limits the ability to distinguish sensory responses from noise. Sensory neuron activity tended to consist of bursts that maintained remarkably consistent statistics during both spontaneous activity and in response to stimulation with sulfated steroids. This, combined with pharmacological and genetic intervention in the signal transduction cascade, indicates that sensory transduction plays a role in shaping overall spontaneous activity. These findings indicate that as-yet unexplored characteristics of the sensory transduction cascade significantly constrain the representation of sensory information by vomeronasal neurons.

Sex- and strain-specific expression and vomeronasal activity of mouse ESP family peptides.

Male mice secrete exocrine-gland-secreting peptide 1 (ESP1) from the extraorbital lacrimal gland into tear fluid [1]. Other mice detect ESP1 through sensory neurons in the vomeronasal organ (VNO), a secondary olfactory system that senses pheromonal information, including sex, strain, and species. ESP1 is now known to be a member of a multigene family that encodes peptides of various lengths. We herein performed genomic and expression analyses of the ESP family. The ESP family consists of 38 members in mice and 10 members in rat but is absent from the human genome, suggesting rapid molecular evolution. In addition to the male-specific ESP1, we discovered one, which we designated ESP36, that, in adult BALB/c mice, is expressed only in the female extraorbital lacrimal gland. The sexually dimorphic expression is ensured by the release of testosterone after puberty. However, we observed dramatic differences in the expression levels of ESPs between strains. Finally, all ESPs elicited an electrical response in the vomeronasal epithelium but not in the main olfactory epithelium. Multielectrode recording of VNO activity demonstrated that ESP1 induces action potentials in vomeronasal neurons, leading to an increase in the spike firing rate, and that ESP1 is recognized by narrowly tuned vomeronasal sensory neurons. Sexual dimorphism and strain differences of ESPs and their reception in the VNO suggest that the ESP family can convey information about sex and individual identity via the vomeronasal system. The chemosensation of this nonvolatile peptide family by direct contact appears to be one of strategies for sociosexual communication in rodent species.