eLife

Fossil central nervous systems (CNS) have illuminated several facets of total-group euarthropod evolution, including their phylogenetic affinities, the origin of the head, and their ancestral neuroanatomical organization. However, the CNS remains unknown in trilobites, the most diverse and earliest group of Palaeozoic euarthropods, but can potentially clarify fundamental questions that remain disputed despite thousands of described species and countless studies spanning over two centuries, such as their affinities relative to modern representatives. Here, we describe the CNS of the cheirurid Anacheirurus adserai from the Lower Ordovician Fezouata Shale. Anacheirurus features a brain with well-developed proto, deuto- and tritocerebral neuromeres, an anteriomedian protocerebral protrusion, a central complex, an esophageal foramen, and a ganglionated ventral nerve cord. The CNS is replicated in framboidal iron that differs in texture and composition from the exoskeleton and rock matrix. The overall organization of Anacheirurus's CNS most closely resembles that of Cambrian stem-group mandibulates, and critically shares the presence of a central complex with a protocerebral bridge and lateral accessory lobes with extant pancrustaceans. Our findings provide new morphological support for the hypothesis that Trilobita belongs to the mandibulate stem lineage and bring new insights into the ancestral morphology and evolutionary history of the euarthropod central complex. More broadly, our findings offer direct anatomical evidence of the key sensorimotor system innovations that originated during the Cambrian Information Revolution.

The mechanism of substrate-ATP coupling is central to the function of ABC transporters. Substrates bind to the specialized transmembrane domains (TMDs), whereas ATP binding and hydrolysis occur in the conserved intracellular nucleotide-binding domains (NBDs). Efficient substrate transport requires coupling between these processes, in which the intracellular coupling helices play an essential role. Here, we probed both coupling helices of the type VI ABC transporter LptB2FG(C), the core complex responsible for intermembrane lipopolysaccharide (LPS) transport in Gram-negative bacteria, using site-specific 19F labeling and ultra-fast MAS NMR. We show that both coupling helices exist in an equilibrium among three major conformational states. Progression through the coupling cycle occurs via conformational selection, as LPS and nucleotide binding shift the conformational equilibrium, providing evidence for bidirectional communication between the TMDs and NBDs. Furthermore, the conformational distributions of the two coupling helices are asymmetric. Finally, complex formation with LptC, a unique feature of type VI ABC transporters, symmetrizes the conformational landscape of the coupling helices and facilitates transitions between conformational states, thereby enhancing the efficiency of coupling ATP hydrolysis to LPS transport. Together, our findings establish the coupling helices as dynamic allosteric elements that integrate nucleotide and substrate binding through conformational selection, providing a mechanistic framework for substrate-ATP coupling in ABC transporters.

Postural control, the capacity of an animal to detect and correct an inappropriate body orientation, is a widespread and fundamental neurobiological function, yet the neural circuits that implement it remain poorly resolved in most species. The larva of Drosophila melanogaster performs a posture control stereotyped self-righting (SR) manoeuvre when turned upside-down, a behaviour previously shown to depend on a pair of segmentally repeated lateral transverse motor neurons (LT1/2-MNs) and on the normal expression of several Hox-targeting microRNAs. Here, we use a connectomics and a functional behavioural approach to trace and test the sensory, interneuronal and motor architecture of the SR circuit along the antero-posterior axis. Starting from the LT1/2-MNs, we identify a set of pre-motor interneurons, their principal upstream partners, and a population of class IV multidendritic sensory neurons as core components of the circuit, and show, through neuron-specific thermogenetic silencing, that inhibiting the great majority of these elements significantly impairs SR performance. We further find substantial overlap between the SR circuit and the previously described nociceptive rolling and touch crawling circuits, converging on shared interneurons including DnB, A02o (Wave-1) and TePn05. Network and axial connectivity analyses reveal a nested, hub-like organisation, a small number of integrator neurons bridging sensory and motor sub-networks, and a consistent decline in synapse number and density towards posterior segments. Together, these data yield the first axial wiring diagram for a postural control circuit in any animal and offer a set of structural principles including: hub organisation, shared sensory-motor structure, and antero-posterior connectivity gradients, which may extend to other segmentally organised nervous systems.

KCNQ2 (Kv7.2) subunits contribute to the channels that carry the neuronal M-current, and variants in KCNQ2 cause epilepsies that range from self-limited neonatal seizures to developmental and epileptic encephalopathy (DEE). Structures of KCNQ2 bound to PIP2 and cannabidiol show a widened activation gate, yet it has remained unclear which point of the activated pore limits K+ flux and how DEE variants at that point act. We simulated the KCNQ2 pore domain (PDB 8J01) in a POPC bilayer under an applied field of about 565 mV, restraining only the terminal residues of S5 and S6 so that G310 and S314 were free. The activated pore conducted K+ at 4.3 pS (95% CI 2.5-6.9; 17 net filter crossings in 1120 ns over four replicas), while the closed-state pore (PDB 7CR3) conducted none in 560 ns. Conduction proceeded by knock-on through a filter that was doubly occupied in 84-92% of frames. Along S6 the narrowest position of the activated pore was not the canonical gate residue S314 but the glycine of the Pro-Ala-Gly motif, G310: activation widens S314 by 0.37 nm but G310 by only 0.07 nm, and restraining G310 to its closed-state width in one replica abolished gate crossings. The DEE variant G310S reduced conduction about six-fold (G310S/wild-type ratio 0.16, 95% CI 0.02-0.66; p = 0.003). Its serine hydroxyls project into the constriction and halve the water column in the G310 plane (8.7 to 4.7 waters), an effect reproduced in CHARMM36m (9.9 to 4.3). The first coordination shell of a K+ ion at the constriction was, however, conserved (6.3 vs 6.5 oxygens), because the serine hydroxyls replace the waters they displace. The hydroxyls did not retain the ion: contacts lasted tens of picoseconds, and ions were instead turned back at the serine ring, fewer reaching the gate plane (45 vs 79 excursions per microsecond) and fewer of those crossing it (5% vs 23%). In mixed tetramers carrying one or two variant subunits the effect on the water column was additive, each subunit removing about a quarter of it (f = 0.22 and 0.58 of the full effect). These results identify G310 as the flux-limiting constriction of the activated KCNQ2 pore in our simulations and point to a permeation defect as one component of G310S loss of function.

Spontaneous voltage oscillations in neuronal ensembles play a critical role in memory formation and storage. Although oscillatory activities arising endogenously within central pattern-generating (CPG) networks underlie many rhythmic motor behaviors, the contribution of such autonomous signals to motor learning and memory remains poorly understood. Previously, we found that the buccal CPG network driving food-seeking behavior in Aplysia contains a subset of electrically-coupled neurons that produces spontaneous, variable-amplitude voltage oscillations instigating infrequent and irregular cycles of patterned motor output. This pattern-initiating activity originates from organelle-derived, inositol triphosphate (IP3) receptor-dependent calcium oscillations in a pair of identified decision-making neurons (B63) within the CPG subset (Bedecarrats et al., 2021). Here, we show that the cycle frequency of this spontaneous pacemaker mechanism based on intracellular calcium store release is persistently increased by operant reward-learning, and in association with increased B63 excitability, constitutes a fundamental memory trace for accelerated and stereotyped rhythmic food-seeking movements. eLife AssessmentSlow voltage oscillations in neuronal ensembles can contribute to learning and memory. This study identifies long-lasting plasticity in the dynamics of oscillatory intracellular calcium store release in key neurons of Aplysias food-seeking network, serving as a sub-cellular substrate for operant-reward learning that leads to the rhythmic expression of motor output producing compulsive-like behavior.

BCR and CD40 signals are integrated to shape B cell transcriptional output, but how the two pathways are coordinated at the level of individual signaling nodes is incompletely understood. Using primary mouse B cells stimulated with anti-IgM, anti-CD40, or both (Both), we defined six DEG classes reflecting costimulation-dependence and cross-pathway antagonism. We also connected a CD40-driven, BCR-suppressed B-cell-identity module (Pax5, Aicda, Bcl6, Cd79a/b, Cd19) to an increase in Blimp1 protein at 24 h. Building on our previous work, which showed that cIAP prolongs IKK/ERK activity after single-pathway stimulation (Shinohara et al., 2016), we added an IAP inhibitor 10 min after Both stimulation and generated transcriptomic data. This profiling showed that cIAP is required not only for late canonical NF-{kappa}B-driven genes, as predicted, but also, unexpectedly, for the entire BCR-dominant, CD40-independent negative-feedback module (Cd5, Il10, Nfkbid, Spry1/2) identified in the first dataset. A separate module was instead amplified by IAP inhibition, with kinetics compatible with non-canonical NF-{kappa}B de-repression, directly addressing a question we left open in that study regarding the BCR-side partners of cIAP. Together, these correlative findings converge on a single model: cIAP is not a CD40-restricted adaptor but a hub shared by BCR and CD40, whose feedback simultaneously sustains canonical, fate-instructive signaling and restrains the non-canonical pathway, thereby coupling receptor-proximal signal dynamics to the B cell's downstream transcriptional and fate decisions.

The past decade has seen remarkable progress in the molecular resolution of the influx and efflux components of mitochondrial Ca2+ handling, but little progress in matrix Ca2+ chelation that is central to mitochondrial calcium signaling. Among possible chelators, inorganic phosphate (Pi) is dynamic, and its uptake can lessen during Ca2+ uptake the rundown of the membrane potential, the primary driving force for Ca2+ uptake. Thus, PiC, the major mitochondrial Pi transporter, is well-positioned to regulate mitochondrial Ca2+ handling. To test this, we depleted (KD) PiC in murine skeletal muscle. We show that PiC depletion enhances the matrix free Ca2+ rise across a range of Ca2+ uptake activities, and surprisingly, causes elevated mitochondrial Ca2+ uptake, which seems to arise from an increased abundance of the mitochondrial Ca2+ uniporter. With protein levels of non-mitochondrial Ca2+ handling proteins unaltered, the greater mitochondrial Ca2+ uptake may drive a suppressed cytoplasmic Ca2+ response to tetanic stimulation, via lesser Ca2+-mediated positive feedback on Ca2+ release channels, contributing to an exercise deficit in KD mice. Alternatively, less buffering of matrix Ca2+ might be beneficial. We test these possibilities by lowering MCU, the uniporters pore forming subunit, in skeletal muscle of adult PiC KD mice, and find a worsened exercise deficit. This study establishes the requirement for PiC to maintain a bound fraction of Ca2+ in the matrix, and reveals a fitness benefit for elevated [Ca2+]m in striated muscle depleted of PiC.

Rod photoreceptors are optimized to detect single photons, but their performance near absolute visual threshold is limited by intrinsic phototransduction noise. The two major components are discrete single-photon-like events and continuous fluctuations in outer-segment current. Although spontaneous rhodopsin activation is established as the source of discrete noise, the molecular origin of continuous noise remains controversial, and how these distinct noise sources influence signal processing at the first visual synapse is not fully understood. To address these questions, we combined genetic manipulation of rhodopsin, phosphodiesterase 6 (PDE6), and glutamic acid rich protein 2 (GARP2) with single-cell recordings from rods and rod-bipolar cells (RBCs) and pharmacological manipulation of mGluR6 signaling. Continuous noise was unaffected by reduced rhodopsin concentration but was strongly influenced by PDE6 expression and regulation, supporting basal PDE6 activity as a major source of continuous noise. Reducing either discrete or continuous noise increased signal-to-noise ratio (SNR) in rods, but these changes were differentially processed at the rod-to-RBC synapse. In WT retina, the nonlinear transformation increased SNR, whereas elevated continuous noise limited this increase. Conversely, reduced continuous noise increased rod SNR, which was maintained in RBCs and associated with a lower detection threshold. Pharmacological reduction of the mGluR6-dependent nonlinearity altered these genotype-dependent differences, indicating that nonlinear filtering is well matched to normal signal and noise distributions but does not fully compensate when presynaptic noise is experimentally altered. Together, our study identifies spontaneous PDE6 activity as a major source of continuous noise and demonstrates that continuous noise interacts with nonlinear processing at the first visual synapse to determine the fidelity and detection threshold of dim-light signals.

During somitogenesis, the vertebrate body axis segments into transient periodic structures known as somites. Somite formation is regulated by a multicellular molecular oscillator known as the segmentation clock. Recent advances in human induced pluripotent stem cell (hiPSC) culture have shown that hiPSC-derived somitogenesis organoids (somitoids) exhibit segmentation clock oscillations and can be produced at scale, making them an excellent model system for high throughput investigation of the mechanisms underpinning the segmentation clock. However, somitoids interact both biochemically and mechanically, and accurate high-throughput sampling of segmentation clock phases is required to exploit this system effectively. Here we address these challenges using an image-based, high-content screening workflow. Individual hiPSC-derived somitoids carrying a segmentation clock reporter are cultured in 384-well plates, and a programmable feeding schedule is used to initiate oscillations that are monitored using fluorescence microscopy. We develop an automated pipeline for image segmentation and data analysis, represent oscillations using a compact set of parameters, and construct predictive mathematical models to interpret data. We find that: (i) a staggered feeding schedule that sequentially initiates oscillations yields large numbers of somitoids at defined stages of the segmentation clock cycle; (ii) media exchange in established oscillations induces a Type 0-like phase response, resetting the segmentation clock to a state characterised by low NOTCH pathway transcription; and (iii) control wells in media exchange experiments exhibit a Type 1 phase response in which the segmentation clock is delayed non-uniformly across the cycle. Using a mathematical model of segmentation clock dynamics along the anterior-posterior axis, we show that periodic activation of a Type 1 phase response could segment a continuous phase gradient -- an insight with potential implications for the determination of somite boundaries in vivo. IMPORTANTO_LIManuscripts submitted to Review Commons are peer reviewed in a journal-agnostic way. C_LIO_LIUpon transfer of the peer reviewed preprint to a journal, the referee reports will be available in full to the handling editor. C_LIO_LIThe identity of the referees will NOT be communicated to the authors unless the reviewers choose to sign their report. C_LIO_LIThe identity of the referee will be confidentially disclosed to any affiliate journals to which the manuscript is transferred. C_LI GUIDELINESO_LIFor reviewers: https://www.reviewcommons.org/reviewers C_LIO_LIFor authors: https://www.reviewcommons.org/authors C_LI CONTACTThe Review Commons office can be contacted directly at: office@reviewcommons.org

While filming Drosophila melanogaster recovering together from xenon anesthesia, we noticed frequent synchronous pauses in flies recovering in the same chamber. To ask whether such coordination could survive physical separation of the animals, we housed flies individually in the opaque wells of a multi-well plate, with no visual or tactile contact. They still coordinated the magnitude of their movements around a shared light*dark startle more closely than the shared stimulus alone can explain. We isolate this excess coordination with a measure that subtracts the stimulus-driven component by comparing movements within versus across transitions. A complementary common-mode statistic shows that flies also remain weakly but reproducibly correlated away from the transitions. The tested conventional channels do not readily account for the effect: vision is excluded by the opaque walls, and self-generated substrate vibration is disfavored for the background common mode because coordination does not scale with movement amplitude and persists when the continuous side-wall path between wells is interrupted by air gaps. The transition signal declined across the material series from plastic toward aluminum. This paNern is not a simple metal-versus-nonmetal split. It is consistent with material-dependent near-field magnetic radiofrequency coupling through the well geometry and transparent imaging aperture. Vector-network measurements also indicate that near-field inter- well coupling differs between steel and aluminum. We have previously shown that Drosophila emit spontaneous, metabolically powered magnetic radiofrequency radiation that tracks nervous-system activity. Together these results are consistent with a near-field magnetic emission that can leak between separated individuals. We discuss radical-pair detection, possible emission mechanisms, and experiments needed to identify the carrier frequency, detector, and biological role. SignificanceFruit flies housed in separate, opaque chambers that block sight and touch nonetheless coordinate their movements. The coordination persists when mechanical vibration paths between chambers are cut. The coordination is absent when the plate is made of aluminum, and intermediate in size when the plate is made of ferromagnetic steel. This ordering, together with direct RF-coupling measurements, points to a near-field magnetic radiofrequency signal passing between individuals, though the carrier frequency and detector remain unknown. Because flies are already known to emit faint, metabolically powered radiofrequency fields that track nervous-system activity, these results raise the possibility that nervous systems both emit and detect such fields. If confirmed, this would identify an unrecognized magnetic radiofrequency channel in animal biology and suggest that brains are less electromagnetically private than assumed.

Effective mentorship is essential for strengthening the STEMM (Science, Technology, Engineering, Mathematics, and Medicine) workforce, yet empirical evidence on how mentorship networks are structured and linked to career success remains limited. Here, we analyze mentorship networks among recipients of NIH career development (K) awards to characterize network size, mentor roles, and their associations with mentee-reported outcomes, including potential variation by sociodemographic characteristics. We found that K-awardees rely on mentors beyond their primary advisor, who play varying roles beyond being a Research mentor. Different mentor roles led to different types of mentoring outcomes; while Research mentors were associated with research-related outcomes such as Publications and Grants, career- and psychosocial-related mentoring outcomes were more likely to come from other types of mentors, such as Coaches, Connectors, and Sponsors. Larger networks, as well as having Peer and Identity mentors are additively beneficial for researchers who identify as underrepresented in science more than their counterparts. This study provides large-scale evidence on how mentorship network configurations relate to early-career grant success.

A dental AI system and a dentist reading the same radiographs form a paired comparison. Published comparative studies often report the two arms separately against a reference standard, leaving the joint pattern of correctness between them unavailable for secondary paired inference. We show what that omission costs. The accuracy difference remains exactly identified; its sampling variance does not, so the report contains the estimate and not its uncertainty. On a study of 282 units, two published accuracies are consistent with 38 distinct joint tables whose confidence intervals differ in width by a factor of 2.5. The consequence is a three-zone decision map rather than a single threshold: differences at or below 1.06 points are non-significant under every compatible table, differences at or above 6.03 points are significant under every compatible table, and in between the published numbers cannot decide. We then show the omission is repairable at negligible cost. One additional integer, the number of units both arms classify correctly, identifies the joint table exactly and restores standard paired inference. For a panel of readers the pairwise dependences must arise from one joint distribution, a constraint that binds once three readers are present; publishing each reader's joint-correct count against a single reference reader cannot widen and may tighten every pairwise bound, and in a 7-arm experiment reduced them by a median of 37% even for pairs excluding that reference. Where the integer was never published we give DentalPair-Cert, an interval with finite-sample coverage uniformly over every admissible within-unit AI-dentist dependence under the independent-sampling-unit model, certified in both the nuisance maximization and the inversion. Across 4,200,000 simulated comparisons an independence analysis falls to 74.5% coverage with 12.2% type-I error; in a purposive sample of 9 recent comparative studies, 1 reported a paired test on discordant units.

Whether synaptic plasticity stabilises or destabilises a recurrent circuit is usually treated as a question about the rule. We find that in a conductance-based model it is not answered by the rule alone. The model is a simulated bursting culture: 48 excitatory and 12 inhibitory stomatogastric-ganglion neurons, sparsely and recurrently connected. The recurrent synapses carry pair-based spike-timing-dependent plasticity under a fixed homeostatic budget on each cell's total incoming excitatory conductance. We ran one net-depressing rule, at one inhibition level, on unselected random wirings, with a survival criterion fixed before the runs. Two excitatory populations built from different published conductance sets gave opposite outcomes. In one, the network collapsed without the rule in 33 of 36 wirings and the rule rescued 31 of those 33. In the other, the network survived without the rule in 11 of 12 wirings and the rule eliminated 6 of those 11. The only model parameter changed between the two is the excitatory conductance set, and that change also alters their intrinsic dynamics. We then asked what carries the reversal. Across seven conductance sets, elimination increases with a cell's own firing rate, and cell identity does not predict it: a follower-type cell firing at pacemaker rates behaves like a pacemaker. But moving the firing rate inside a single set by injected current reverses the same quantity, which rules out firing rate as a sufficient explanation. Substituting one conductance at a time from the rescued set into the other, over the six channels that differ, no single substitution transferred the rescue (2 of 42 pooled) while substituting all eight did so in 6 of 6 (Fisher exact p = 2.3 x 10^-6). The sign reversal cannot be attributed to any single tested conductance, and firing rate alone cannot account for it. All results are computational and concern one circuit model, one plasticity rule and one homeostatic constraint. Predictions and falsifiers were committed to a version-controlled record before the corresponding runs, with three departures from that order recorded in the manuscript's pre-registration index.

Axon collateral branching is a fundamental determinant of neuronal connectivity, enabling individual neurons to innervate multiple targets and establish functional neural circuits. Although branch initiation requires local actin remodelling within the axon shaft, the mechanisms that restrict actin assembly to prevent excessive branching remain poorly understood. Here, we identify Kaptin (Kptn) as a conserved negative regulator of axon collateral formation that limits the maturation of axonal actin patches. Using primary neuronal cultures, quantitative live-cell imaging, and zebrafish genetics, we show that loss of Kptn enhances the conversion of actin patches into filopodial protrusions, leading to excessive collateral branching. Kptn-deficient zebrafish exhibit increased motor axon arborisation, elevated neuromuscular junction density and impaired motor behaviour. Mechanistically, Kptn antagonises the actin elongation factor Formin-2 (Fmn2) to regulate actin filament barbed-end dynamics, thereby controlling the threshold for productive branch initiation. Importantly, these branching defects occur independently of Kptns established role in mTORC1 signalling, revealing a distinct physiological function during neuronal development. Our findings identify Kptn as a key molecular brake that constrains axon collateral branching and establish negative regulation of actin patch maturation as a fundamental mechanism controlling neuronal circuit assembly. This work provides a mechanistic framework for understanding how KPTN mutations associated with intellectual disability and epilepsy disrupt neuronal connectivity.

The rapidly increasing number of video tracking-based behavioral summary tools and methods raises the question as to the most suitable approaches for pharmacological fingerprinting in pre-clinical research. We have recently shown that social context has a strong effect on behavioral syntax in mice, further suggesting that treatment effects can be context dependent. Here, we aim to answer the question whether there is an optimal combination of context and behavioral summary method for effect detection and discrimination of different psychoactive substances in a controlled environment. To this end, we applied eight different treatment-dosage pairs (amphetamine 1.5,3,6 mg/kg; modafinil 5,10,50 mg/kg; seltorexant 3,10 mg/kg) and evaluated five different approaches to behavioral summary: parametric aggregation, unsupervised segmentation in Keypoint-MoSeq (KPMS) & Variational Animal Motion Encoding (VAME), and supervised segmentation in Simple Behavioral Analysis (SimBA) & A-SOiD, across two different contexts (Solitary & Social) in 314 recordings of freely moving mice in an open-field arena. Surprisingly, our results show no significant differences in performance across models and context. Across treatment effect detection to treatment-dose discrimination, all models showed performance significantly above chance that was insensitive to various data limitations and extensions. Overall, our study shows that under the tested conditions and treatments, the choice of a behavioral summary model does not meaningfully affect the description of treatment effects. Simple aggregate measures from tracking data and machine learning based behavioral summary approaches that are expensive, in terms of training data and computational resources, performed equally well. Our findings taken together with literature suggest that the fuller decomposition of complex behavior through unsupervised machine learning might be necessary for the description of large-scale datasets but does not necessarily align with the goals present in smaller-scale treatment discrimination tasks common in pre-clinical research.

Many falls and balance disorders in older adults originate in the otolith organs, the gravity sensors of the inner ear. These sensors maintain upright posture through otoconia, calcium carbonate crystals that mass-load the sensory maculae. Otoconia dislodgement causes the most common form of vertigo, and their age-related loss reduces gravity sensation and undermines balance. Yet the cellular mechanisms of otoconia formation and maintenance -and how they fail in disease- remain unknown. Using mineral-preserving histology, crystal-sensitive imaging, volume electron microscopy, and immunolabeling in human and animal otolith organs, we found otoconia biogenesis-related proteins and early crystallization at the pole opposing the macula, the roof domain. We discovered filigree extracellular matrix scaffolds bridging roof and macula, loaded with nascent otoconia, suggesting scaffold-guided transport across the organ. Single-cell transcriptomics nominated a specialized roof mesenchyme, signaling to the roof epithelium, as a driver of otoconia and scaffold production. In guinea pigs with endolymphatic hydrops, fluid expansion of the organs ruptured the otoconia-trafficking scaffolds as roof and macula separated, followed by a decline in macular otoconial mass. We propose a new disease model for common vertigo and balance disorders in which disruption of the otoconia-generating and -trafficking epithelial-mesenchymal roof niche leads to displacement and depletion of otoconia.

Opsins are light-activated receptors. A basic question is how the molecular properties of an opsin shape its effects on the organism. In mammals, melanopsin regulates perception, cognition, physiology, and mood. Because these actions are vital, an international standard has been formulated to express light in terms of melanopsin stimulation. Short wavelengths are absorbed best by melanopsin and should drive its effects most strongly. We report that melanopsin's cellular and behavioral effects show little or even opposite wavelength sensitivity in some regimes. This divergence stems from melanopsin's persistent activity, which generates adaptation. Wavelengths absorbed strongly by melanopsin produce high persistent activity and adaptation, giving net desensitization, whereas wavelengths absorbed weakly can produce sensitization instead. We provide a quantitative model that captures these transformations. This work indicates how the downstream actions of a molecule may diverge starkly from its intrinsic properties.

Drosophila s-LNv clock neurons promote nighttime sleep by releasing the neuropeptide sNPF to activate sNPF receptors (sNPF-Rs) on l-LNv clock neurons. Behavior is controlled by synaptic transmission, but s-LNv and l-LNv neurons are not connected directly by chemical synapses. To investigate the basis of sNPF/sNPF-R communication between LNv neurons, the spread of sNPF was imaged in the adult brain. We report the daily midmorning burst of sNPF released from s-LNv terminals does not reach l-LNv neurons or s-LNv somata. Instead, sNPF released by the s-LNv soma late at night in response to sleep-promoting IP3 signaling reaches l-LNv somata, but not their terminals. In addition to communication by neuropeptide diffusion, analysis of fly connectomes revealed that adult s-LNv and l-LNv neurons form non-synaptic direct contacts mediated by cytonemes. Remarkably, genetically perturbing LNv neuron cytonemes alters sleep latency, but not nighttime sleep, the target of somatic sNPF release, or circadian behavior, which depends on PDF neuropeptide released by LNv terminals. Therefore, three distinct aspects of adult rhythmic behavior are produced by terminals, the soma and cytonemes, with the latter possibly acting via contacts that are not currently annotated in the connectome.

In his seminal 1920s studies, Otto Warburg found the retina to generate large amounts of lactate. However, it was unclear what retinal cells produced lactate and whether it was a metabolic waste product or used further. Here, we show that lactate produced by rod photoreceptors fuels the energy-intensive function and viability of cone photoreceptors. In an initial expression analysis, we found monocarboxylate transporter-1 (MCT1), lactate-producing lactate-dehydrogenase-A (LDHA), and pyruvate carboxykinase-1 (PCK1) localized to rod photoreceptors, while high-affinity MCT2, pyruvate-producing LDHB, and PCK2 were expressed in cones. We then exposed retina to defined media containing either glucose or lactate as caloric component, and applied specific MCT inhibitors. In glucose-containing medium, 1H-NMR metabolomics showed rod MCT1 inhibition to increase retinal lactate, suggesting rods as a major source of lactate. In lactate-only medium, functional recordings using micro-electroretinography showed decreased rod function, while cone function was maintained. In glucose-containing medium, blocking rod MCT1 abolished cone function. Long-term treatment with MCT inhibitors selectively decreased photoreceptor viability. Conversely, supplementing the defined medium with lactate preserved cone viability in the rd1 mouse model for Retinitis Pigmentosa. Together, our data suggest that lactate shuttling from rods is crucial for cone function and viability. This may explain cone degeneration seen in various retinal diseases and provides an entirely new avenue for metabolism-based treatment development. The discovery of a lactate-shuttle between two functionally similar, yet distinct types of neurons may have far-reaching implications for our understanding of the central nervous system in general.

Many G protein-coupled receptors (GPCRs) are capable of activating multiple heterotrimeric G protein subtypes. The angiotensin II type 1 receptor (AT1R) couples to both Gq and Gi family members, and the two families mediate distinct aspects of the effects of angiotensin II (AngII) on blood pressure and fluid balance. Previous work has pointed to fundamental differences in the strength and ligand-dependence of Gq versus Gi activation by AT1R. Here we resolve key aspects of the mechanisms underlying these subtype-specific patterns of G protein coupling to AT1R. Using cellular biosensors, we show that AT1R activates Gq more efficaciously than Gi. Leveraging mini-G proteins, engineered versions of G subunits that diverge only in the C-terminal 5 helix, we identify four residues dispersed throughout the 5 helix that are sufficient to account for these differences. We employ radioligand binding to demonstrate that subtype-specific differences originate at the level of allosteric coupling between ligand and transducer binding, and we use Systematic Protein Affinity Strength Modulation biosensors to show that Gq-specific residues in the 5 helix increase affinity for AngII-activated AT1R. We provide evidence that the 5 helix residue with the greatest effect on coupling makes interactions with AT1R intracellular loop 3 that were not resolved in a previously reported AT1R-Gq cryo-electron microscopy structure. These findings enhance our understanding of how AT1R differentially activates Gq and Gi and may ultimately guide pharmacological approaches to modulate certain G proteins downstream of AT1R and other GPCRs.