Inhibiting endocytosis in CGRP+ nociceptors attenuates inflammatory pain-like behavior

Powell, Rasheen; Young, Violet A.; Pryce, Kerri D.; Sheehan, Garrett D.; Bonsu, Kwaku; Ahmed, Abdulelah; Bhattacharjee, Abhishek

doi:10.1038/s41467-021-26100-6

Cited by 33 publications

(10 citation statements)

References 72 publications

(98 reference statements)

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“…There are some points that I would like to comment on: It would be nice if the authors discuss and compare these results with previous reports. For example, CGRP was known to play an important role in noxious heat sensitivity or inflammatory pain 1 , 2 . TrkB-positive neurons were also ablated to investigate its role in mechanical pain 3 .…”

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confidence: 99%

Tools for analysis and conditional deletion of subsets of sensory neurons

et al. 2021

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Background: Somatosensation depends on primary sensory neurons of the trigeminal and dorsal root ganglia (DRG). Transcriptional profiling of mouse DRG sensory neurons has defined at least 18 distinct neuronal cell types. Using an advillin promoter, we have generated a transgenic mouse line that only expresses diphtheria toxin A (DTA) in sensory neurons in the presence of Cre recombinase. This has allowed us to ablate specific neuronal subsets within the DRG using a range of established and novel Cre lines that encompass all sets of sensory neurons. Methods: A floxed-tdTomato-stop-DTA bacterial artificial chromosome (BAC) transgenic reporter line (AdvDTA) under the control of the mouse advillin DRG promoter was generated. The line was first validated using a Nav1.8Cre and then crossed to CGRPCreER (Calca), ThCreERT2, Tmem45bCre, Tmem233Cre, Ntng1Cre and TrkBCreER (Ntrk2) lines. Pain behavioural assays included Hargreaves’, hot plate, Randall-Selitto, cold plantar, partial sciatic nerve ligation and formalin tests. Results: Motor activity, as assessed by the rotarod test, was normal for all lines tested. Noxious mechanosensation was significantly reduced when either Nav1.8 positive neurons or Tmem45b positive neurons were ablated whilst acute heat pain was unaffected. In contrast, noxious mechanosensation was normal following ablation of CGRP-positive neurons but acute heat pain thresholds were significantly elevated and a reduction in nocifensive responses was observed in the second phase of the formalin test. Ablation of TrkB-positive neurons led to significant deficits in mechanical hypersensitivity in the partial sciatic nerve ligation neuropathic pain model. Conclusions: Ablation of specific DRG neuronal subsets using the AdvDTA line will be a useful resource for further functional characterization of somatosensory processing, neuro-immune interactions and chronic pain disorders.

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confidence: 99%

Tools for analysis and conditional deletion of subsets of sensory neurons

et al. 2021

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“…2 CGRP+ve nociceptors express AP2α2, a key endocytosis protein that contributes to nociception. 28 Satellite glial cells of sensory ganglia were identified by immunostaining for GS. Dnm1 and Dnm3 mRNAs were expressed in neurons of DRG and TG (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The a subunit of the AP2 complex (AP2a2) is expressed in CGRP1ve nociceptors and AP2 small hairpin RNA (shRNA) suppressed nociception in mice. 28 However, the distribution of Dnm and AAK1 isoforms in nociceptive circuits is unknown, and their role in nociception is unexplored. We used genetic and pharmacological approaches to investigate the contribution of endocytosis to nociception.…”

Section: Introductionmentioning

confidence: 99%

The contribution of endocytosis to sensitization of nociceptors and synaptic transmission in nociceptive circuits

Tonello

Anderson

Davidson

et al. 2022

Pain

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Chronic pain involves sensitization of nociceptors and synaptic transmission of painful signals in nociceptive circuits in the dorsal horn of the spinal cord. We investigated the contribution of clathrin-dependent endocytosis to sensitization of nociceptors by G protein-coupled receptors (GPCRs) and to synaptic transmission in spinal nociceptive circuits. We determined whether therapeutic targeting of endocytosis could ameliorate pain. mRNA encoding dynamin (Dnm) 1 to 3 and adaptor-associated protein kinase 1 (AAK1), which mediate clathrin-dependent endocytosis, were localized to primary sensory neurons of dorsal root ganglia of mouse and human and to spinal neurons in the dorsal horn of the mouse spinal cord by RNAScope. When injected intrathecally to mice, Dnm and AAK1 siRNA or shRNA knocked down Dnm and AAK1 mRNA in dorsal root ganglia neurons, reversed mechanical and thermal allodynia and hyperalgesia, and normalized nonevoked behavior in preclinical models of inflammatory and neuropathic pain. Intrathecally administered inhibitors of clathrin, Dnm, and AAK1 also reversed allodynia and hyperalgesia. Disruption of clathrin, Dnm, and AAK1 did not affect normal motor functions of behaviors. Patch clamp recordings of dorsal horn neurons revealed that Dnm1 and AAK1 disruption inhibited synaptic transmission between primary sensory neurons and neurons in lamina I/II of the spinal cord dorsal horn by suppressing release of synaptic vesicles from presynaptic primary afferent neurons. Patch clamp recordings from dorsal root ganglion nociceptors indicated that Dnm siRNA prevented sustained GPCR-mediated sensitization of nociceptors. By disrupting synaptic transmission in the spinal cord and blunting sensitization of nociceptors, endocytosis inhibitors offer a therapeutic approach for pain treatment.

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“…While endocytosis is a critical mechanism in maintaining cell and thus host well-being, there is growing evidence that transient modulation is not only feasible but may have positive therapeutic outcomes. Chew et al ( 2020 ) report on blocking CME as an effective strategy to improve the clinical response to antibody dependent cellular cytotoxicity mediating antibodies in humans; Tremblay et al ( 2020 ) report that the dynamin inhibitor, dynole 34-4 is highly synergistic with existing chemotherapy in mouse models of T-ALL and AML; Powell et al ( 2021 ) report that endocytosis inhibition attenuates inflammatory pain like behaviour; and Jensen et al ( 2017 ) also reported on an endocytosis mediated effect as a viable therapeutic target for prolonged pain relief. While these findings should be considered positive, it is prudent to temper the enthusiasm that may be apparent with the realisation that the path to the clinic is long and winding.…”

Section: Discussionmentioning

confidence: 99%

Role of Clathrin and Dynamin in Clathrin Mediated Endocytosis/Synaptic Vesicle Recycling and Implications in Neurological Diseases

Prichard

O'Brien²,

Murcia³

et al. 2022

Front. Cell. Neurosci.

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Endocytosis is a process essential to the health and well-being of cell. It is required for the internalisation and sorting of “cargo”—the macromolecules, proteins, receptors and lipids of cell signalling. Clathrin mediated endocytosis (CME) is one of the key processes required for cellular well-being and signalling pathway activation. CME is key role to the recycling of synaptic vesicles [synaptic vesicle recycling (SVR)] in the brain, it is pivotal to signalling across synapses enabling intracellular communication in the sensory and nervous systems. In this review we provide an overview of the general process of CME with a particular focus on two key proteins: clathrin and dynamin that have a central role to play in ensuing successful completion of CME. We examine these two proteins as they are the two endocytotic proteins for which small molecule inhibitors, often of known mechanism of action, have been identified. Inhibition of CME offers the potential to develop therapeutic interventions into conditions involving defects in CME. This review will discuss the roles and the current scope of inhibitors of clathrin and dynamin, providing an insight into how further developments could affect neurological disease treatments.

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Inhibiting endocytosis in CGRP+ nociceptors attenuates inflammatory pain-like behavior

Cited by 33 publications

References 72 publications

Tools for analysis and conditional deletion of subsets of sensory neurons

Tools for analysis and conditional deletion of subsets of sensory neurons

The contribution of endocytosis to sensitization of nociceptors and synaptic transmission in nociceptive circuits

Role of Clathrin and Dynamin in Clathrin Mediated Endocytosis/Synaptic Vesicle Recycling and Implications in Neurological Diseases

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