Divergent changes in PBN excitability in a mouse model of neuropathic pain.

The transition from acute to chronic pain involves maladaptive plasticity in central nociceptive pathways. Growing evidence suggests that changes within the parabrachial nucleus (PBN), an important component of the spino-parabrachio-amygdaloid pain pathway, are key contributors to the development and maintenance of chronic pain. In animal models of chronic pain, PBN neurons become sensitive to normally innocuous stimuli and responses to noxious stimuli become amplified and more often produce after-discharges that outlast the stimulus. Using ex vivo slice electrophysiology and two mouse models of neuropathic pain, sciatic cuff and chronic constriction of the infraorbital nerve (CCI-ION), we find that changes in the firing properties of PBN neurons and a shift in inhibitory synaptic transmission may underlie this phenomenon. Compared to PBN neurons from shams, a larger proportion of PBN neurons from mice with a sciatic cuff were spontaneously active at rest, and these same neurons showed increased excitability relative to shams. In contrast, quiescent PBN neurons from cuff mice were less excitable than those from shams. Despite an increase in excitability in a subset of PBN neurons, the presence of after-discharges frequently observed in vivo were largely absent ex vivo in both injury models. However, GABAB-mediated presynaptic inhibition of GABAergic terminals is enhanced in PBN neurons after CCI-ION. These data suggest that the amplified activity of PBN neurons observed in rodent models of chronic pain arise through a combination of changes in firing properties and network excitability.Significance Statement Hyperactivity of neurons in the parabrachial nucleus (PBN) is causally linked to exaggerated pain behaviors in rodent models of chronic pain but the underlying mechanisms remain unknown. Using two mouse models of neuropathic pain, we show the intrinsic properties of PBN neurons are largely unaltered following injury. However, subsets of PBN neurons become more excitable and GABAB receptor mediated suppression of inhibitory terminals is enhanced after injury. Thus, shifts in network excitability may be a contributing factor in injury induced potentiation of PBN activity.

Elevated Serotonin in Mouse Spinal Dorsal Horn Is Pronociceptive.

Serotonergic neurons in the rostral ventral medulla (RVM) contribute to bidirectional control of pain through modulation of spinal and trigeminal nociceptive networks. Deficits in this pathway are believed to contribute to pathologic pain states, but whether changes in serotonergic mechanisms are pro- or antinociceptive is debated. We used a combination of optogenetics and fiber photometry to examine these mechanisms more closely. We find that optogenetic activation of RVM serotonergic afferents in the spinal cord of naive mice produces mechanical hypersensitivity and conditioned place aversion (CPA). Neuropathic pain, produced by chronic constriction injury of the infraorbital nerve (CCI-ION), evoked a tonic increase in serotonin (5HT) concentrations within the spinal trigeminal nucleus caudalis (SpVc), measured with liquid chromatography-tandem mass spectroscopy (LC-MS/MS). By contract, CCI-ION had no effect on the phasic serotonin transients in SpVc, evoked by noxious pinch, and measured with fiber photometry of a serotonin sensor. These findings suggest that serotonin release in the spinal cord is pronociceptive and that an increase in sustained serotonin signaling, rather than phasic or event driven increases, potentiate nociception in models of chronic pain.

Divergent changes in PBN excitability in a mouse model of neuropathic pain.

The transition from acute to chronic pain involves maladaptive plasticity in central nociceptive pathways. Growing evidence suggests that changes within the parabrachial nucleus (PBN), an important component of the spino-parabrachio-amygdaloid pain pathway, are key contributors to the development and maintenance of chronic pain. In animal models of chronic pain, PBN neurons become sensitive to normally innocuous stimuli and responses to noxious stimuli become amplified and more often produce after-discharges that outlast the stimulus. Using ex vivo slice electrophysiology and two mouse models of neuropathic pain, sciatic cuff and chronic constriction of the infraorbital nerve (CCI-ION), we find that changes in the firing properties of PBN neurons and a shift in inhibitory synaptic transmission may underlie this phenomenon. Compared to PBN neurons from shams, a larger proportion of PBN neurons from mice with a sciatic cuff were spontaneously active at rest, and these same neurons showed increased excitability relative to shams. In contrast, quiescent PBN neurons from cuff mice were less excitable than those from shams. Despite an increase in excitability in a subset of PBN neurons, the presence of after-discharges frequently observed in vivo were largely absent ex vivo in both injury models. However, GABAB-mediated presynaptic inhibition of GABAergic terminals is enhanced in PBN neurons after CCIION. These data suggest that the amplified activity of PBN neurons observed in rodent models of chronic pain arise through a combination of changes in firing properties and network excitability.

Elevated serotonin in mouse spinal dorsal horn is pronociceptive.

Serotonergic neurons in the rostral ventral medulla (RVM) contribute to bidirectional control of pain through modulation of spinal and trigeminal nociceptive networks. Deficits in this pathway are believed to contribute to pathological pain states, but whether changes in serotonergic mechanisms are pro or anti-nociceptive are debated. We used a combination of optogenetics and fiber photometry to examine these mechanisms more closely. We find that optogenetic activation of RVM serotonergic afferents in the spinal cord of naïve mice produces mechanical hypersensitivity and conditioned place aversion. Neuropathic pain, produced by chronic constriction injury of the infraorbital nerve (CCI-ION), evoked a tonic increase in serotonin concentrations within the spinal trigeminal nucleus caudalis (SpVc), measured with liquid chromatography-tandem mass spectroscopy (LC-MS/MS). By contract, CCI-ION had no effect on the phasic serotonin transients in SpVc, evoked by noxious pinch, and measured with fiber photometry of a serotonin sensor. These findings suggest that serotonin release in the spinal cord is pronociceptive and that an increase is sustained serotonin signaling, rather than phasic or event driven increases, potentiate nociception in models of chronic pain.

Parabrachial Nucleus Activity in Nociception and Pain in Awake Mice.

The parabrachial nuclear complex (PBN) is a nexus for aversion and for the sensory and affective components of pain perception. We have previously shown that during chronic pain PBN neurons in anesthetized rodents have amplified activity. We report a method to record from PBN neurons of behaving, head-restrained mice while applying reproducible noxious stimuli. We find that both spontaneous and evoked activity are higher in awake animals compared with urethane anesthetized mice. Fiber photometry of calcium responses from calcitonin-gene-related peptide-expressing PBN neurons demonstrates that these neurons respond to noxious stimuli. In both males and females with neuropathic or inflammatory pain, responses of PBN neurons remain amplified for at least 5 weeks, in parallel with increased pain metrics. We also show that PBN neurons can be rapidly conditioned to respond to innocuous stimuli after pairing with noxious stimuli. Finally, we demonstrate that changes in PBN neuronal activity are correlated with changes in arousal, measured as changes in pupil area.SIGNIFICANCE STATEMENT The parabrachial complex is a nexus of aversion, including pain. We report a method to record from parabrachial nucleus neurons of behaving mice while applying reproducible noxious stimuli. This allowed us to track parabrachial activity over time in animals with neuropathic or inflammatory pain. It also allowed us to show that the activity of these neurons correlates with arousal states and that these neurons can be conditioned to respond to innocuous stimuli.

Noradrenergic Input from Nucleus of the Solitary Tract Regulates Parabrachial Activity in Mice.

The parabrachial complex (PB) is critically involved in aversive processes, and chronic pain is associated with amplified activity of PB neurons in rodent models of neuropathic pain. Here, we demonstrate that catecholaminergic input from the caudal nucleus of the solitary tract (cNTScat), a stress responsive region that integrates interoceptive and exteroceptive signals, causes amplification of PB activity and their sensory afferents. We used a virally mediated expression of a norepinephrine (NE) sensor, NE2h, fiber photometry, and extracellular recordings in anesthetized mice to show that noxious mechanical and thermal stimuli activate cNTS neurons. These stimuli also produce prolonged NE transients in PB that far outlast the noxious stimuli. Similar NE transients can be evoked by focal electrical stimulation of cNTS, a region that contains the noradrenergic A2 cell group that projects densely on PB. In vitro, optical stimulation of cNTScat terminals depolarized PB neurons and caused a prolonged increase the frequency of excitatory synaptic activity. A dual opsin approach showed that sensory afferents from the caudal spinal trigeminal nucleus are potentiated by cNTScat terminal activation. This potentiation was coupled with a decrease in the paired pulse ratio (PPR), consistent with an cNTScat-mediated increase in the probability of release at SpVc synapses. Together, these data suggest that A2 neurons of the cNTS generate long lasting NE transients in PB which increase excitability and potentiate responses of PB neurons to sensory inputs. These reveal a mechanism through which stressors from multiple modalities may potentiate the aversiveness of nociceptive stimuli.

Parabrachial nucleus activity in nociception and pain in awake mice.

The parabrachial nuclear complex (PBN) is a nexus for aversion, and for the sensory and affective components of pain perception. We have previously shown that, during chronic pain, PBN neurons in anesthetized rodents have amplified activity. We report a method to record from PBN neurons of behaving, head-restrained mice, while applying reproducible noxious stimuli. We find that both spontaneous and evoked activity are higher in awake animals, compared to urethane anesthetized mice. Fiber photometry of calcium responses from CGRP-expressing PBN neurons demonstrates that these neurons respond to nociceptive stimuli. In both males and females with neuropathic or inflammatory pain, responses of PBN neurons remain amplified for at least 5 weeks, in parallel with increased pain metrics. We also show that PBN neurons can be rapidly conditioned to respond to innocuous stimuli, after pairing with nociceptive stimuli. Finally, we demonstrate that changes in PBN neuronal activity are correlated with changes in arousal, measured as changes in pupil diameter. Significance Statement: The parabrachial complex is a nexus of aversion, including pain. We report a method to record from parabrachial nucleus neurons of behaving mice, while applying reproducible noxious stimuli. This allowed, for the first time, tracking the activity of these neurons over time in animals with neuropathic or inflammatory pain. It also allowed us to show that the activity of these neurons correlates with arousal states, and that these neurons can be conditioned to respond to innocuous stimuli.

Limited role of generation time changes in driving the evolution of the mutation spectrum in humans.

Recent studies have suggested that the human germline mutation rate and spectrum evolve rapidly. Variation in generation time has been linked to these changes, though its contribution remains unclear. We develop a framework to characterize temporal changes in polymorphisms within and between populations, while controlling for the effects of natural selection and biased gene conversion. Application to the 1000 Genomes Project dataset reveals multiple independent changes that arose after the split of continental groups, including a previously reported, transient elevation in TCC>TTC mutations in Europeans and novel signals of divergence in C>Gand T>A mutation rates among population samples. We also find a significant difference between groups sampled in and outside of Africa in old T>C polymorphisms that predate the out-of-Africa migration. This surprising signal is driven by TpG>CpG mutations and stems in part from mis-polarized CpG transitions, which are more likely to undergo recurrent mutations. Finally, by relating the mutation spectrum of polymorphisms to parental age effects on de novo mutations, we show that plausible changes in the generation time cannot explain the patterns observed for different mutation types jointly. Thus, other factors - genetic modifiers or environmental exposures - must have had a non-negligible impact on the human mutation landscape.

Each human has 23 pairs of chromosomes, one set inherited from each parent. But the child's chromosomes are not an exact copy of their parents' chromosomes. Spontaneous changes or mutations in the DNA during the formation of the egg or sperm cells, or early development of the embryo, can change a small fraction of the nucleotides or 'letters' that make up the DNA. These modifications are an important source of genetic diversity in human populations and contribute to the evolution of new traits. Each genetic variant in present-day human populations represents a mutation in one of their ancestors. The types and frequencies of variants vary across human populations and have changed over time, suggesting that mutation patterns have evolved in the past. But the processes driving these population-level differences remain elusive. One possible factor may be changes in the average age of reproduction or the generation time in a population . For example, older parents contribute more ­ and also different types of ­ new mutations to their children than younger parents do. Populations, where it is customary to have children at older ages, may therefore have a different mutation landscape. To find out if this is indeed the case, Gao et al. used computer algorithms to analyze the genomes of hundreds of people living on three continents who participated in 'the 1,000 Genomes Project'. The analysis identified differences in mutation patterns across continental groups and estimated when these changes occurred. Further, they showed that although the age of reproduction had an impact on the mutation landscape, differences in generation time alone could not explain the observed changes in the human mutation spectrum. Factors other than generation time, such as environmental exposures, may have played a role in shifting these patterns. The study provides new insights into the changes in the mutation landscape over the course of human evolution. Mapping these patterns in humans worldwide may help scientists understand the causes underlying these changes. The techniques used by Gao et al. may also help analyze changes in mutation patterns in other organisms.

Molecular, Circuit, and Stress Response Characterization of Ventral Pallidum Npas1-Neurons.

Altered activity of the ventral pallidum (VP) underlies disrupted motivation in stress and drug exposure. The VP is a very heterogeneous structure composed of many neuron types with distinct physiological properties and projections. Neuronal PAS 1-positive (Npas1+) VP neurons are thought to send projections to brain regions critical for motivational behavior. While Npas1+ neurons have been characterized in the globus pallidus external, there is limited information on these neurons in the VP. To address this limitation, we evaluated the projection targets of the VP Npas1+ neurons and performed RNA-sequencing on ribosome-associated mRNA from VP Npas1+ neurons to determine their molecular identity. Finally, we used a chemogenetic approach to manipulate VP Npas1+ neurons during social defeat stress (SDS) and behavioral tasks related to anxiety and motivation in Npas1-Cre mice. We used a similar approach in females using the chronic witness defeat stress (CWDS). We identified VP Npas1+ projections to the nucleus accumbens, ventral tegmental area, medial and lateral habenula, lateral hypothalamus, thalamus, medial and lateral septum, and periaqueductal gray area. VP Npas1+ neurons displayed distinct translatome representing distinct biological processes. Chemogenetic activation of hM3D(Gq) receptors in VP Npas1+ neurons increased susceptibility to a subthreshold SDS and anxiety-like behavior in the elevated plus maze and open field while the activation of hM4D(Gi) receptors in VP Npas1+ neurons enhanced resilience to chronic SDS and CWDS. Thus, the activity of VP Npas1+ neurons modulates susceptibility to social stressors and anxiety-like behavior. Our studies provide new information on VP Npas1+ neuron circuitry, molecular identity, and their role in stress response.SIGNIFICANCE STATEMENT The ventral pallidum (VP) is a structure connected to both reward-related and aversive brain centers. It is a key brain area that signals the hedonic value of natural rewards. Disruption in the VP underlies altered motivation in stress and substance use disorder. However, VP is a very heterogeneous area with multiple neuron subtypes. This study characterized the projection pattern and molecular signatures of VP Neuronal PAS 1-positive (Npas1+) neurons. We further used tools to alter receptor signaling in VP Npas1+ neurons in stress to demonstrate a role for these neurons in stress behavioral outcomes. Our studies have implications for understanding brain cell type identities and their role in brain disorders, such as depression, a serious disorder that is precipitated by stressful events.

Medullary kappa-opioid receptor neurons inhibit pain and itch through a descending circuit.

In perilous and stressful situations, the ability to suppress pain can be critical for survival. The rostral ventromedial medulla contains neurons that robustly inhibit nocioception at the level of the spinal cord through a top-down modulatory pathway. Although much is known about the role of the rostral ventromedial medulla in the inhibition of pain, the precise ability to directly manipulate pain-inhibitory neurons in the rostral ventromedial medulla has never been achieved. We now expose a cellular circuit that inhibits nocioception and itch in mice. Through a combination of molecular, tracing and behavioural approaches, we found that rostral ventromedial medulla neurons containing the kappa-opioid receptor inhibit itch and nocioception. With chemogenetic inhibition, we uncovered that these neurons are required for stress-induced analgesia. Using intersectional chemogenetic and pharmacological approaches, we determined that rostral ventromedial medulla kappa-opioid receptor neurons inhibit nocioception and itch through a descending circuit. Lastly, we identified a dynorphinergic pathway arising from the periaqueductal grey that modulates nociception within the rostral ventromedial medulla. These discoveries highlight a distinct population of rostral ventromedial medulla neurons capable of broadly and robustly inhibiting itch and nocioception.

Single-cell multiomics defines tolerogenic extrathymic Aire-expressing populations with unique homology to thymic epithelium.

The autoimmune regulator (Aire), a well-defined transcriptional regulator in the thymus, is also found in extrathymic Aire-expressing cells (eTACs) in the secondary lymphoid organs. eTACs are hematopoietic antigen-presenting cells and inducers of immune tolerance, but their precise identity has remained unclear. Here, we use single-cell multiomics, transgenic murine models, and functional approaches to define eTACs at the transcriptional, genomic, and proteomic level. We find that eTACs consist of two similar cell types: CCR7+ Aire-expressing migratory dendritic cells (AmDCs) and an Airehi population coexpressing Aire and retinoic acid receptor­related orphan receptor γt (RORγt) that we term Janus cells (JCs). Both JCs and AmDCs have the highest transcriptional and genomic homology to CCR7+ migratory dendritic cells. eTACs, particularly JCs, have highly accessible chromatin and broad gene expression, including a range of tissue-specific antigens, as well as remarkable homology to medullary thymic epithelium and RANK-dependent Aire expression. Transgenic self-antigen expression by eTACs is sufficient to induce negative selection and prevent autoimmune diabetes. This transcriptional, genomic, and functional symmetry between eTACs (both JCs and AmDCs) and medullary thymic epithelium­the other principal Aire-expressing population and a key regulator of central tolerance­identifies a core program that may influence self-representation and tolerance across the spectrum of immune development.

Perinatal Fentanyl Exposure Leads to Long-Lasting Impairments in Somatosensory Circuit Function and Behavior.

One consequence of the opioid epidemic are lasting neurodevelopmental sequelae afflicting adolescents exposed to opioids in the womb. A translationally relevant and developmentally accurate preclinical model is needed to understand the behavioral, circuit, network, and molecular abnormalities resulting from this exposure. By employing a novel preclinical model of perinatal fentanyl exposure, our data reveal that fentanyl has several dose-dependent, developmental consequences to somatosensory function and behavior. Newborn male and female mice exhibit signs of withdrawal and sensory-related deficits that extend at least to adolescence. As fentanyl exposure does not affect dams' health or maternal behavior, these effects result from the direct actions of perinatal fentanyl on the pups' developing brain. At adolescence, exposed mice exhibit reduced adaptation to sensory stimuli, and a corresponding impairment in primary somatosensory (S1) function. In vitro electrophysiology demonstrates a long-lasting reduction in S1 synaptic excitation, evidenced by decreases in release probability, NMDA receptor-mediated postsynaptic currents, and frequency of miniature excitatory postsynaptic currents (mEPSCs), as well as increased frequency of miniature inhibitory postsynaptic currents (mIPSCs). In contrast, anterior cingulate cortical neurons exhibit an opposite phenotype, with increased synaptic excitation. Consistent with these changes, electrocorticograms (ECoGs) reveal suppressed ketamine-evoked γ oscillations. Morphologic analysis of S1 pyramidal neurons indicate reduced dendritic complexity, dendritic length, and soma size. Further, exposed mice exhibited abnormal cortical mRNA expression of key receptors involved in synaptic transmission and neuronal growth and development, changes that were consistent with the electrophysiological and morphologic changes. These findings demonstrate the lasting sequelae of perinatal fentanyl exposure on sensory processing and function.SIGNIFICANCE STATEMENT This is the first study to show that exposure to fentanyl in the womb results in behavioral, circuitry, and synaptic effects that last at least to adolescence. We also show, for the first time, that this exposure has different, lasting effects on synapses in different cortical areas.

Single-cell transcriptional profiling of human thymic stroma uncovers novel cellular heterogeneity in the thymic medulla.

The thymus' key function in the immune system is to provide the necessary environment for the development of diverse and self-tolerant T lymphocytes. While recent evidence suggests that the thymic stroma is comprised of more functionally distinct subpopulations than previously appreciated, the extent of this cellular heterogeneity in the human thymus is not well understood. Here we use single-cell RNA sequencing to comprehensively profile the human thymic stroma across multiple stages of life. Mesenchyme, pericytes and endothelial cells are identified as potential key regulators of thymic epithelial cell differentiation and thymocyte migration. In-depth analyses of epithelial cells reveal the presence of ionocytes as a medullary population, while the expression of tissue-specific antigens is mapped to different subsets of epithelial cells. This work thus provides important insight on how the diversity of thymic cells is established, and how this heterogeneity contributes to the induction of immune tolerance in humans.

Divergent profiles of fentanyl withdrawal and associated pain in mice and rats.

Opioid abuse has devastating effects on patients, their families, and society. Withdrawal symptoms are severely unpleasant, prolonged, and frequently hinder recovery or lead to relapse. The sharp increase in abuse and overdoses arising from the illicit use of potent and rapidly-acting synthetic opioids, such as fentanyl, highlights the urgency of understanding the withdrawal mechanisms related to these drugs. Progress is impeded by inconsistent reports on opioid withdrawal in different preclinical models. Here, using rats and mice of both sexes, we quantified withdrawal behaviors during spontaneous and naloxone-precipitated withdrawal, following two weeks of intermittent fentanyl exposure. We found that both mice and rats lost weight during exposure and showed increased signs of distress during spontaneous and naloxone precipitated withdrawal. However, these species differed in their expression of withdrawal associated pain, a key contributor to relapse in humans. Spontaneous or ongoing pain was preferentially expressed in rats in both withdrawal conditions, while no change was observed in mice. In contrast, withdrawal associated thermal hyperalgesia was found only in mice. These data suggest that rats and mice diverge in how they experience withdrawal and which aspects of the human condition they most accurately model. These differences highlight each species' strengths as model systems and can inform experimental design in studies of opioid withdrawal.

Control of synaptic transmission and neuronal excitability in the parabrachial nucleus.

The parabrachial nucleus (PB) is a hub for aversive behaviors, including those related to pain. We have shown that the expression of chronic pain is causally related to amplified activity of PB neurons, and to changes in synaptic inhibition of these neurons. These findings indicate that regulation of synaptic activity in PB may modulate pain perception and be involved in the pathophysiology of chronic pain. Here, we identify the roles in PB of signaling pathways that modulate synaptic functions. In pharmacologically isolated lateral PB neurons in acute mouse slices we find that baclofen, a GABAB receptor agonist, suppresses the frequency of miniature inhibitory and excitatory postsynaptic currents (mIPSCs and mEPSC). Activation of µ-opioid peptide receptors with DAMGO had similar suppressive effects on excitatory and inhibitory synapses, while the κ-opioid peptide receptor agonist U-69593 suppressed mIPSC release but had no consistent effects on mEPSCs. Activation of cannabinoid type 1 receptors with WIN 55,212-2 reduced the frequency of both inhibitory and excitatory synaptic events, while the CB1 receptor inverse agonist AM251 had opposite effects on mIPSC and mEPSC frequencies. AM251 increased the frequency of inhibitory events but led to a reduction in excitatory events through a GABAB mediated mechanism. Although none of the treatments produced a consistent effect on mIPSC or mEPSC amplitudes, baclofen and DAMGO both reliably activated a postsynaptic conductance. These results demonstrate that multiple signaling pathways can alter synaptic transmission and neuronal excitability in PB and provide a basis for investigating the contributions of these systems to the development and maintenance of chronic pain.

An Amygdalo-Parabrachial Pathway Regulates Pain Perception and Chronic Pain.

The parabrachial (PB) complex mediates both ascending nociceptive signaling and descending pain modulatory information in the affective/emotional pain pathway. We have recently reported that chronic pain is associated with amplified activity of PB neurons in a rat model of neuropathic pain. Here we demonstrate that similar activity amplification occurs in mice, and that this is related to suppressed inhibition to lateral parabrachial (LPB) neurons from the CeA in animals of either sex. Animals with pain after chronic constriction injury of the infraorbital nerve (CCI-Pain) displayed higher spontaneous and evoked activity in PB neurons, and a dramatic increase in after-discharges, responses that far outlast the stimulus, compared with controls. LPB neurons in CCI-Pain animals showed a reduction in inhibitory, GABAergic inputs. We show that, in both rats and mice, LPB contains few GABAergic neurons, and that most of its GABAergic inputs arise from CeA. These CeA GABA neurons express dynorphin, somatostatin, and/or corticotropin releasing hormone. We find that the efficacy of this CeA-LPB pathway is suppressed in chronic pain. Further, optogenetically stimulating this pathway suppresses acute pain, and inhibiting it, in naive animals, evokes pain behaviors. These findings demonstrate that the CeA-LPB pathway is critically involved in pain regulation, and in the pathogenesis of chronic pain.SIGNIFICANCE STATEMENT We describe a novel pathway, consisting of inhibition by dynorphin, somatostatin, and corticotropin-releasing hormone-expressing neurons in the CeA that project to the parabrachial nucleus. We show that this pathway regulates the activity of pain-related neurons in parabrachial nucleus, and that, in chronic pain, this inhibitory pathway is suppressed, and that this suppression is causally related to pain perception. We propose that this amygdalo-parabrachial pathway is a key regulator of both chronic and acute pain, and a novel target for pain relief.

Chronic Exposure to High Altitude: Synaptic, Astroglial and Memory Changes.

Long-term operations carried out at high altitude (HA) by military personnel, pilots, and astronauts may trigger health complications. In particular, chronic exposure to high altitude (CEHA) has been associated with deficits in cognitive function. In this study, we found that mice exposed to chronic HA (5000 m for 12 weeks) exhibited deficits in learning and memory associated with hippocampal function and were linked with changes in the expression of synaptic proteins across various regions of the brain. Specifically, we found decreased levels of synaptophysin (SYP) (p < 0.05) and spinophilin (SPH) (p < 0.05) in the olfactory cortex, post synaptic density-95 (PSD-95) (p < 0.05), growth associated protein 43 (GAP43) (p < 0.05), glial fibrillary acidic protein (GFAP) (p < 0.05) in the cerebellum, and SYP (p < 0.05) and PSD-95 (p < 0.05) in the brainstem. Ultrastructural analyses of synaptic density and morphology in the hippocampus did not reveal any differences in CEHA mice compared to SL mice. Our data are novel and suggest that CEHA exposure leads to cognitive impairment in conjunction with neuroanatomically-based molecular changes in synaptic protein levels and astroglial cell marker in a region specific manner. We hypothesize that these new findings are part of highly complex molecular and neuroplasticity mechanisms underlying neuroadaptation response that occurs in brains when chronically exposed to HA.

Neuronal and vascular deficits following chronic adaptation to high altitude.

We sought to understand the mechanisms underlying cognitive deficits that are reported to affect non-native subjects following their prolonged stay and/or work at high altitude (HA). We found that mice exposed to a simulated environment of 5000 m exhibit deficits in hippocampal learning and memory accompanied by abnormalities in brain MR imaging. Exposure (1-8 months) to HA led to an increase in brain ventricular volume, a reduction in relative cerebral blood flow and changes in diffusion tensor imaging (DTI) derived parameters within the hippocampus and corpus callosum. Furthermore, neuropathological examination revealed significant expansion of the neurovascular network, microglia activation and demyelination within the corpus callosum. Electrophysiological recordings from the corpus callosum indicated that axonal excitabilities are increased while refractory periods are longer despite a lack of change in action potential conduction velocities of both myelinated and unmyelinated fibers. Next generation RNA-sequencing identified alterations in hippocampal and amygdala transcriptome signaling pathways linked to angiogenesis, neuroinflammation and myelination. Our findings reveal that exposure to hypobaric-hypoxia triggers maladaptive responses inducing cognitive deficits and suggest potential mechanisms underlying the adverse impacts of staying or traveling at high altitude.

Experimental Traumatic Brain Injury Identifies Distinct Early and Late Phase Axonal Conduction Deficits of White Matter Pathophysiology, and Reveals Intervening Recovery.

Traumatic brain injury (TBI) patients often exhibit slowed information processing speed that can underlie diverse symptoms. Processing speed depends on neural circuit function at synapses, in the soma, and along axons. Long axons in white matter (WM) tracts are particularly vulnerable to TBI. We hypothesized that disrupted axon-myelin interactions that slow or block action potential conduction in WM tracts may contribute to slowed processing speed after TBI. Concussive TBI in male/female mice was used to produce traumatic axonal injury in the corpus callosum (CC), similar to WM pathology in human TBI cases. Compound action potential velocity was slowed along myelinated axons at 3 d after TBI with partial recovery by 2 weeks, suggesting early demyelination followed by remyelination. Ultrastructurally, dispersed demyelinated axons and disorganized myelin attachment to axons at paranodes were apparent within CC regions exhibiting traumatic axonal injury. Action potential conduction is exquisitely sensitive to paranode abnormalities. Molecular identification of paranodes and nodes of Ranvier detected asymmetrical paranode pairs and abnormal heminodes after TBI. Fluorescent labeling of oligodendrocyte progenitors in NG2CreER;mTmG mice showed increased synthesis of new membranes extended along axons to paranodes, indicating remyelination after TBI. At later times after TBI, an overall loss of conducting axons was observed at 6 weeks followed by CC atrophy at 8 weeks. These studies identify a progression of both myelinated axon conduction deficits and axon-myelin pathology in the CC, implicating WM injury in impaired information processing at early and late phases after TBI. Furthermore, the intervening recovery reveals a potential therapeutic window.SIGNIFICANCE STATEMENT Traumatic brain injury (TBI) is a major global health concern. Across the spectrum of TBI severities, impaired information processing can contribute to diverse functional deficits that underlie persistent symptoms. We used experimental TBI to exploit technical advantages in mice while modeling traumatic axonal injury in white matter tracts, which is a key pathological feature of human TBI. A combination of approaches revealed slowed and failed signal conduction along with damage to the structure and molecular composition of myelinated axons in the white matter after TBI. An early regenerative response was not sustained yet reveals a potential time window for intervention. These insights into white matter abnormalities underlying axon conduction deficits can inform strategies to improve treatment options for TBI patients.

Down Syndrome Developmental Brain Transcriptome Reveals Defective Oligodendrocyte Differentiation and Myelination.

Trisomy 21, or Down syndrome (DS), is the most common genetic cause of developmental delay and intellectual disability. To gain insight into the underlying molecular and cellular pathogenesis, we conducted a multi-region transcriptome analysis of DS and euploid control brains spanning from mid-fetal development to adulthood. We found genome-wide alterations in the expression of a large number of genes, many of which exhibited temporal and spatial specificity and were associated with distinct biological processes. In particular, we uncovered co-dysregulation of genes associated with oligodendrocyte differentiation and myelination that were validated via cross-species comparison to Ts65Dn trisomy mice. Furthermore, we show that hypomyelination present in Ts65Dn mice is in part due to cell-autonomous effects of trisomy on oligodendrocyte differentiation and results in slower neocortical action potential transmission. Together, these results identify defects in white matter development and function in DS, and they provide a transcriptional framework for further investigating DS neuropathogenesis.