Protocol for dissection and culture of murine dorsal root ganglia neurons to study neuropeptide release

Perner, Caroline; Sokol, Caroline L.

doi:10.1016/j.xpro.2021.100333

Cited by 27 publications

(17 citation statements)

References 6 publications

(9 reference statements)

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“…DRG were dissected from naïve Swiss mice, digested, and seeded on Nunc™ Glass Bottom Dishes at a density of 1 × 10 6 per mL (1 × 10 5 /plate). Plates were incubated overnight in neurobasal medium, as previously described [ 38 , 52 ]. Neurons were treated either with vehicle (2% DMSO in culture media) or KA (50 and 150 μM) for 1 h at 37 °C and 5% CO 2 atmosphere.…”

Section: Methodsmentioning

confidence: 99%

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

et al. 2023

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Kaurenoic acid (KA) is a diterpene extracted from Sphagneticola trilobata (L.) Pruski. KA presents analgesic properties. However, the analgesic activity and mechanisms of action of KA in neuropathic pain have not been investigated so far; thus, we addressed these points in the present study. A mouse model of neuropathic pain was induced by chronic constriction injury (CCI) of the sciatic nerve. Acute (at the 7th-day post-CCI surgery) and prolonged (from 7–14th days post-CCI surgery) KA post-treatment inhibited CCI-induced mechanical hyperalgesia at all evaluated time points, as per the electronic version of von Frey filaments. The underlying mechanism of KA was dependent on activating the NO/cGMP/PKG/ATP-sensitive potassium channel signaling pathway since L-NAME, ODQ, KT5823, and glibenclamide abolished KA analgesia. KA reduced the activation of primary afferent sensory neurons, as observed by a reduction in CCI-triggered colocalization of pNF-κB and NeuN in DRG neurons. KA treatment also increased the expression of neuronal nitric oxide synthase (nNOS) at the protein level as well as the intracellular levels of NO in DRG neurons. Therefore, our results provide evidence that KA inhibits CCI neuropathic pain by activating a neuronal analgesic mechanism that depends on nNOS production of NO to silence the nociceptive signaling that generates analgesia.

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Section: Methodsmentioning

confidence: 99%

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

et al. 2023

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“…Animals were killed by CO 2 inhalation, followed by cervical dislocation and postmortem decapitation. Dissociated DRG cultures were prepared as previously described (Perner & Sokol, 2021 ). Briefly, DRGs were dissected from 1‐ to 2‐month‐old mice in Hank's buffered salt solution (HBSS) and digested with Collagenase A and Dispase II (Sigma‐Aldrich) for 25 min at 37°, centrifuged at 200× g for 5 min and washed with Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS), glutamate, sodium pyruvate, 1% penicillin and 1% streptomycin.…”

Section: Methodsmentioning

confidence: 99%

Exploring pharmacological inhibition of G_q/11 as an analgesic strategy

Marwari

Kowalski

Martemyanov

2022

British J Pharmacology

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Background and Purpose Misuse of opioids has greatly affected our society. One potential solution is to develop analgesics that act at targets other than opioid receptors. These can be used either as stand‐alone therapeutics or to improve the safety profile of opioid drugs. Previous research showed that activation of Gq/11 proteins by G‐protein coupled receptors has pro‐nociceptive properties, suggesting that blockade of Gq/11 signalling could be beneficial for pain control. The aim of this study was to test this hypothesis pharmacologically by using potent and selective Gq/11 inhibitor YM‐254890. Experimental Approach We used a series of behavioural assays to evaluate the acute responses of mice to painful thermal stimulation while administering YM‐254890 alone and in combination with morphine. We then used electrophysiological recordings to evaluate the effects of YM‐254890 on the excitability of dorsal root ganglion (DRG) nociceptor neurons. Key Results We found that systemic administration of YM‐254890 produced anti‐nociceptive effects and also augmented morphine analgesia in both hotplate and tail flick paradigms. However, it also caused substantial inhibition of locomotion, which may limit its therapeutic utility. To circumvent these issues, we explored the local administration of YM‐254890. Intrathecal injections of YM‐254890 produced lasting analgesia in a tail flick test and greatly augmented the anti‐nociceptive effects of morphine without any significant effects on locomotor behaviour. Electrophysiological studies showed that YM‐254890 reduced the excitability of DRG nociceptors and augmented their opioid‐induced inhibition. Conclusion and Implications These findings indicate that pharmacological inhibition of Gq/11 could be explored as an analgesic strategy.

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“…Primary DRG neurons were prepared from 8 to 10-week old mice as described previously [ 28 ]. Briefly, lumber DRG (L1 to L6) were isolated from mice and incubated with Dispase-II Solution (Sigma-Aldrich, St. Louis, MO, USA) and collagenase type II (Worthington, Columbus, OH, USA) for 70 min and with 0.25% trypsin for 5 min.…”

Section: Methodsmentioning

confidence: 99%

Oral Administration of Glutathione Trisulfide Increases Reactive Sulfur Levels in Dorsal Root Ganglion and Ameliorates Paclitaxel-Induced Peripheral Neuropathy in Mice

et al. 2022

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Peripheral neuropathy is a dose-limiting side effect of chemotherapy with paclitaxel. Paclitaxel-induced peripheral neuropathy (PIPN) is typically characterized by a predominantly sensory neuropathy presenting with allodynia, hyperalgesia and spontaneous pain. Oxidative mitochondrial damage in peripheral sensory neurons is implicated in the pathogenesis of PIPN. Reactive sulfur species, including persulfides (RSSH) and polysulfides (RSnH), are strong nucleophilic and electrophilic compounds that exert antioxidant effects and protect mitochondria. Here, we examined the potential neuroprotective effects of glutathione trisulfide (GSSSG) in a mouse model of PIPN. Intraperitoneal administration of paclitaxel at 4 mg/kg/day for 4 days induced mechanical allodynia and thermal hyperalgesia in mice. Oral administration of GSSSG at 50 mg/kg/day for 28 days ameliorated mechanical allodynia, but not thermal hyperalgesia. Two hours after oral administration, 34S-labeled GSSSG was detected in lumber dorsal root ganglia (DRG) and in the lumber spinal cord. In mice treated with paclitaxel, GSSSG upregulated expression of genes encoding antioxidant proteins in lumber DRG, prevented loss of unmyelinated axons and inhibited degeneration of mitochondria in the sciatic nerve. In cultured primary neurons from cortex and DRG, GSSSG mitigated paclitaxel-induced superoxide production, loss of axonal mitochondria, and axonal degeneration. These results indicate that oral administration of GSSSG mitigates PIPN by preventing axonal degeneration and mitochondria damage in peripheral sensory nerves. The findings suggest that administration of GSSSG may be an approach to the treatment or prevention of PIPN and other peripheral neuropathies.

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Protocol for dissection and culture of murine dorsal root ganglia neurons to study neuropeptide release

Cited by 27 publications

References 6 publications

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

Exploring pharmacological inhibition of G_q/11 as an analgesic strategy

Oral Administration of Glutathione Trisulfide Increases Reactive Sulfur Levels in Dorsal Root Ganglion and Ameliorates Paclitaxel-Induced Peripheral Neuropathy in Mice

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Protocol for dissection and culture of murine dorsal root ganglia neurons to study neuropeptide release

Cited by 27 publications

References 6 publications

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

Kaurenoic Acid Reduces Ongoing Chronic Constriction Injury-Induced Neuropathic Pain: Nitric Oxide Silencing of Dorsal Root Ganglia Neurons

Exploring pharmacological inhibition of Gq/11 as an analgesic strategy

Oral Administration of Glutathione Trisulfide Increases Reactive Sulfur Levels in Dorsal Root Ganglion and Ameliorates Paclitaxel-Induced Peripheral Neuropathy in Mice

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Exploring pharmacological inhibition of G_q/11 as an analgesic strategy