Biology subjects

Anderson, D. J.

Publications and source records attributed to Anderson, D. J..

At least 19 recordsLinked to original sources

bioRxiv2026

The Human Contraceptive Antibody Targets A Semenogelin Galectin-3 N-Linked Glycan Complex on the Sperm Surface

The human contraception antibody (HCA) is a non-hormonal topical contraceptive entering Phase II clinical trials that functions by agglutinating and immobilizing sperm. To date, the HCA epitope has been described as an N-linked polylactosamine glycan motif on CD52, but this assignment has been inferred only by colocalization with anti-CD52 and S19 antibodies in western blots. Here, we report that HCA may instead bind semenogelin-galectin-3 complexes associated with the sperm surface. Semenogelins are gel forming, highly glycosylated proteins, with molecular weights of 50-70 kDa that are cleaved into smaller segments that adhere to sperm during semen liquefaction. In this study, we isolated the HCA antigen from seminal plasma and sperm using an HCA affinity column and identified semenogelin by mass spectrometry. Western blot analysis of seminal plasma showed that HCA binds to a large smear of proteins with a higher apparent molecular mass than CD52, and that these may correspond to semenogelin sequences. Flow cytometric analysis of enzymatically deglycosylated sperm demonstrated that HCA binding requires sperm surface N-linked glycans. However, despite this apparent glycan dependence, HCA failed to bind any glycans represented on a comprehensive glycan microarray. In addition to semenogelin, Galectin-3 was identified in the HCA purified sperm lysates, supporting a model in which HCA recognizes semenogelin-derived glycopeptides that are tethered to the sperm surface via a galectin mediated interaction with an N-linked glycan. We found that O-linked glycan modifications on the semenogelin proteins contain LacNAc motifs that can bind to galectins. Finally, we found that depletion of galectins bound to the sperm surface by competitive inhibitors reduced HCA-mediated agglutination, and this could be rescued by addition of recombinant galectin-3/semenogelin-2 complexes to sperm cultures. Collectively, our findings suggest that HCA's binding site involves a semenogelin-galectin-3 complex anchored to the sperm surface via an N-linked glycan.

immunology↗

bioRxiv2026

Widespread introgression and a potential role for a neo-sex chromosome in booby diversification and speciation

Speciation in marine taxa is dynamic and complex, often occurring in the absence of absolute geographic barriers to gene flow. Boobies comprise a clade of seven highly mobile seabird species with high diversity in the eastern Pacific Ocean, where no land barriers separate extant sister species and hybridization is widespread. They exhibit striking diversity in bare-part coloration, an important signal of mate quality, and possess a novel multiple sex chromosome system (Z1Z2Z1Z2 / Z1Z2W), likely resulting from a fusion between an ancestral autosomal microchromosome and the W chromosome. To understand speciation processes in the context of gene flow, we sequenced and analyzed 29 short-read booby genomes and assembled a reference northern gannet genome to (1) test for introgression, (2) characterize genomic patterns of divergence across species, and (3) investigate the gene content and evolution of the neo-sex chromosome. We found that divergence among eastern Pacific sister taxa is temporally associated with periods of glacial maxima. Between periods of glacial maxima, two pairs of sister species (blue-footed and Peruvian boobies; masked and Nazca boobies) exhibit strong signatures of episodic introgression, indicating that speciation has occurred over multiple periods of divergence followed by gene flow. However, in the third pair of sister species, genomic signatures showed higher divergence within brown booby populations than between brown and Cocos boobies. We also identified the epidermal differentiation complex - a gene cassette involved in skin, feather, beak, and claw development - on the neo-sex region of the W chromosome, where it spans the boundary between the putatively recombining and non-recombining regions, with interspecific variation in the exact recombination suppression boundary. We inferred that this complex may contribute to diversity in feather and bare part coloration across boobies. Together, these results reveal that diversification and speciation in a clade of highly mobile seabirds emerged from a complex evolutionary history involving historical climate dynamics, introgression, and sex chromosome evolution.

evolutionary biology↗

bioRxiv2026

Complement protein concentrations and activity in human cervical mucus

ProblemComplement, a system of over 30 interacting proteins, functions as a critical immune mediator at mucosal surfaces including the intestine, airway, and nasal mucosae, where it orchestrates complement-dependent cytotoxicity (CDC), complement-dependent phagocytosis (CDP), and inflammatory responses. While complement components have been detected at low levels in genital tract fluids, including cervical mucus, the physiologic dynamics of complement in the female reproductive tract remain poorly characterized. Notably, systematic quantification of complement component levels across the menstrual cycle has not been conducted, limiting our understanding of how hormonal fluctuations may influence complement-mediated protection at the vaginal mucosa. Method of studyTen healthy women of reproductive age were recruited to provide paired samples of cervical mucus (CM) and serum (S) during each phase of the menstrual cycle: follicular, ovulatory, and luteal. Samples were analyzed using bead-based multiplex assays to quantify 13 primary complement components. ResultsAll 13 complement proteins tested were detectable in CM and S; C3b/iC3b was the predominant complement component in CM followed by C4 and C3. In contrast, C4 was the dominant component in S followed by C1q and C3. There were significantly higher levels of C2 in CM during the follicular phase of the menstrual cycle vs. the ovulatory phase (1.15{+/-}0.80 vs 0.24{+/-}0.22{micro}g/mL), C4b (0.56{+/-}0.35 vs 0.22{+/-}0.36 {micro}g/mL), C5a (1.45{+/-}1.09 vs 0.38{+/-}0.48 {micro}g/mL) In contrast, no differences were found in serum complement levels were during the menstrual cycle. Complement concentrations were on average 308-fold (median= 84) lower in CM than in S, except for C3b/iC3b which was only 5 to 7.5-fold lower in CM, and C2 which was higher in CM at the luteal and follicular phases. Midcycle cervical mucus was capable of inducing complement-mediated hemolysis at about 1/3 the potency of serum. ConclusionCervical mucus contains detectable and functional levels of complement proteins. These normative values can provide a foundation for future studies on immune mechanisms in the FRT.

immunology↗

bioRxiv2025

Neural control of coordinated wing and leg movements during a terrestrial threat display

Threat displays are a common form of social communication. In flying species, such displays often involve stereotypical, choreographed wing and leg movements. How the brain coordinates characteristic movements of these appendages to transmit social signals is poorly understood. In male Drosophila, threat displays flexibly combine wing displays with rapid turns and charges. Here we identify two brain modules downstream of a superordinate threat command center. Each contains two neurons that combinatorially generate threat-specific appendicular movements: one comprises two descending neurons synergistically controlling wing threat; the other contains two interneurons that antagonistically control turns and charges. Within-module neuronal coactivation evokes appendage-specific actions recapitulating those evoked by the upstream center. These data uncover a hierarchical combinatorial circuit that coordinates wing and leg movements during a terrestrial social display. More generally, our findings identify an instantiation of a Tinbergenian hierarchical behavioral control system for social communication and reveal a novel underlying implementation logic.

neuroscience↗

bioRxiv2025

A line attractor maintains aggressiveness during feeding in hangry mice

Aggression evolved to protect resources such as food from competitors, but animals must balance fighting and feeding so that they facilitate rather than hinder re-establishment of energy homeostasis. How this balancing is computed is not well understood. We have approached this problem at the level of neural population-coding by examining the effect of progressive starvation on a hypothalamic line attractor that encodes an internal state of aggressiveness. Moderate fasting yielded "hangry" mice, decreasing attack latency and increasing attack frequency. In parallel, line attractor ramping rate and stability were increased, suggesting that hunger enhances aggressiveness by modifying neural dynamics. In contrast, prolonged starvation inhibited aggression and eliminated the line attractor. In satiated mice, titrated acute chemogenetic activation of arcuate AgRP neurons recapitulated the biphasic effects of progressive starvation, suggesting that a continuous increase in hunger exerts bi-directional influences at different intensities. When confronted with food and an intruder, hangry mice alternated between feeding and fighting. During eating, population neural activity moved out of the line attractor while activity in the attractor dimension remained unchanged. Following feeding, activity rapidly relaxed back into the attractor and aggression resumed. Thus, the line attractor may serve to keep hungry animals primed for aggression during intermittent feeding bouts.

animal behavior and cognition↗

bioRxiv2025

Natural evolution of intermale sexual behavior by multiple pheromone switches among Drosophila species

We have identified a Drosophila species which exhibits spontaneous and robust intermale sexual behavior. D. santomea males distinguish conspecific sexes but court both vigorously and seldom attack. Elevated intermale courtship stems from at least three evolutionarily derived pheromonal changes. In males, the sexually monomorphic cuticular pheromone 7-tricosene promotes rather than inhibits courtship and the courtship-inhibiting olfactory pheromone cVA is reduced 84-92% compared to close relatives. The third switch is in D. santomea females, where cVA suppresses rather than promotes sexual receptivity. Female cVA aversion and male cVA reduction may have co-evolved to maintain efficient intraspecific mating but prevent hybridization with the sympatric sibling species D. yakuba. High intermale courtship and low cVA also co-occur and appear selectively derived in a distant monomorphic species D. persimilis, implying pheromonal and social behavioral convergence. Changes in pheromone valence and levels may therefore explain the recent evolutionary emergence of intermale sexual behavior in Drosophila.

evolutionary biology↗

bioRxiv2025

Revolutionizing male contraception: Personal lubricants as a novel way to deliver the Human Contraception Antibody

Study QuestionCan personal lubricants effectively deliver the sperm-agglutinating Human Contraception Antibody (HCA) to achieve on-demand male contraception? Summary AnswerThis study demonstrates that several water-based lubricants can effectively deliver bioactive HCA, and that a dimethicone-containing silicone lubricant can be modified into a stable emulsion suitable for antibody delivery. What Is Known AlreadyThe HCA-based vaginal film ZB-06 was shown to be safe and effective in a Phase I clinical trial for female contraception. Male contraceptive options remain limited. Using sexual lubricants as a delivery platform for HCA represents a novel and practical approach to male contraception. Study Design, Size, DurationWe tested the stability of HCA for up to two years in a variety of commercially available sexual lubricants, and its delivery from a penile device using a simulated intercourse model. Participants/Materials, Setting, MethodsFourteen commercial lubricants were initially screened for impact on sperm motility and viability, miscibility with HCA solution, and preservation of HCA activity (sperm agglutination test). From these, three water-based and one silicone-based lubricant were selected for further study. The silicone-based product was engineered into a novel emulsion (KY-DE) by incorporating 6% w/w PEG-10 dimethicone and 0.005% v/v TWEEN-20 for HCA integration. Formulations were tested for HCA functional stability and contraceptive efficacy over time using kinetic sperm agglutination assays and a capillary tube sperm penetration test. A novel simulated intercourse model was developed using a 3D-printed penile device and a vaginal analog (Fleshlight(R)). HCA-lube was applied to the "penis", and concentrations of HCA delivered to the "vagina" during intercourse were measured by ELISA and sperm agglutination assay. Safety evaluations were conducted using a vaginal tissue model (EpiVaginal, VEC-100-FT). Viability (MTT assay), tight junction integrity (TEER), and cytokine release were assessed after application of HCA-lube. Main Results and the Role of ChanceAll selected lubricants were nontoxic and did not affect HCAs ability to agglutinate sperm. In the capillary tube sperm penetration assay, HCA-formulated lubricants significantly impaired sperm penetration; few motile sperm penetrated past a depth of 1 cm after 90 minutes in the HCA-lubricants vs >80_in most of the untreated lubricants. Importantly, the simulated intercourse assay confirmed successful delivery of HCA to the vagina. Water-based lubricants delivered an average of 24 {+/-} 8.4 {micro}g/mL of HCA, the commercial silicone lubricant 11 {+/-} 5.2 {micro}g/mL, and the KY-DE emulsion 27.4 {+/-} 22.4 {micro}g/mL. Futhermore, the formulations did not compromise tissue integrity or viability, or induce significant inflammatory responses. Large Scale DataNot applicable. Limitations, Reasons for CautionWhile these in vitro results support the feasibility of using lubricants as HCA delivery vehicles, translation into clinical application requires further validation and clinical trials. Wider Implications of the FindingsThis work presents a new direction in male contraceptive development, leveraging the widespread use of personal lubricants to introduce antibody-based, reversible, on-demand contraception. Engineering a silicone-based lubricant that retains HCA activity under varied conditions and ensures higher mucosal delivery could further advance this approach. Study Funding/Competing Interest(s)This work was supported by a Sokol Grant from the Male Contraceptive Initiative, a Sexual Medicine Grant from Boston University Chobanian & Avedisian School of Medicine, and grant P50HD096957 from the National Institutes of Health. Kevin Whaley (ZabBio) intends to commercialize HCA for contraception. None of the other authors have competing interests.

bioengineering↗

bioRxiv2025

Female sexual agency and frequent extra-pair copulations, but no extra-pair paternity, in Nazca boobies (Sula granti)

Extra-pair copulations (EPCs) are the poorly known antecedents of extra-pair fertilizations (EPFs) in birds. EPFs occur in most bird species that have been examined, but sexual conflict will generally reward females hiding their EPCs from males attempting to protect their paternity. EPCs will be difficult for researchers to document, and necessarily underestimated, in that case. We measured EPC behaviors and EPF frequency in a colonial seabird, the Nazca booby Sula granti, in which all copulations occur in a visually open setting with numerous possible copulatory partners readily available. Females are larger and more physically powerful than males, and are the numerically limiting sex, perhaps limiting options for males to control females. We found that all copulations were voluntary, and females sexual activities were wholly unconstrained by male coercion. Most females had multiple copulatory partners in the weeks preceding egg-laying. Despite the commonness of EPC, EPFs did not occur. The different schedules of EPC and within-pair copulation (WPC) provided a sufficient explanation for this outcome: during the ovulation window days before laying, WPC rate increased and EPC rate approached zero. To our knowledge, this is the first robust evidence of complete sexual agency in a female bird aside from lek-mating species, contributing a valuable exemplar to the literature on sexual conflict over reproduction.

animal behavior and cognition↗

bioRxiv2025

Engineering microbial consortia for distributed signal processing

A critical goal in biology is deducing input signals from measurable readouts. Genetic circuits have successfully been designed to respond to specific analytes and produce quantifiable outputs; however, multiplexed signal processing remains challenging. This limitation is partially due to crosstalk, or sensors non-specific responses to unintended signals. While strategies to achieve orthogonality are promising, they are time-intensive and context-dependent. Here, we introduce a new, generalizable approach that leverages microbial consortia to distribute sensory functions and computational methods to disentangle signals. Compartmentalizing sensor circuits within distinct populations simplifies experimental optimization by allowing individual populations to be exchanged without requiring genetic modifications. Our computational pipeline combines mechanistic modeling with machine learning to decode microbial communities unique temporal responses and predict multiple input concentrations. We demonstrated this platforms versatility in a variety of contexts: measuring signals with high crosstalk, detecting antibiotics with natural microbial communities, and quantifying chemicals in hospital sink wastewater. Our findings highlight how combining microbial engineering with computational strategies can produce robust, scalable biosensors for diverse applications.

synthetic biology↗

bioRxiv2025

Sialidases derived from Gardnerella vaginalis remodel the sperm glycocalyx and impair sperm function

Bacterial vaginosis (BV), a dysbiosis of the vaginal microbiome, affects approximately 30 percent of women worldwide (up to 50% in some regions) and is associated with several adverse health outcomes including preterm birth and increased incidence of sexually transmitted infections (STIs). BV-associated bacteria such as Gardnerella vaginalis and Prevotella timonensis damage the vaginal mucosa through the activity of sialidase enzymes that remodel the epithelial glycocalyx and degrade mucin glycoproteins. This damage may contribute to adverse health outcomes. However, whether BV-associated glycolytic enzymes also damage sperm has not yet been determined. Here, we show that sialidase-mediated glycocalyx remodeling of human sperm increases sperm susceptibility to damage and adversely affects their function in vitro. Specifically, we report that sperm motility was not adversely affected by sialidase treatment, but desialylated human sperm demonstrate increased susceptibility to agglutination and complement-mediated cytotoxicity as well as impaired transit through cervical mucus. Our results demonstrate mechanisms by which BV-associated sialidases affect sperm survival and function and potentially contribute to adverse reproductive outcomes such as infertility.

immunology↗

bioRxiv2025

Keystone engineering enables collective range expansion in microbial communities

Keystone engineers profoundly influence microbial communities by altering their shared environment, often by modifying key resources. Here, we show that in an antibiotic-treated microbial community, bacterial spread is controlled by keystone engineering affecting dispersal--an effect hidden in well-mixed environments. Focusing on two pathogens, non-motile Klebsiella pneumoniae and motile Pseudomonas aeruginosa, we found that both tolerate a {beta}-lactam antibiotic, with Pseudomonas being more resilient and dominating in well-mixed cultures. During range expansion, however, the antibiotic inhibits Pseudomonas ability to spread unless it is near Klebsiella--Klebsiella degrades the antibiotic to create a "clear zone" that allows Pseudomonas to expand, at the expense of Klebsiellas own growth, thus acting as a keystone engineer. As Pseudomonas spreads, it competitively suppresses Klebsiella. Our modeling and experimental analyses reveal that this keystone effect operates at a millimeter scale. We also observed similar keystone engineering by a Bacillus species isolated from a hospital sink, in both pairwise and eight-member bacterial communities with its co-isolates. These findings suggest that spatially explicit experiments are essential to understand certain keystone engineering mechanisms and have implications for surface-associated microbial communities like biofilms, as well as for diagnosing and treating polymicrobial infections involving drug-degrading, non-motile (e.g., Klebsiella), and drug-tolerant, motile (e.g., Pseudomonas) bacteria.

ecology↗

bioRxiv2024

Population coding of predator imminence in the hypothalamus

Hypothalamic VMHdmSF1 neurons are activated by predator cues and are necessary and sufficient for instinctive defensive responses. However, such data do not distinguish which features of a predator encounter are encoded by VMHdmSF1 neural activity. To address this issue, we imaged VMHdmSF1 neurons at single-cell resolution in freely behaving mice exposed to a natural predator in varying contexts. Our results reveal that VMHdmSF1 neurons do not represent different defensive behaviors, but rather encode predator identity and multiple predator-evoked internal states, including threat-evoked fear/anxiety; neophobia or arousal; predator imminence; and safety. Notably, threat and safety are encoded bi-directionally by anti-correlated subpopulations. Finally, individual differences in predator defensiveness are correlated with differences in VMHdmSF1 response dynamics. Thus, different threat-related internal state variables are encoded by distinct neuronal subpopulations within a genetically defined, anatomically restricted hypothalamic cell class. HighlightsO_LIDistinct subsets of VMHdmSF1 neurons encode multiple predator-evoked internal states. C_LIO_LIAnti-correlated subsets encode safety vs. threat in a bi-directional manner C_LIO_LIA population code for predator imminence is identified using a novel assay C_LIO_LIVMHdmSF1 dynamics correlate with individual variation in predator defensiveness. C_LI

neuroscience↗

bioRxiv2024

Intrinsic dynamics and neural implementation of a hypothalamic line attractor encoding an internal behavioral state

Line attractors are emergent population dynamics hypothesized to encode continuous variables such as head direction and internal states. In mammals, direct evidence of neural implementation of a line attractor has been hindered by the challenge of targeting perturbations to specific neurons within contributing ensembles. Estrogen receptor type 1 (Esr1)-expressing neurons in the ventrolateral subdivision of the ventromedial hypothalamus (VMHvl) show line attractor dynamics in male mice during fighting. We hypothesized that these dynamics may encode continuous variation in the intensity of an internal aggressive state. Here, we report that these neurons also show line attractor dynamics in head-fixed mice observing aggression. We exploit this finding to identify and perturb line attractor-contributing neurons using 2-photon calcium imaging and holographic optogenetic perturbations. On-manifold perturbations demonstrate that integration and persistent activity are intrinsic properties of these neurons which drive the system along the line attractor, while transient off-manifold perturbations reveal rapid relaxation back into the attractor. Furthermore, stimulation and imaging reveal selective functional connectivity among attractor-contributing neurons. Intriguingly, individual differences among mice in line attractor stability were correlated with the degree of functional connectivity among contributing neurons. Mechanistic modelling indicates that dense subnetwork connectivity and slow neurotransmission are required to explain our empirical findings. Our work bridges circuit and manifold paradigms, shedding light on the intrinsic and operational dynamics of a behaviorally relevant mammalian line attractor.

neuroscience↗

bioRxiv2023

Oblique Line Scan Illumination Enables Expansive, Accurate and Sensitive Single Protein Measurements in Solution and in Living Cells

Single-molecule localization microscopy (SMLM) techniques, such as single-molecule tracking (SMT), enable in situ measurements in cells from which data-rich metrics can be extracted. SMT has been successfully applied to a variety of biological questions and model systems, aiming to unravel the spatiotemporal regulation of molecular mechanisms that govern protein function, downstream pathway effects, and cellular function. While powerful, SMLM often suffers from low throughput and illumination inhomogeneity, along with microscope and user-induced technical biases. Due to technical limitations in scaling SMLM techniques, a tradeoff between spatiotemporal resolution and throughput has been made historically, restricting broad application of these technologies. Here we address these limitations using Oblique Line Scan (OLS), a robust single-objective light-sheet based illumination and detection modality that achieves nanoscale spatial resolution and sub-millisecond temporal resolution across a 250 x 190 m field of view. We demonstrate OLS-enabled SMT on Halo-Tagged proteins in living cells capturing protein motion up to 14 m2 /s. By exploiting the adaptability of the acquisition frame rate and the improved rejection of out of focus light, we extend the utility of OLS beyond cellular compartments with in-solution SMT (isSMT) for single-molecule measurement of ligand-protein interactions and disruption of protein-protein interactions (PPI). We illustrate the versatility of OLS by showcasing two-color SMT, STORM, and single molecule fluorescence recovery after photobleaching (FRAP). OLS expands the range of SMLM applications and paves the way for robust, high-throughput single-molecule investigations of protein dynamics required for drug screening and systems biology studies, both in cells and in solution.

biophysics↗

bioRxiv2023

WRN Inhibition Leads to its Chromatin-Associated Degradation Via the PIAS4-RNF4-p97/VCP Axis

Synthetic lethality, the concept in which the co-occurrence of two genetic events leads to cell death while either single event alone does not, is an attractive strategy for targeted cancer therapies. A recent example of synthetic lethality as a therapeutic paradigm is the observation that cancer cells with high levels of microsatellite instability (MSI-H) are dependent on the Werner (WRN) RecQ helicase for survival. However, the mechanisms that regulate WRN spatiotemporal dynamics are not fully understood. In this study, we used our single molecule tracking (SMT) platform in combination with a recently disclosed WRN inhibitor to gain insights into WRNs dynamic localization within the nuclei of live cancer cells. We observe that WRN inhibition results in the helicase becoming trapped on chromatin, requiring p97/VCP for extraction and shuttling to the proteasome for degradation. Interestingly, this sequence of events resulting in WRN degradation appears to be MSI-H dependent. Using a phenotypic screen, we identify the PIAS4-RNF4 axis as the pathway responsible for WRN degradation and show that co-inhibition of WRN and SUMOylation has an additive toxic effect in MSI-H cells. Taken together, our work elucidates a novel regulatory mechanism for WRN. Gaining a deeper understanding into this regulatory pathway for WRN can aid in the identification of new high value targets for targeted cancer therapies.

molecular biology↗

bioRxiv2023

Distributed representations of innate behaviors in the hypothalamus do not predict specialized functional centers

Artificial activation of anatomically localized, genetically defined hypothalamic neuron populations is known to trigger distinct innate behaviors, suggesting a hypothalamic nucleus-centered organization of behavior control. To assess whether the encoding of behavior is similarly anatomically confined, we performed simultaneous neuron recordings across twenty hypothalamic regions in freely moving animals. Here we show that distinct but anatomically distributed neuron ensembles encode the social and fear behavior classes, primarily through mixed selectivity. While behavior class-encoding ensembles were spatially distributed, individual ensembles exhibited strong localization bias. Encoding models identified that behavior actions, but not motion-related variables, explained a large fraction of hypothalamic neuron activity variance. These results identify unexpected complexity in the hypothalamic encoding of instincts and provide a foundation for understanding the role of distributed neural representations in the expression of behaviors driven by hardwired circuits.

neuroscience↗

bioRxiv2023

Neuropeptide Signaling Is Required To Implement A Line Attractor Encoding A Persistent Internal Behavioral State

Internal states drive survival behaviors, but their neural implementation is not well understood. Recently we identified a line attractor in the ventromedial hypothalamus (VMH) that represents an internal state of aggressiveness. Line attractors can be implemented by recurrent connectivity and/or neuromodulatory signaling, but evidence for the latter is scant. Here we show that neuropeptidergic signaling is necessary for line attractor dynamics in this system, using a novel approach that integrates cell type-specific, anatomically restricted CRISPR/Cas9-based gene editing with microendoscopic calcium imaging. Co-disruption of receptors for oxytocin and vasopressin in adult VMH Esr1+ neurons that control aggression suppressed attack, reduced persistent neural activity and eliminated line attractor dynamics, while only modestly impacting neural activity and sex- or behavior-tuning. These data identify a requisite role for neuropeptidergic signaling in implementing a behaviorally relevant line attractor. Our approach should facilitate mechanistic studies in neuroscience that bridge different levels of biological function and abstraction.

neuroscience↗

bioRxiv2023

HI-FISH: WHOLE BRAIN IN SITU MAPPING OF NEURONAL ACTIVATION IN DROSOPHILA DURING SOCIAL BEHAVIORS AND OPTOGENETIC STIMULATION

Monitoring neuronal activity at single-cell resolution in freely moving Drosophila engaged in social behaviors is challenging because of their small size and lack of transparency. Extant methods, such as Flyception, are highly invasive. Whole-brain calcium imaging in head-fixed, walking flies is feasible but the animals cannot perform the consummatory phases of social behaviors like aggression or mating under these conditions. This has left open the fundamental question of whether neurons identified as functionally important for such behaviors using loss- or gain-of-function screens are actually active during the natural performance of such behaviors, and if so during which phase(s). Here we perform brain-wide mapping of active cells expressing the Immediate Early Gene hr38 using a high-sensitivity/low background FISH amplification method called HCR-3.0. Using double-labeling for hr38 mRNA and for GFP, we describe the activity of several classes of aggression-promoting neurons during courtship and aggression, including P1a cells, an intensively studied population of male-specific interneurons. Using HI-FISH in combination with optogenetic activation of aggression-promoting neurons (opto-HI-FISH) we identify candidate downstream functional targets of these cells in a brain-wide, unbiased manner. Finally we compare the activity of P1a neurons during sequential performance of courtship and aggression, using intronic vs. exonic hr38 probes to differentiate newly synthesized nuclear transcripts from cytoplasmic transcripts synthesized at an earlier time. These data provide evidence suggesting that different subsets of P1a neurons may be active during courtship vs. aggression. HI-FISH and associated methods may help to fill an important lacuna in the armamentarium of tools for neural circuit analysis in Drosophila.

neuroscience↗