Patrik Ernfors

Although inflammatory processes in rheumatoid arthritis have been described, mechanisms driving pain are poorly defined. Here, we used a multitude of approaches to uncover the neural basis and causes of inflammatory pain. We show in mice with cartilage autoantibody-induced arthritis that early immune activation and a cytokine storm were mainly driven by vascular cells and monocytes/macrophages in the dorsal root ganglion. However, persistently elevated interferons and receptor activation of the MNK1/MNK2–eIF4E signaling pathway at all disease phases caused sensory–motor dysfunction and pain by inducing hyperexcitability and sensitization of a GFRA3 ⁺ C-fiber subtype of joint-innervating sensory neurons. Signaling pathway inhibition in vivo reversed pain and restored limb function. Like mice, human sensory neurons expressed interferon receptors, and type 1 interferons and signaling were increased only in individuals with painful rheumatoid arthritis. The finding that joint pain before and during arthritis is caused by a defined cytokine and signaling pathway holds promise for targeted therapies for pain relief in arthritis.

Heterogeneity among somatosensory neurons is necessary for internal and external sensation. Precise patterns of gene transcription orchestrated through enhancer activation maintain heterogeneity. Thus, high-resolution cell type classification, chromatin accessibility and its relation to enhancer activation can explain the governing principles for sensory neuron heterogeneity. Here, we present an integrated atlas from published high-quality scRNA-seq datasets and resequencing the dorsal root ganglion, including over 44,000 neurons. MERSCOPE spatial transcriptomics confirms cell types in situ, including previously unrecognized neuronal types, and a spatial zonation of both neurons and non-neuronal cells. We present a cell type specific open chromatin atlas revealing enhancer driven regulons and gene-regulatory networks organized into co-regulated gene-programs that together define sensory neuron diversity. Cell type complexity is shown to be generated by layered co-regulated transcriptional modules representing shared functions across different scales of the neuronal type hierarchy with cell type specific contribution as the exception.

Acute pain is an unpleasant experience caused by noxious stimuli. How the spinal neural circuits attribute differences in quality of noxious information remains unknown. By means of genetic capturing, activity manipulation and single-cell RNA sequencing, we identified distinct neural ensembles in the adult mouse spinal cord encoding mechanical and heat pain. Reactivation or silencing of these ensembles potentiated or stopped, respectively, paw shaking, lifting and licking within but not across the stimuli modalities. Within ensembles, polymodal Gal ⁺ inhibitory neurons with monosynaptic contacts to A-fiber sensory neurons gated pain transmission independent of modality. Peripheral nerve injury led to inferred microglia-driven inflammation and an ensemble transition with decreased recruitment of Gal ⁺ inhibitory neurons and increased excitatory drive. Forced activation of Gal ⁺ neurons reversed hypersensitivity associated with neuropathy. Our results reveal the existence of a spinal representation that forms the neural basis of the discriminative and defensive qualities of acute pain, and these neurons are under the control of a shared feed-forward inhibition.

While the inflammatory processes in rheumatoid arthritis have been described, mechanisms driving pain are poorly defined. Here, we used a multitude of approaches to uncover the neural basis, mediators, intracellular signaling pathway and the mechanism of inflammatory pain. In cartilage autoantibody-induced arthritis mice, an early immune-activation and a cytokine storm were mainly driven by vascular cells and monocyte/macrophages in the dorsal root ganglion. However, persistently elevated interferons and receptor-activation of the MNK1/2-eIF4E signaling pathway at all disease phases caused sensory-motor dysfunction and pain by inducing hyperexcitability and sensitization of Gfra3 + sensory neurons. Like mice, human sensory neurons expressed interferon receptors and interferons were elevated only in individuals with painful rheumatoid arthritis. Signaling pathway inhibition in vivo reversed pain and restored limb function. The finding that joint pain before and during arthritis is caused by a defined cytokine and signaling pathway holds promise for targeted therapies for pain relief in arthritis.

The versatility of somatosensation arises from heterogeneous dorsal root ganglion (DRG) neurons. However, soma transcriptomes of individual human (h)DRG neurons—critical information to decipher their functions—are lacking due to technical difficulties. In this study, we isolated somata from individual hDRG neurons and conducted deep RNA sequencing (RNA-seq) to detect, on average, over 9,000 unique genes per neuron, and we identified 16 neuronal types. These results were corroborated and validated by spatial transcriptomics and RNAscope in situ hybridization. Cross-species analyses revealed divergence among potential pain-sensing neurons and the likely existence of human-specific neuronal types. Molecular-profile-informed microneurography recordings revealed temperature-sensing properties across human sensory afferent types. In summary, by employing single-soma deep RNA-seq and spatial transcriptomics, we generated an hDRG neuron atlas, which provides insights into human somatosensory physiology and serves as a foundation for translational work.

Opioids are widely used, effective analgesics to manage severe acute and chronic pain, although they have recently come under scrutiny because of epidemic levels of abuse. While these compounds act on numerous central and peripheral pain pathways, the neuroanatomical substrate for opioid analgesia is not fully understood. By means of single-cell transcriptomics and manipulation of morphine-responsive neurons, we have identified an ensemble of neurons in the rostral ventromedial medulla (RVM) that regulates mechanical nociception in mice. Among these, forced activation or silencing of excitatory RVM ^BDNF projection neurons mimicked or completely reversed morphine-induced mechanical antinociception, respectively, via a brain-derived neurotrophic factor (BDNF)/tropomyosin receptor kinase B (TrkB)–dependent mechanism and activation of inhibitory spinal galanin-positive neurons. Our results reveal a specific RVM-spinal circuit that scales mechanical nociception whose function confers the antinociceptive properties of morphine.

Acute pain is an unpleasant experience caused by noxious stimuli. How the spinal neural circuits attribute differences in quality of noxious information remains unknown. By means of genetic capturing, activity manipulation and single cell RNA sequencing, we identified distinct neural ensembles in mouse spinal cord encoding mechanical and heat pain. Re-activation or silencing of these ensembles potentiated or stopped, respectively, affective but not reflex behaviour without altering pain behaviour to cross stimuli modality. Within ensembles, polymodal Gal ⁺ inhibitory neurons with monosynaptic contacts to A-fiber sensory neurons gated affective pain independent of modality. Peripheral nerve injury led to microglia driven inflammation and an ensemble transition with decreased recruitment of Gal ⁺ inhibitory neurons and increased excitatory drive. However, activating Gal ⁺ neurons reversed hypersensitivity associated with neuropathy. Our results reveal the existence of a spinal representation which forms the neural basis of the discriminative and affective qualities of acute pain and that these neurons are under the control of a shared feed-forward inhibition.