RESUMO
Placebo effects are notable demonstrations of mind-body interactions1,2. During pain perception, in the absence of any treatment, an expectation of pain relief can reduce the experience of pain-a phenomenon known as placebo analgesia3-6. However, despite the strength of placebo effects and their impact on everyday human experience and the failure of clinical trials for new therapeutics7, the neural circuit basis of placebo effects has remained unclear. Here we show that analgesia from the expectation of pain relief is mediated by rostral anterior cingulate cortex (rACC) neurons that project to the pontine nucleus (rACCâPn)-a precerebellar nucleus with no established function in pain. We created a behavioural assay that generates placebo-like anticipatory pain relief in mice. In vivo calcium imaging of neural activity and electrophysiological recordings in brain slices showed that expectations of pain relief boost the activity of rACCâPn neurons and potentiate neurotransmission in this pathway. Transcriptomic studies of Pn neurons revealed an abundance of opioid receptors, further suggesting a role in pain modulation. Inhibition of the rACCâPn pathway disrupted placebo analgesia and decreased pain thresholds, whereas activation elicited analgesia in the absence of placebo conditioning. Finally, Purkinje cells exhibited activity patterns resembling those of rACCâPn neurons during pain-relief expectation, providing cellular-level evidence for a role of the cerebellum in cognitive pain modulation. These findings open the possibility of targeting this prefrontal cortico-ponto-cerebellar pathway with drugs or neurostimulation to treat pain.
Assuntos
Analgesia , Giro do Cíngulo , Dor , Efeito Placebo , Camundongos , Animais , Masculino , Giro do Cíngulo/fisiopatologia , Dor/fisiopatologia , Vias Neurais , Neurônios/fisiologia , Feminino , Células de Purkinje/fisiologia , Manejo da Dor/métodos , Receptores Opioides/metabolismo , Camundongos Endogâmicos C57BL , Limiar da Dor , Transmissão Sináptica , Percepção da Dor/fisiologiaRESUMO
Opioids are potent analgesics broadly used for pain management; however, they can produce dangerous side effects including addiction and respiratory depression. These harmful effects have led to an epidemic of opioid abuse and overdose deaths, creating an urgent need for the development of both safer pain medications and treatments for opioid use disorders. Both the analgesic and addictive properties of opioids are mediated by the mu opioid receptor (MOR), making resolution of the cell types and neural circuits responsible for each of the effects of opioids a critical research goal. Single-cell RNA sequencing (scRNA-seq) technology is enabling the identification of MOR-expressing cell types throughout the nervous system, creating new opportunities for mapping distinct opioid effects onto newly discovered cell types. Here, we describe molecularly defined MOR-expressing neuronal cell types throughout the peripheral and central nervous systems and their potential contributions to opioid analgesia and addiction.
Assuntos
Analgésicos Opioides , Transtornos Relacionados ao Uso de Opioides , Humanos , Analgésicos Opioides/efeitos adversos , Receptores Opioides mu/genética , Receptores Opioides mu/metabolismo , Dor/metabolismo , Analgésicos , Transtornos Relacionados ao Uso de Opioides/epidemiologiaRESUMO
Axon regeneration after spinal cord injury (SCI) is limited by both a decreased intrinsic ability of neurons to grow axons and the growth-hindering effects of extrinsic inhibitory molecules expressed around the lesion. Deletion of phosphatase and tensin homolog (Pten) augments mechanistic target of rapamycin (mTOR) signaling and enhances the intrinsic regenerative response of injured corticospinal neurons after SCI. Because of the variety of growth-restrictive extrinsic molecules, it remains unclear how inhibition of conserved inhibitory signaling elements would affect axon regeneration and rewiring after SCI. Moreover, it remains unknown how a combinatorial approach to modulate both extrinsic and intrinsic mechanisms can enhance regeneration and rewiring after SCI. In the present study, we deleted RhoA and RhoC, which encode small GTPases that mediate growth inhibition signals of a variety of extrinsic molecules, to remove global extrinsic pathways. RhoA/RhoC double deletion in mice suppressed retraction or dieback of corticospinal axons after SCI. In contrast, Pten deletion increased regrowth of corticospinal axons into the lesion core. Although deletion of both RhoA and Pten did not promote axon regrowth across the lesion or motor recovery, it additively promoted rewiring of corticospinal circuits connecting the cerebral cortex, spinal cord, and hindlimb muscles. Our genetic findings, therefore, reveal that a combinatorial approach to modulate both intrinsic and extrinsic factors can additively promote neural circuit rewiring after SCI.SIGNIFICANCE STATEMENT SCI often causes severe motor deficits because of damage to the corticospinal tract (CST), the major neural pathway for voluntary movements. Regeneration of CST axons is required to reconstruct motor circuits and restore functions; however, a lower intrinsic ability to grow axons and extrinsic inhibitory molecules severely limit axon regeneration in the CNS. Here, we investigated whether suppression of extrinsic inhibitory cues by genetic deletion of Rho as well as enhancement of the intrinsic pathway by deletion of Pten could enable axon regrowth and rewiring of the CST after SCI. We show that simultaneous elimination of extrinsic and intrinsic signaling pathways can additively promote axon sprouting and rewiring of the corticospinal circuits. Our data demonstrate a potential molecular approach to reconstruct motor pathways after SCI.
Assuntos
Regeneração Nervosa/fisiologia , PTEN Fosfo-Hidrolase/metabolismo , Tratos Piramidais/fisiopatologia , Traumatismos da Medula Espinal/fisiopatologia , Proteínas rho de Ligação ao GTP/metabolismo , Animais , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Peripheral nerve injury induces long-term pro-inflammatory responses in spinal cord glial cells that facilitate neuropathic pain, but the identity of endogenous cells that resolve spinal inflammation has not been determined. Guided by single-cell RNA sequencing (scRNA-seq), we found that MRC1+ spinal cord macrophages proliferated and upregulated the anti-inflammatory mediator Cd163 in mice following superficial injury (SI; nerve intact), but this response was blunted in nerve-injured animals. Depleting spinal macrophages in SI animals promoted microgliosis and caused mechanical hypersensitivity to persist. Conversely, expressing Cd163 in spinal macrophages increased Interleukin 10 expression, attenuated micro- and astrogliosis, and enduringly alleviated mechanical and thermal hypersensitivity in nerve-injured animals. Our data indicate that MRC1+ spinal macrophages actively restrain glia to limit neuroinflammation and resolve mechanical pain following a superficial injury. Moreover, we show that spinal macrophages from nerve-injured animals mount a dampened anti-inflammatory response but can be therapeutically coaxed to promote long-lasting recovery of neuropathic pain.
Assuntos
Hiperalgesia/metabolismo , Macrófagos/fisiologia , Neuralgia/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Medula Espinal/metabolismo , Animais , Modelos Animais de Doenças , Inflamação/metabolismo , Camundongos , Nociceptividade/fisiologia , Medição da DorRESUMO
Axon regeneration is limited in the central nervous system, which hinders the reconstruction of functional circuits following spinal cord injury (SCI). Although various extrinsic molecules to repel axons following SCI have been identified, the role of semaphorins, a major class of axon guidance molecules, has not been thoroughly explored. Here we show that expression of semaphorins, including Sema5a and Sema6d, is elevated after SCI, and genetic deletion of either molecule or their receptors (neuropilin1 and plexinA1, respectively) suppresses axon retraction or dieback in injured corticospinal neurons. We further show that Olig2+ cells are essential for SCI-induced semaphorin expression, and that Olig2 binds to putative enhancer regions of the semaphorin genes. Finally, conditional deletion of Olig2 in the spinal cord reduces the expression of semaphorins, alleviating the axon retraction. These results demonstrate that semaphorins function as axon repellents following SCI, and reveal a novel transcriptional mechanism for controlling semaphorin levels around injured neurons to create zones hostile to axon regrowth.
Assuntos
Regulação da Expressão Gênica/fisiologia , Regeneração Nervosa/fisiologia , Fator de Transcrição 2 de Oligodendrócitos/metabolismo , Semaforinas/biossíntese , Traumatismos da Medula Espinal/metabolismo , Animais , Axônios/patologia , Camundongos , Camundongos Endogâmicos C57BL , Tratos Piramidais/lesões , Tratos Piramidais/metabolismo , Traumatismos da Medula Espinal/patologiaRESUMO
Topoisomerase 1 (TOP1) relieves torsional stress in DNA during transcription and facilitates the expression of long (>100 kb) genes, many of which are important for neuronal functions. To evaluate how loss of Top1 affected neurons in vivo, we conditionally deleted (cKO) Top1 in postmitotic excitatory neurons in the mouse cerebral cortex and hippocampus. Top1 cKO neurons develop properly, but then show biased transcriptional downregulation of long genes, signs of DNA damage, neuroinflammation, increased poly(ADP-ribose) polymerase-1 (PARP1) activity, single-cell somatic mutations, and ultimately degeneration. Supplementation of nicotinamide adenine dinucleotide (NAD+) with nicotinamide riboside partially blocked neurodegeneration, and increased the lifespan of Top1 cKO mice by 30%. A reduction of p53 also partially rescued cortical neuron loss. While neurodegeneration was partially rescued, behavioral decline was not prevented. These data indicate that reducing neuronal loss is not sufficient to limit behavioral decline when TOP1 function is disrupted.
Assuntos
DNA Topoisomerases Tipo I/deficiência , Instabilidade Genômica , Doenças Neurodegenerativas/enzimologia , Neurônios/enzimologia , Animais , Apoptose/efeitos dos fármacos , Córtex Cerebral/enzimologia , Córtex Cerebral/patologia , Dano ao DNA , DNA Topoisomerases Tipo I/genética , Hipocampo/enzimologia , Hipocampo/patologia , Inflamação , Camundongos , Camundongos Knockout , Mortalidade Prematura , Atividade Motora , Mutação , NAD/administração & dosagem , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/patologia , Doenças Neurodegenerativas/fisiopatologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Niacinamida/administração & dosagem , Niacinamida/análogos & derivados , Poli(ADP-Ribose) Polimerase-1/metabolismo , Compostos de PiridínioRESUMO
Subsets of small-diameter dorsal root ganglia (DRG) neurons detect pruritogenic (itch-causing) and algogenic (pain-causing) stimuli and can be activated or sensitized by chemical mediators. Many of these chemical mediators activate receptors that are coupled to lipid hydrolysis and diacylglycerol (DAG) production. Diacylglycerol kinase iota (DGKI) can phosphorylate DAG and is expressed at high levels in small-diameter mouse DRG neurons. Given the importance of these neurons in sensing pruritogenic and algogenic chemicals, we sought to determine if loss of DGKI impaired responses to itch- or pain-producing stimuli. Using male and female Dgki-knockout mice, we found that in vivo sensitivity to histamine-but not other pruritogens-was enhanced. In contrast, baseline pain sensitivity and pain sensitization following inflammatory or neuropathic injury were equivalent between wild type and Dgki-/- mice. In vitro calcium responses in DRG neurons to histamine was enhanced, while responses to algogenic ligands were unaffected by Dgki deletion. These data suggest Dgki regulates sensory neuron and behavioral responses to histamine, without affecting responses to other pruritogenic or algogenic agents.
Assuntos
Diacilglicerol Quinase/deficiência , Histamina/efeitos adversos , Prurido/induzido quimicamente , Prurido/enzimologia , Animais , Comportamento Animal , Cálcio/farmacologia , Diacilglicerol Quinase/metabolismo , Modelos Animais de Doenças , Feminino , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Inflamação/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nociceptividade , Dor/enzimologia , Dor/patologia , Dor/fisiopatologia , Prurido/patologia , Prurido/fisiopatologia , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/patologiaRESUMO
The development of the mammalian cerebral cortex depends on careful orchestration of proliferation, maturation, and migration events, ultimately giving rise to a wide variety of neuronal and non-neuronal cell types. To better understand cellular and molecular processes that unfold during late corticogenesis, we perform single-cell RNA-seq on the mouse cerebral cortex at a progenitor driven phase (embryonic day 14.5) and at birth-after neurons from all six cortical layers are born. We identify numerous classes of neurons, progenitors, and glia, their proliferative, migratory, and activation states, and their relatedness within and across age. Using the cell-type-specific expression patterns of genes mutated in neurological and psychiatric diseases, we identify putative disease subtypes that associate with clinical phenotypes. Our study reveals the cellular template of a complex neurodevelopmental process, and provides a window into the cellular origins of brain diseases.