ANO1 (TMEM16A) is a Ca2+-activated Cl− channel (CaCC) expressed in peripheral somatosensory neurons that are activated by painful (noxious) stimuli. These neurons also express the Ca2+-permeable channel and noxious heat sensor TRPV1, which can activate ANO1. Here, we revealed an intricate mechanism of TRPV1-ANO1 channel coupling in rat dorsal root ganglion (DRG) neurons. Simultaneous optical monitoring of CaCC activity and Ca2+ dynamics revealed that the TRPV1 ligand capsaicin activated CaCCs. However, depletion of endoplasmic reticulum (ER) Ca2+ stores reduced capsaicin-induced Ca2+ increases and CaCC activation, suggesting that ER Ca2+ release contributed to TRPV1-induced CaCC activation. ER store depletion by plasma membrane–localized TRPV1 channels was demonstrated with an ER-localized Ca2+ sensor in neurons exposed to a cell-impermeable TRPV1 ligand. Proximity ligation assays established that ANO1, TRPV1, and the IP3 receptor IP3R1 were often found in close proximity to each other. Stochastic optical reconstruction microscopy (STORM) confirmed the close association between all three channels in DRG neurons. Together, our data reveal the existence of ANO1-containing multichannel nanodomains in DRG neurons and suggest that coupling between TRPV1 and ANO1 requires ER Ca2+ release, which may be necessary to enhance ANO1 activation.

Accumulating observations suggest that peripheral somatosensory ganglia may regulate nociceptive transmission, yet direct evidence is sparse. Here, in experiments on rats and mice, we show that the peripheral afferent nociceptive information in mice undergoes dynamic filtering within the dorsal root ganglion (DRG) and suggest that this filtering occurs at the axonal bifurcations (t-junctions). Using synchronous in vivo electrophysiological recordings from the peripheral and central processes of sensory neurons (in the spinal nerve and dorsal root), ganglionic transplantation of GABAergic progenitor cells, and optogenetics, we demonstrate existence of tonic and dynamic filtering of action potentials traveling through the DRG. Filtering induced by focal application of GABA or optogenetic GABA release from the DRG-transplanted GABAergic progenitor cells was specific to nociceptive fibers. Light-sheet imaging and computer modeling demonstrated that, compared to other somatosensory fiber types, nociceptors have shorter stem axons, making somatic control over t-junctional filtering more efficient. Optogenetically induced GABA release within DRG from the transplanted GABAergic cells enhanced filtering and alleviated hypersensitivity to noxious stimulation produced by chronic inflammation and neuropathic injury in vivo. These findings support “gating” of pain information by DRGs and suggest new therapeutic approaches for pain relief.

Graphical abstract

The actions of animals provide a window into how their minds work. Recent advances in deep learning are providing powerful approaches to recognize patterns of animal movement from video recordings, including markerless pose estimation models. However, tools to efficiently parse coordinates of animal position and pose into meaningful semantic behavioral labels are lacking. Here, we present PoseRecognition (PoseR), a behavioral decoder leveraging state-of-the-art action recognition models using spatio-temporal graph convolutional networks. We show that it can be used to decode animal behavior quickly and accurately from pose estimations, using zebrafish larvae and mice as model organisms. PoseR can be accessed using a Napari plugin, which facilitates efficient behavioral extraction, annotation, model training and deployment. We have simplified the workflow of behavioral analysis after pose estimation, transforming coordinates of animal position and pose into meaningful semantic behavioral labels, using methods designed for fast and accurate behavioral extraction, annotation, model training and deployment. Furthermore, we contribute a novel method for unsupervised clustering of behaviors and provide open-source access to our zebrafish datasets and models. The design of our tool ensures scalability and versatility for use across multiple species and contexts, improving the efficiency of behavioral analysis across fields.

Kv3 channels contribute to the excitability of sub-populations of spinal cord neurons in lamina VII.