RESUMO
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.
Assuntos
Dor Crônica , Núcleos Parabraquiais , Masculino , Feminino , Camundongos , Animais , Núcleos Parabraquiais/fisiologia , Nociceptividade , Vigília , Peptídeo Relacionado com Gene de Calcitonina/metabolismoRESUMO
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.
Assuntos
Prosencéfalo Basal , Feminino , Camundongos , Animais , Prosencéfalo Basal/fisiologia , Neurônios/fisiologia , Área Tegmentar Ventral/fisiologia , Núcleo Accumbens/metabolismo , Recompensa , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genéticaRESUMO
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.
Assuntos
Bulbo , Neurônios , Dor , Prurido , Receptores Opioides kappa , Animais , Bulbo/citologia , Camundongos , Neurônios/fisiologia , Dor/fisiopatologia , Prurido/fisiopatologia , Receptores Opioides kappa/metabolismoRESUMO
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.
Assuntos
Analgésicos Opioides/toxicidade , Potenciais Somatossensoriais Evocados , Fentanila/toxicidade , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Potenciais Sinápticos , Adaptação Fisiológica , Animais , Comportamento Animal , Feminino , Ritmo Gama , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurogênese , Percepção , Gravidez , Células Piramidais/efeitos dos fármacos , Células Piramidais/patologia , Células Piramidais/fisiologia , Córtex Somatossensorial/citologia , Córtex Somatossensorial/efeitos dos fármacos , Córtex Somatossensorial/crescimento & desenvolvimentoRESUMO
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.
Assuntos
Tonsila do Cerebelo/fisiopatologia , Dor Crônica/fisiopatologia , Neuralgia/fisiopatologia , Percepção da Dor/fisiologia , Núcleos Parabraquiais/fisiopatologia , Potenciais de Ação/fisiologia , Animais , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Vias Neurais/fisiopatologia , Neurônios/fisiologia , Medição da Dor , Limiar da Dor/fisiologiaRESUMO
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.
Assuntos
Potenciais de Ação , Axônios/fisiologia , Lesões Encefálicas Traumáticas/fisiopatologia , Bainha de Mielina/fisiologia , Substância Branca/fisiopatologia , Animais , Lesões Encefálicas Traumáticas/patologia , Corpo Caloso/patologia , Corpo Caloso/fisiopatologia , Modelos Animais de Doenças , Feminino , Masculino , Camundongos Endogâmicos C57BL , Bainha de Mielina/patologia , Bainha de Mielina/ultraestrutura , Oligodendroglia/patologia , Oligodendroglia/fisiologia , Substância Branca/patologia , Substância Branca/ultraestruturaRESUMO
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.
RESUMO
Due to their interactions with the neurovasculature, microglia are implicated in maladaptive responses to hypobaric hypoxia at high altitude (HA). To explore these interactions at HA, pharmacological depletion of microglia with the colony-stimulating factor-1 receptor inhibitor, PLX5622, was employed in male C57BL/6J mice maintained at HA or sea level (SL) for 3-weeks, followed by assessment of ex-vivo hippocampal long-term potentiation (LTP), fear memory recall and microglial dynamics/physiology. Our findings revealed that microglia depletion decreased LTP and reduced glucose levels by 25% at SL but did not affect fear memory recall. At HA, the absence of microglia did not significantly alter HA associated deficits in fear memory or HA mediated decreases in peripheral glucose levels. In regard to microglial dynamics in the cortex, HA enhanced microglial surveillance activity, ablation of microglia resulted in increased chemotactic responses and decreased microglia tip proliferation during ball formation. In contrast, vessel ablation increased cortical microglia tip path tortuosity. In the hippocampus, changes in microglial dynamics were only observed in response to vessel ablation following HA. As the hippocampus is critical for learning and memory, poor hippocampal microglial context-dependent adaptation may be responsible for some of the enduring neurological deficits associated with HA.
Assuntos
Altitude , Cognição , Hipocampo , Potenciação de Longa Duração , Camundongos Endogâmicos C57BL , Microglia , Neurônios , Animais , Microglia/metabolismo , Microglia/fisiologia , Masculino , Camundongos , Hipocampo/metabolismo , Cognição/fisiologia , Neurônios/fisiologia , Neurônios/metabolismo , Aclimatação/fisiologia , Medo/fisiologia , Memória/fisiologia , Glucose/metabolismo , Compostos OrgânicosRESUMO
Denervation-induced plastic changes impair locomotor recovery after spinal cord injury (SCI). Spinal motoneurons become hyperexcitable after SCI, but the plastic responses of locomotor network interneurons (INs) after SCI have not been studied. Using an adult mouse SCI model, we analyzed the effects of complete spinal cord lesions on the intrinsic electrophysiological properties, excitability, and neuromodulatory responses to serotonin (5-HT) in mouse lumbar V2a spinal INs, which help regulate left-right alternation during locomotion. Four weeks after SCI, V2a INs showed almost no changes in baseline excitability or action potential properties; the only parameter that changed was a reduced input resistance. However, V2a INs became 100- to 1000-fold more sensitive to 5-HT. Immunocytochemical analysis showed that SCI caused a coordinated loss of serotonergic fibers and the 5-HT transporter (SERT). Blocking the SERT with citalopram in intact mice did not increase 5-HT sensitivity to the level seen after SCI. SCI also evoked an increase in 5-HT(2C) receptor cluster number and intensity, suggesting that several plastic changes cooperate in increasing 5-HT sensitivity. Our results suggest that different components of the spinal neuronal network responsible for coordinating locomotion are differentially affected by SCI, and highlight the importance of understanding these changes when considering therapies targeted at functional recovery.
Assuntos
Fenômenos Biofísicos/efeitos dos fármacos , Interneurônios/efeitos dos fármacos , Potenciais da Membrana/efeitos dos fármacos , Serotonina/farmacologia , Traumatismos da Medula Espinal/patologia , Medula Espinal/patologia , Animais , Citalopram/farmacologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Proteínas de Homeodomínio/genética , Técnicas In Vitro , Locomoção/efeitos dos fármacos , Locomoção/fisiologia , Masculino , Camundongos , Camundongos Transgênicos , Técnicas de Patch-Clamp , Receptor 5-HT2C de Serotonina/metabolismo , Serotonina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Inibidores Seletivos de Recaptação de Serotonina/farmacologia , Medula Espinal/metabolismo , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Estatísticas não Paramétricas , Fatores de Tempo , Fatores de Transcrição/genéticaRESUMO
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.
Assuntos
Mutação em Linhagem Germinativa , Taxa de Mutação , Humanos , Mutação , Genoma , Seleção GenéticaRESUMO
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.
Assuntos
Dor Crônica , Núcleo Solitário , Camundongos , Animais , Neurônios/fisiologia , Nervo Vago , NorepinefrinaRESUMO
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.
RESUMO
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.
Assuntos
Neuralgia , Serotonina , Camundongos , Animais , Serotonina/metabolismo , Hiperalgesia/metabolismo , Cromatografia Líquida , Espectrometria de Massas em Tandem , Corno Dorsal da Medula Espinal , Medula Espinal/metabolismo , Neuralgia/metabolismoRESUMO
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.
RESUMO
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.
RESUMO
Down syndrome (DS) is caused by the overexpression of genes on triplicated regions of human chromosome 21 (Hsa21). While the resulting physiological and behavioral phenotypes vary in their penetrance and severity, all individuals with DS have variable but significant levels of cognitive disability. At the core of cognitive processes is the phenomenon of synaptic plasticity, a functional change in the strength at points of communication between neurons. A wide variety of evidence from studies on DS individuals and mouse models of DS indicates that synaptic plasticity is adversely affected in human trisomy 21 and mouse segmental trisomy 16, respectively, an outcome that almost certainly extensively contributes to the cognitive impairments associated with DS. In this review, we will highlight some of the neurophysiological changes that we believe reduce the ability of trisomic neurons to undergo neuroplasticity-related adaptations. We will focus primarily on hippocampal networks which appear to be particularly impacted in DS and where consequently the majority of cellular and neuronal network research has been performed using DS animal models, in particular the Ts65Dn mouse. Finally, we will postulate on how altered plasticity may contribute to the DS cognitive disability.
Assuntos
Transtornos Cognitivos/fisiopatologia , Síndrome de Down/fisiopatologia , Hipocampo/fisiopatologia , Plasticidade Neuronal/fisiologia , Sinapses/fisiologia , Animais , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/psicologia , Transtornos Cognitivos/terapia , Terapia Cognitivo-Comportamental , Síndrome de Down/complicações , Síndrome de Down/psicologia , Síndrome de Down/terapia , Humanos , Camundongos , Modelos Neurológicos , Transdução de Sinais/fisiologiaRESUMO
It has been very difficult to record from interneurons in acute slices of the lumbar spinal cord from mice >3 wk of age. The low success rate and short recording times limit in vitro experimentation on mouse spinal networks to neonatal and early postnatal periods when locomotor networks are still developmentally immature. To overcome this limitation and enable investigation of mature locomotor network neurons, we have established a reliable procedure to record from spinal cord neurons in slices from adult, behaviorally mature mice of any age. Two key changes to the established neonate procedure were implemented. First, we remove the cord by a dorsal laminectomy from a deeply anesthetized animal. This enables respiration and other vital functions to continue up to the moment the maximally oxygenated lumbar spinal cord is removed, improving the health of the slices. Second, since adult spinal cord interneurons appear more sensitive to the intracellular dialysis that occurs during whole cell recordings, we introduced perforated patch recordings to the procedure. Stable recordings up to 12 h in duration were obtained with our new method. This will allow investigation of changes in mature neuronal properties in disease states or after spinal cord injury and allow prolonged recordings of responses to drug application that were previously impossible.
Assuntos
Interneurônios/fisiologia , Técnicas de Cultura de Órgãos/métodos , Técnicas de Patch-Clamp/métodos , Medula Espinal/citologia , Medula Espinal/fisiologia , Potenciais de Ação/fisiologia , Fatores Etários , Anestesia , Animais , Dissecação/métodos , Laminectomia/métodos , Vértebras Lombares , Camundongos , Medula Espinal/crescimento & desenvolvimento , Fatores de TempoRESUMO
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.
RESUMO
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.
Assuntos
Analgésicos Opioides/efeitos adversos , Fentanila/efeitos adversos , Dor/metabolismo , Síndrome de Abstinência a Substâncias/metabolismo , Analgésicos Opioides/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Feminino , Fentanila/farmacologia , Humanos , Hiperalgesia/induzido quimicamente , Locomoção/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Entorpecentes/efeitos adversos , Entorpecentes/farmacologia , Dor/tratamento farmacológico , Ratos , Ratos Wistar , Síndrome de Abstinência a Substâncias/tratamento farmacológicoRESUMO
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.