Population genomic structure of a widespread, urban-dwelling mammal: The eastern grey squirrel (Sciurus carolinensis).

Urbanization is a persistent and widespread driver of global environmental change, potentially shaping evolutionary processes due to genetic drift and reduced gene flow in cities induced by habitat fragmentation and small population sizes. We tested this prediction for the eastern grey squirrel (Sciurus carolinensis), a common and conspicuous forest-dwelling rodent, by obtaining 44K SNPs using reduced representation sequencing (ddRAD) for 403 individuals sampled across the species' native range in eastern North America. We observed moderate levels of genetic diversity, low levels of inbreeding, and only a modest signal of isolation-by-distance. Clustering and migration analyses show that estimated levels of migration and genetic connectivity were higher than expected across cities and forested areas, specifically within the eastern portion of the species' range dominated by urbanization, and genetic connectivity was less than expected within the western range where the landscape is fragmented by agriculture. Landscape genetic methods revealed greater gene flow among individual squirrels in forested regions, which likely provide abundant food and shelter for squirrels. Although gene flow appears to be higher in areas with more tree cover, only slight discontinuities in gene flow suggest eastern grey squirrels have maintained connected populations across urban areas in all but the most heavily fragmented agricultural landscapes. Our results suggest urbanization shapes biological evolution in wildlife species depending strongly on the composition and habitability of the landscape matrix surrounding urban areas.

Seasonality and uncertainty in global COVID-19 growth rates.

The virus causing COVID-19 has spread rapidly worldwide and threatens millions of lives. It remains unknown, as of April 2020, whether summer weather will reduce its spread, thereby alleviating strains on hospitals and providing time for vaccine development. Early insights from laboratory studies and research on related viruses predicted that COVID-19 would decline with higher temperatures, humidity, and ultraviolet (UV) light. Using current, fine-scaled weather data and global reports of infections, we develop a model that explains 36% of the variation in maximum COVID-19 growth rates based on weather and demography (17%) and country-specific effects (19%). UV light is most strongly associated with lower COVID-19 growth. Projections suggest that, without intervention, COVID-19 will decrease temporarily during summer, rebound by autumn, and peak next winter. Validation based on data from May and June 2020 confirms the generality of the climate signal detected. However, uncertainty remains high, and the probability of weekly doubling rates remains >20% throughout summer in the absence of social interventions. Consequently, aggressive interventions will likely be needed despite seasonal trends.

Evolutionary origins for ecological patterns in space.

Historically, many biologists assumed that evolution and ecology acted independently because evolution occurred over distances too great to influence most ecological patterns. Today, evidence indicates that evolution can operate over a range of spatial scales, including fine spatial scales. Thus, evolutionary divergence across space might frequently interact with the mechanisms that also determine spatial ecological patterns. Here, we synthesize insights from 500 eco-evolutionary studies and develop a predictive framework that seeks to understand whether and when evolution amplifies, dampens, or creates ecological patterns. We demonstrate that local adaptation can alter everything from spatial variation in population abundances to ecosystem properties. We uncover 14 mechanisms that can mediate the outcome of evolution on spatial ecological patterns. Sometimes, evolution amplifies environmental variation, especially when selection enhances resource uptake or patch selection. The local evolution of foundation or keystone species can create ecological patterns where none existed originally. However, most often, we find that evolution dampens existing environmental gradients, because local adaptation evens out fitness across environments and thus counteracts the variation in associated ecological patterns. Consequently, evolution generally smooths out the underlying heterogeneity in nature, making the world appear less ragged than it would be in the absence of evolution. We end by highlighting the future research needed to inform a fully integrated and predictive biology that accounts for eco-evolutionary interactions in both space and time.

Cool microrefugia accumulate and conserve biodiversity under climate change.

A major challenge in climate change biology is to explain why the impacts of climate change vary around the globe. Microclimates could explain some of this variation, but climate change biologists often overlook microclimates because they are difficult to map. Here, we map microclimates in a freshwater rock pool ecosystem and evaluate how accounting for microclimates alters predictions of climate change impacts on aquatic invertebrates. We demonstrate that average maximum temperature during the growing season can differ by 9.9-11.6°C among microclimates less than a meter apart and this microclimate variation might increase by 21% in the future if deeper pools warm less than shallower pools. Accounting for this microclimate variation significantly alters predictions of climate change impacts on aquatic invertebrates. Predictions that exclude microclimates predict low future occupancy (0.08-0.32) and persistence probabilities (2%-73%) for cold-adapted taxa, and therefore predict decreases in gamma richness and a substantial shift toward warm-adapted taxa in local communities (i.e., thermophilization). However, predictions incorporating microclimates suggest cool locations will remain suitable for cold-adapted taxa in the future, no change in gamma richness, and 825% less thermophilization. Our models also suggest that cool locations will become suitable for warm-adapted taxa and will therefore accumulate biodiversity in the future, which makes cool locations essential for biodiversity conservation. Simulated protection of the 10 coolest microclimates (9% of locations on the landscape) results in a 100% chance of conserving all focal taxa in the future. In contrast, protecting the 10 currently most biodiverse locations, a commonly employed conservation strategy, results in a 3% chance of conserving all focal taxa in the future. Our study suggests that we must account for microclimates if we hope to understand the future impacts of climate change and design effective conservation strategies to limit biodiversity loss.

Regional neutrality evolves through local adaptive niche evolution.

Biodiversity in natural systems can be maintained either because niche differentiation among competitors facilitates stable coexistence or because equal fitness among neutral species allows for their long-term cooccurrence despite a slow drift toward extinction. Whereas the relative importance of these two ecological mechanisms has been well-studied in the absence of evolution, the role of local adaptive evolution in maintaining biological diversity through these processes is less clear. Here we study the contribution of local adaptive evolution to coexistence in a landscape of interconnected patches subject to disturbance. Under these conditions, early colonists to empty patches may adapt to local conditions sufficiently fast to prevent successful colonization by other preadapted species. Over the long term, the iteration of these local-scale priority effects results in niche convergence of species at the regional scale even though species tend to monopolize local patches. Thus, the dynamics evolve from stable coexistence through niche differentiation to neutral cooccurrence at the landscape level while still maintaining strong local niche segregation. Our results show that neutrality can emerge at the regional scale from local, niche-based adaptive evolution, potentially resolving why ecologists often observe neutral distribution patterns at the landscape level despite strong niche divergence among local communities.

LAR inhibitory peptide promotes recovery of diaphragm function and multiple forms of respiratory neural circuit plasticity after cervical spinal cord injury.

Chondroitin sulfate proteoglycans (CSPGs), up-regulated in and around the lesion after traumatic spinal cord injury (SCI), are key extracellular matrix inhibitory molecules that limit axon growth and consequent recovery of function. CSPG-mediated inhibition occurs via interactions with axonal receptors, including leukocyte common antigen- related (LAR) phosphatase. We tested the effects of a novel LAR inhibitory peptide in rats after hemisection at cervical level 2, a SCI model in which bulbospinal inspiratory neural circuitry originating in the medullary rostral ventral respiratory group (rVRG) becomes disconnected from phrenic motor neuron (PhMN) targets in cervical spinal cord, resulting in persistent partial-to-complete diaphragm paralysis. LAR peptide was delivered by a soaked gelfoam, which was placed directly over the injury site immediately after C2 hemisection and replaced at 1 week post-injury. Axotomized rVRG axons originating in ipsilateral medulla or spared rVRG fibers originating in contralateral medulla were separately assessed by anterograde tracing via AAV2-mCherry injection into rVRG. At 8 weeks post-hemisection, LAR peptide significantly improved ipsilateral hemidiaphragm function, as assessed in vivo with electromyography recordings. LAR peptide promoted robust regeneration of ipsilateral-originating rVRG axons into and through the lesion site and into intact caudal spinal cord to reach PhMNs located at C3-C5 levels. Furthermore, regenerating rVRG axons re-established putative monosynaptic connections with their PhMNs targets. In addition, LAR peptide stimulated robust sprouting of both modulatory serotonergic axons and contralateral-originating rVRG fibers within the PhMN pool ipsilateral/caudal to the hemisection. Our study demonstrates that targeting LAR-based axon growth inhibition promotes multiple forms of respiratory neural circuit plasticity and provides a new peptide-based therapeutic strategy to ameliorate the devastating respiratory consequences of SCI.

Adaptation reduces competitive dominance and alters community assembly.

A growing body of theory predicts that evolution of an early-arriving species in a new environment can produce a competitive advantage against later arriving species, therefore altering community assembly (i.e. the community monopolization hypothesis). Applications of the community monopolization hypothesis are increasing. However, experimental tests of the hypothesis are rare. Here, we provide a rare experimental demonstration of the community monopolization hypothesis using two archaeal species. We first expose one species to low- and high-temperature environments for 135 days. Populations in the high-temperature treatment evolved a 20% higher median performance when grown at high temperature. We then demonstrate that early arrival and adaptation reduce the abundance of a late-arriving species in the high-temperature environment by 63% relative to when both species arrive simultaneously and neither species is adapted to high temperature. These results are consistent with the community monopolization hypothesis and suggest that adaptation can reduce competitive dominance to alter community assembly. Hence, community monopolization might be much more common in nature than previously assumed. Our results strongly support the idea that patterns of biodiversity might often stem from a race between local adaptation and colonization of pre-adapted species.

The circadian clock gene Per1 modulates context fear memory formation within the retrosplenial cortex in a sex-specific manner.

Context memory formation is a complex process that requires transcription in many subregions of the brain including the dorsal hippocampus and retrosplenial cortex. One critical gene necessary for memory formation is the circadian gene Period1 (Per1), which has been shown to function in the dorsal hippocampus to modulate spatial memory in addition to its well-documented role in regulating the diurnal clock within the suprachiasmatic nucleus (SCN). We recently found that alterations in Per1 expression in the dorsal hippocampus can modulate spatial memory formation, with reduced hippocampal Per1 impairing memory and overexpression of Per1 ameliorating age-related impairments in spatial memory. Whether Per1 similarly functions within other memory-relevant brain regions is currently unknown. Here, to test whether Per1 is a general mechanism that modulates memory across the brain, we tested the role of Per1 in the retrosplenial cortex (RSC), a brain region necessary for context memory formation. First, we demonstrate that context fear conditioning drives a transient increase in Per1 mRNA expression within the anterior RSC that peaks 60 m after training. Next, using HSV-CRISPRi-mediated knockdown of Per1, we show that reducing Per1 within the anterior RSC before context fear acquisition impairs memory in both male and female mice. In contrast, overexpressing Per1 with either HSV-CRISPRa or HSV-Per1 before context fear acquisition drives a sex-specific memory impairment; males show impaired context fear memory whereas females are not affected by Per1 overexpression. Finally, as Per1 levels are known to rhythmically oscillate across the day/night cycle, we tested the possibility that Per1 overexpression might have different effects on memory depending on the time of day. In contrast to the impairment in memory we observed during the daytime, Per1 overexpression has no effect on context fear memory during the night in either male or female mice. Together, our results indicate that Per1 modulates memory in the anterior retrosplenial cortex in addition to its documented role in regulating memory within the dorsal hippocampus, although this role may differ between males and females.

Balancing selection and drift in a polymorphic salamander metapopulation.

Understanding how genetic variation is maintained in a metapopulation is a longstanding problem in evolutionary biology. Historical resurveys of polymorphisms have offered efficient insights about evolutionary mechanisms, but are often conducted on single, large populations, neglecting the more comprehensive view afforded by considering all populations in a metapopulation. Here, we resurveyed a metapopulation of spotted salamanders (Ambystoma maculatum) to understand the evolutionary drivers of frequency variation in an egg mass colour polymorphism. We found that this metapopulation was demographically, phenotypically and environmentally stable over the last three decades. However, further analysis revealed evidence for two modes of evolution in this metapopulation-genetic drift and balancing selection. Although we cannot identify the balancing mechanism from these data, our findings present a clear view of contemporary evolution in colour morph frequency and demonstrate the importance of metapopulation-scale studies for capturing a broad range of evolutionary dynamics.

Microgeographic divergence of functional responses among salamanders under antagonistic selection from apex predators.

A predator's functional response determines predator-prey interactions by describing the relationship between the number of prey available and the number eaten. Its shape and parameters fundamentally govern the dynamic equilibrium of predator-prey interactions and their joint abundances. Yet, estimates of these key parameters generally assume stasis in space and time and ignore the potential for local adaptation to alter feeding responses and the stability of trophic dynamics. Here, we evaluate if functional responses diverge among populations of spotted salamander (Ambystoma maculatum) larvae that face antagonistic selection on feeding strategies based on their own risk of predation. Common garden experiments revealed that spotted salamander from ponds with varying predation risks differed in their functional responses, suggesting an evolutionary response. Applying mechanistic equations, we discovered that the combined changes in attack rates, handling times and shape of the functional response enhanced feeding rate in environments with high densities of gape-limited predators. We suggest how these parameter changes could alter community equilibria and other emergent properties of food webs. Community ecologists might often need to consider how local evolution at fine scales alters key relationships in ways that alter local diversity patterns, food web dynamics, resource gradients and community responses to disturbance.

AAV2-BDNF promotes respiratory axon plasticity and recovery of diaphragm function following spinal cord injury.

More than half of spinal cord injury (SCI) cases occur in the cervical region, leading to respiratory dysfunction due to damaged neural circuitry that controls critically important muscles such as the diaphragm. The C3-C5 spinal cord is the location of phrenic motor neurons (PhMNs) that are responsible for diaphragm activation; PhMNs receive bulbospinal excitatory drive predominately from supraspinal neurons of the rostral ventral respiratory group (rVRG). Cervical SCI results in rVRG axon damage, PhMN denervation, and consequent partial-to-complete paralysis of hemidiaphragm. In a rat model of C2 hemisection SCI, we expressed the axon guidance molecule, brain-derived neurotrophic factor (BDNF), selectively at the location of PhMNs (ipsilateral to lesion) to promote directed growth of rVRG axons toward PhMN targets by performing intraspinal injections of adeno-associated virus serotype 2 (AAV2)-BDNF vector. AAV2-BDNF promoted significant functional diaphragm recovery, as assessed by in vivo electromyography. Within the PhMN pool ipsilateral to injury, AAV2-BDNF robustly increased sprouting of both spared contralateral-originating rVRG axons and serotonergic fibers. Furthermore, AAV2-BDNF significantly increased numbers of putative monosynaptic connections between PhMNs and these sprouting rVRG and serotonergic axons. These findings show that targeting circuit plasticity mechanisms involving the enhancement of synaptic inputs from spared axon populations is a powerful strategy for restoring respiratory function post-SCI.-Charsar, B. A., Brinton, M. A., Locke, K., Chen, A. Y., Ghosh, B., Urban, M. W., Komaravolu, S., Krishnamurthy, K., Smit, R., Pasinelli, P., Wright, M. C., Smith, G. M., Lepore, A. C. AAV2-BDNF promotes respiratory axon plasticity and recovery of diaphragm function following spinal cord injury.

Developmental temperature influences color polymorphism but not hatchling size in a woodland salamander.

Phenotypic plasticity can be an important adaptive response to climate change, particularly for dispersal-limited species. Temperature frequently alters developmental and phenotypic traits including morphology, behavior, and reproductive cycles. We often lack crucial information about if and how thermal conditions during development will interact with genetic responses and facilitate persistence or adaptation under climate change. Polymorphic species offer an ideal test for this, as alternative morphs often confer differential adaptive advantages. However, few studies have examined the effects of incubation temperature on color expression or development in polymorphic taxa. Here we test if developmental temperature mediates morph frequency in the polymorphic salamander Plethodon cinereus. Although previous research suggests geographic variation in morph proportions results from differential climate adaptation, it remains unknown if plasticity also contributes to this variation. We used a split-clutch common garden experiment to determine the effects of developmental temperature on the color and development of P. cinereus. Our results indicate developmental temperature affects coloration in P. cinereus, either via plasticity or differential mortality, with eggs incubated at warmer temperatures yielding a higher proportion of unstriped individuals than those from cooler temperatures. This temperature response may contribute to the spatial variation in morph frequencies in natural populations. Surprisingly, we found neither temperature nor egg size affected hatchling size. Our study provides important insights into the potential for climate-induced responses to preserve diversity in dispersal-limited species, like P. cinereus, and enable time for adaptive evolution.

Local BDNF Delivery to the Injured Cervical Spinal Cord using an Engineered Hydrogel Enhances Diaphragmatic Respiratory Function.

We developed an innovative biomaterial-based approach to repair the critical neural circuitry that controls diaphragm activation by locally delivering brain-derived neurotrophic factor (BDNF) to injured cervical spinal cord. BDNF can be used to restore respiratory function via a number of potential repair mechanisms; however, widespread BDNF biodistribution resulting from delivery methods such as systemic injection or lumbar puncture can lead to inefficient drug delivery and adverse side effects. As a viable alternative, we developed a novel hydrogel-based system loaded with polysaccharide-BDNF particles self-assembled by electrostatic interactions that can be safely implanted in the intrathecal space for achieving local BDNF delivery with controlled dosing and duration. Implantation of BDNF hydrogel after C4/C5 contusion-type spinal cord injury (SCI) in female rats robustly preserved diaphragm function, as assessed by in vivo recordings of compound muscle action potential and electromyography amplitudes. However, BDNF hydrogel did not decrease lesion size or degeneration of cervical motor neuron soma, suggesting that its therapeutic mechanism of action was not neuroprotection within spinal cord. Interestingly, BDNF hydrogel significantly preserved diaphragm innervation by phrenic motor neurons (PhMNs), as assessed by detailed neuromuscular junction morphological analysis and retrograde PhMN labeling from diaphragm using cholera toxin B. Furthermore, BDNF hydrogel enhanced the serotonergic axon innervation of PhMNs that plays an important role in modulating PhMN excitability. Our findings demonstrate that local BDNF hydrogel delivery is a robustly effective and safe strategy to restore diaphragm function after SCI. In addition, we demonstrate novel therapeutic mechanisms by which BDNF can repair respiratory neural circuitry.SIGNIFICANCE STATEMENT Respiratory compromise is a leading cause of morbidity and mortality following traumatic spinal cord injury (SCI). We used an innovative biomaterial-based drug delivery system in the form of a hydrogel that can be safely injected into the intrathecal space for achieving local delivery of brain-derived neurotrophic factor (BDNF) with controlled dosing and duration, while avoiding side effects associated with other delivery methods. In a clinically relevant rat model of cervical contusion-type SCI, BDNF hydrogel robustly and persistently improved diaphragmatic respiratory function by enhancing phrenic motor neuron (PhMN) innervation of the diaphragm neuromuscular junction and by increasing serotonergic innervation of PhMNs in ventral horn of the cervical spinal cord. These exciting findings demonstrate that local BDNF hydrogel delivery is a safe and robustly effective strategy to maintain respiratory function after cervical SCI.

Astrocyte progenitor transplantation promotes regeneration of bulbospinal respiratory axons, recovery of diaphragm function, and a reduced macrophage response following cervical spinal cord injury.

Stem/progenitor cell transplantation delivery of astrocytes is a potentially powerful strategy for spinal cord injury (SCI). Axon extension into SCI lesions that occur spontaneously or in response to experimental manipulations is often observed along endogenous astrocyte "bridges," suggesting that augmenting this response via astrocyte lineage transplantation can enhance axon regrowth. Given the importance of respiratory dysfunction post-SCI, we transplanted glial-restricted precursors (GRPs)-a class of lineage-restricted astrocyte progenitors-into the C2 hemisection model and evaluated effects on diaphragm function and the growth response of descending rostral ventral respiratory group (rVRG) axons that innervate phrenic motor neurons (PhMNs). GRPs survived long term and efficiently differentiated into astrocytes in injured spinal cord. GRPs promoted significant recovery of diaphragm electromyography amplitudes and stimulated robust regeneration of injured rVRG axons. Although rVRG fibers extended across the lesion, no regrowing axons re-entered caudal spinal cord to reinnervate PhMNs, suggesting that this regeneration response-although impressive-was not responsible for recovery. Within ipsilateral C3-5 ventral horn (PhMN location), GRPs induced substantial sprouting of spared fibers originating in contralateral rVRG and 5-HT axons that are important for regulating PhMN excitability; this sprouting was likely involved in functional effects of GRPs. Finally, GRPs reduced the macrophage response (which plays a key role in inducing axon retraction and limiting regrowth) both within the hemisection and at intact caudal spinal cord surrounding PhMNs. These findings demonstrate that astrocyte progenitor transplantation promotes significant plasticity of rVRG-PhMN circuitry and restoration of diaphragm function and suggest that these effects may be in part through immunomodulation.

A hydrogel engineered to deliver minocycline locally to the injured cervical spinal cord protects respiratory neural circuitry and preserves diaphragm function.

We tested a biomaterial-based approach to preserve the critical phrenic motor circuitry that controls diaphragm function by locally delivering minocycline hydrochloride (MH) following cervical spinal cord injury (SCI). MH is a clinically-available antibiotic and anti-inflammatory drug that targets a broad range of secondary injury mechanisms via its anti-inflammatory, anti-oxidant and anti-apoptotic properties. However, MH is only neuroprotective at high concentrations that cannot be achieved by systemic administration, which limits its clinical efficacy. We have developed a hydrogel-based MH delivery system that can be injected into the intrathecal space for local delivery of high concentrations of MH, without damaging spinal cord tissue. Implantation of MH hydrogel after unilateral level-C4/5 contusion SCI robustly preserved diaphragm function, as assessed by in vivo recordings of compound muscle action potential (CMAP) and electromyography (EMG) amplitudes. MH hydrogel also decreased lesion size and degeneration of cervical motor neuron somata, demonstrating its central neuroprotective effects within the injured cervical spinal cord. Furthermore, MH hydrogel significantly preserved diaphragm innervation by the axons of phrenic motor neurons (PhMNs), as assessed by both detailed neuromuscular junction (NMJ) morphological analysis and retrograde PhMN labeling from the diaphragm using cholera toxin B (CTB). In conclusion, our findings demonstrate that local MH hydrogel delivery to the injured cervical spinal cord is effective in preserving respiratory function after SCI by protecting the important neural circuitry that controls diaphragm activation.

Maladapted Prey Subsidize Predators and Facilitate Range Expansion.

Dispersal of prey from predator-free patches frequently supplies a trophic subsidy to predators by providing more prey than are produced locally. Prey arriving from predator-free patches might also have evolved weaker defenses against predators and thus enhance trophic subsidies by providing easily captured prey. Using local models assuming a linear or accelerating trade-off between defense and population growth rate, we demonstrate that immigration of undefended prey increased predator abundances and decreased defended prey through eco-evolutionary apparent competition. In individual-based models with spatial structure, explicit genetics, and gene flow along an environmental gradient, prey became maladapted to predators at the predator's range edge, and greater gene flow enhanced this maladaptation. The predator gained a subsidy from these easily captured prey, which enhanced its abundance, facilitated its persistence in marginal habitats, extended its range extent, and enhanced range shifts during environmental changes, such as climate change. Once the predator expanded, prey adapted to it and the advantage disappeared, resulting in an elastic predator range margin driven by eco-evolutionary dynamics. Overall, the results indicate a need to consider gene flow-induced maladaptation and species interactions as mutual forces that frequently determine ecological and evolutionary dynamics and patterns in nature.

Calcineurin Dysregulation Underlies Spinal Cord Injury-Induced K+ Channel Dysfunction in DRG Neurons.

Dysfunction of the fast-inactivating Kv3.4 potassium current in dorsal root ganglion (DRG) neurons contributes to the hyperexcitability associated with persistent pain induced by spinal cord injury (SCI). However, the underlying mechanism is not known. In light of our previous work demonstrating modulation of the Kv3.4 channel by phosphorylation, we investigated the role of the phosphatase calcineurin (CaN) using electrophysiological, molecular, and imaging approaches in adult female Sprague Dawley rats. Pharmacological inhibition of CaN in small-diameter DRG neurons slowed repolarization of the somatic action potential (AP) and attenuated the Kv3.4 current. Attenuated Kv3.4 currents also exhibited slowed inactivation. We observed similar effects on the recombinant Kv3.4 channel heterologously expressed in Chinese hamster ovary cells, supporting our findings in DRG neurons. Elucidating the molecular basis of these effects, mutation of four previously characterized serines within the Kv3.4 N-terminal inactivation domain eliminated the effects of CaN inhibition on the Kv3.4 current. SCI similarly induced concurrent Kv3.4 current attenuation and slowing of inactivation. Although there was little change in CaN expression and localization after injury, SCI induced upregulation of the native regulator of CaN 1 (RCAN1) in the DRG at the transcript and protein levels. Consistent with CaN inhibition resulting from RCAN1 upregulation, overexpression of RCAN1 in naive DRG neurons recapitulated the effects of pharmacological CaN inhibition on the Kv3.4 current and the AP. Overall, these results demonstrate a novel regulatory pathway that links CaN, RCAN1, and Kv3.4 in DRG neurons. Dysregulation of this pathway might underlie a peripheral mechanism of pain sensitization induced by SCI.SIGNIFICANCE STATEMENT Pain sensitization associated with spinal cord injury (SCI) involves poorly understood maladaptive modulation of neuronal excitability. Although central mechanisms have received significant attention, recent studies have identified peripheral nerve hyperexcitability as a driver of persistent pain signaling after SCI. However, the ion channels and signaling molecules responsible for this change in primary sensory neuron excitability are still not well defined. To address this problem, this study used complementary electrophysiological and molecular methods to determine how Kv3.4, a voltage-gated K+ channel robustly expressed in dorsal root ganglion neurons, becomes dysfunctional upon calcineurin (CaN) inhibition. The results strongly suggest that CaN inhibition underlies SCI-induced dysfunction of Kv3.4 and the associated excitability changes through upregulation of the native regulator of CaN 1 (RCAN1).

Range position and climate sensitivity: The structure of among-population demographic responses to climatic variation.

Species' distributions will respond to climate change based on the relationship between local demographic processes and climate and how this relationship varies based on range position. A rarely tested demographic prediction is that populations at the extremes of a species' climate envelope (e.g., populations in areas with the highest mean annual temperature) will be most sensitive to local shifts in climate (i.e., warming). We tested this prediction using a dynamic species distribution model linking demographic rates to variation in temperature and precipitation for wood frogs (Lithobates sylvaticus) in North America. Using long-term monitoring data from 746 populations in 27 study areas, we determined how climatic variation affected population growth rates and how these relationships varied with respect to long-term climate. Some models supported the predicted pattern, with negative effects of extreme summer temperatures in hotter areas and positive effects on recruitment for summer water availability in drier areas. We also found evidence of interacting temperature and precipitation influencing population size, such as extreme heat having less of a negative effect in wetter areas. Other results were contrary to predictions, such as positive effects of summer water availability in wetter parts of the range and positive responses to winter warming especially in milder areas. In general, we found wood frogs were more sensitive to changes in temperature or temperature interacting with precipitation than to changes in precipitation alone. Our results suggest that sensitivity to changes in climate cannot be predicted simply by knowing locations within the species' climate envelope. Many climate processes did not affect population growth rates in the predicted direction based on range position. Processes such as species-interactions, local adaptation, and interactions with the physical landscape likely affect the responses we observed. Our work highlights the need to measure demographic responses to changing climate.

Identification of second-generation P2X3 antagonists for treatment of pain.

A second-generation small molecule P2X3 receptor antagonist has been developed. The lead optimization strategy to address shortcomings of the first-generation preclinical lead compound is described herein. These studies were directed towards the identification and amelioration of preclinical hepatobiliary findings, reducing potential for drug-drug interactions, and decreasing the projected human dose of the first-generation lead.

The road to higher permanence and biodiversity in exurban wetlands.

Exurban areas are expanding throughout the world, yet their effects on local biodiversity remain poorly understood. Wetlands, in particular, face ongoing and substantial threats from exurban development. We predicted that exurbanization would reduce the diversity of wetland amphibian and invertebrate communities and that more spatially aggregated residential development would leave more undisturbed natural land, thereby promoting greater local diversity. Using structural equation models, we tested a series of predictions about the direct and indirect pathways by which exurbanization extent, spatial pattern, and wetland characteristics might affect diversity patterns in 38 wetlands recorded during a growing season. We used redundancy, indicator species, and nested community analyses to evaluate how exurbanization affected species composition. In contrast to expectations, we found higher diversity in exurban wetlands. We also found that housing aggregation did not significantly affect diversity. Exurbanization affected biodiversity indirectly by increasing roads and development, which promoted permanent wetlands with less canopy cover and more aquatic vegetation. These pond characteristics supported greater diversity. However, exurbanization was associated with fewer temporary wetlands and fewer of the species that depend on these habitats. Moreover, the best indicator species for an exurban wetland was the ram's head snail, a common disease vector in disturbed ponds. Overall, results suggest that exurbanization is homogenizing wetlands into more permanent water bodies. These more permanent, exurban ponds support higher overall animal diversity, but exclude temporary wetland specialists. Conserving the full assemblage of wetland species in expanding exurban regions throughout the world will require protecting and creating temporary wetlands.