Abstract:Nerve injury-induced downregulation of voltage-gated potassium channel subunit Kcna2 in the dorsal root ganglion (DRG) is critical for DRG neuronal excitability and neuropathic pain genesis. However, how nerve injury causes this downregulation is still elusive. Euchromatic histone-lysine N-methyltransferase 2, also known as G9a, methylates histone H3 on lysine residue 9 to predominantly produce a dynamic histone dimethylation, resulting in condensed chromatin and gene transcriptional repression. We showed here… Show more
“…No cell bodies of spinal cord neurons were labeled. Consistent with previous reports, 13–16,18 the injected DRG, stained with hematoxylin and eosin, retained its structural integrity and contained no visible leukocytes (data not shown). Figure 5.EGFP-labeled AAV-DJ is limited in L4 DRG and its fibers and terminals on the ipsilateral side after AAV-EGFP injection into the unilateral L4 DRG.…”
Section: Resultssupporting
confidence: 92%
“…AAV-DJ expressing EGFP (AAV-EGFP) was used as a control. Like AAV5 reported previously, 13–16,18 AAV-DJ requires three to four weeks to become expressed and its expression lasts for at least six weeks (Figure 5). EGFP-labeled AAV-DJ was limited to the ipsilateral L 4 DRG neurons and their fibers and terminals four weeks after microinjection (Figure 5(e), (g), and (h)).…”
Section: Resultssupporting
confidence: 52%
“…15,16,19 Briefly, animals were anesthetized with 2–3% isoflurane. The left fifth lumbar transverse process was identified and then was separated from its muscle attachments.…”
Section: Methodsmentioning
confidence: 99%
“…13–16,18 Briefly, mice were sacrificed under isoflurane anesthesia, and bilateral L 4 DRGs was rapidly collected. To obtain enough RNA, L 4 DRGs from four mice per time point were pooled.…”
Section: Methodsmentioning
confidence: 99%
“…All data were normalized to Tuba1a mRNA, which has been demonstrated to be stable even after peripheral nerve injury insult in mice. 14–16 …”
Peripheral nerve injury-caused hyperexcitability and abnormal ectopic discharges in the primary sensory neurons of dorsal root ganglion (DRG) play a key role in neuropathic pain development and maintenance. The two-pore domain background potassium (K2P) channels have been identified as key determinants of the resting membrane potential and neuronal excitability. However, whether K2P channels contribute to neuropathic pain is still elusive. We reported here that K2P1.1, the first identified mammalian K2P channel, was highly expressed in mouse DRG and distributed in small-, medium-, and large-sized DRG neurons. Unilateral lumbar (L) 4 spinal nerve ligation led to a significant and time-dependent reduction of K2P1.1 mRNA and protein in the ipsilateral L4 DRG, but not in the contralateral L4 or ipsilateral L3 DRG. Rescuing this reduction through microinjection of adeno-associated virus-DJ expressing full-length K2P1.1 mRNA into the ipsilateral L4 DRG blocked spinal nerve ligation-induced mechanical, thermal, and cold pain hypersensitivities during the development and maintenance periods. This DRG viral microinjection did not affect acute pain and locomotor function. Our findings suggest that K2P1.1 participates in neuropathic pain development and maintenance and may be a potential target in the management of this disorder.
“…No cell bodies of spinal cord neurons were labeled. Consistent with previous reports, 13–16,18 the injected DRG, stained with hematoxylin and eosin, retained its structural integrity and contained no visible leukocytes (data not shown). Figure 5.EGFP-labeled AAV-DJ is limited in L4 DRG and its fibers and terminals on the ipsilateral side after AAV-EGFP injection into the unilateral L4 DRG.…”
Section: Resultssupporting
confidence: 92%
“…AAV-DJ expressing EGFP (AAV-EGFP) was used as a control. Like AAV5 reported previously, 13–16,18 AAV-DJ requires three to four weeks to become expressed and its expression lasts for at least six weeks (Figure 5). EGFP-labeled AAV-DJ was limited to the ipsilateral L 4 DRG neurons and their fibers and terminals four weeks after microinjection (Figure 5(e), (g), and (h)).…”
Section: Resultssupporting
confidence: 52%
“…15,16,19 Briefly, animals were anesthetized with 2–3% isoflurane. The left fifth lumbar transverse process was identified and then was separated from its muscle attachments.…”
Section: Methodsmentioning
confidence: 99%
“…13–16,18 Briefly, mice were sacrificed under isoflurane anesthesia, and bilateral L 4 DRGs was rapidly collected. To obtain enough RNA, L 4 DRGs from four mice per time point were pooled.…”
Section: Methodsmentioning
confidence: 99%
“…All data were normalized to Tuba1a mRNA, which has been demonstrated to be stable even after peripheral nerve injury insult in mice. 14–16 …”
Peripheral nerve injury-caused hyperexcitability and abnormal ectopic discharges in the primary sensory neurons of dorsal root ganglion (DRG) play a key role in neuropathic pain development and maintenance. The two-pore domain background potassium (K2P) channels have been identified as key determinants of the resting membrane potential and neuronal excitability. However, whether K2P channels contribute to neuropathic pain is still elusive. We reported here that K2P1.1, the first identified mammalian K2P channel, was highly expressed in mouse DRG and distributed in small-, medium-, and large-sized DRG neurons. Unilateral lumbar (L) 4 spinal nerve ligation led to a significant and time-dependent reduction of K2P1.1 mRNA and protein in the ipsilateral L4 DRG, but not in the contralateral L4 or ipsilateral L3 DRG. Rescuing this reduction through microinjection of adeno-associated virus-DJ expressing full-length K2P1.1 mRNA into the ipsilateral L4 DRG blocked spinal nerve ligation-induced mechanical, thermal, and cold pain hypersensitivities during the development and maintenance periods. This DRG viral microinjection did not affect acute pain and locomotor function. Our findings suggest that K2P1.1 participates in neuropathic pain development and maintenance and may be a potential target in the management of this disorder.
Nerve injury‐induced change in gene expression in primary sensory neurons of dorsal root ganglion (DRG) is critical for neuropathic pain genesis. N
6
‐methyladenosine (m
6
A) modification of RNA represents an additional layer of gene regulation. Here, it is reported that peripheral nerve injury increases the expression of the m
6
A demethylase fat‐mass and obesity‐associated proteins (FTO) in the injured DRG via the activation of Runx1, a transcription factor that binds to the
Fto
gene promoter. Mimicking this increase erases m
6
A in euchromatic histone lysine methyltransferase 2 (
Ehmt2
) mRNA (encoding the histone methyltransferase G9a) and elevates the level of G9a in DRG and leads to neuropathic pain symptoms. Conversely, blocking this increase reverses a loss of m
6
A sites in
Ehmt2
mRNA and destabilizes the nerve injury‐induced G9a upregulation in the injured DRG and alleviates nerve injury‐associated pain hypersensitivities. FTO contributes to neuropathic pain likely through stabilizing nerve injury‐induced upregulation of G9a, a neuropathic pain initiator, in primary sensory neurons.
Nerve injury-induced maladaptive changes of gene expression in dorsal root ganglion (DRG) neurons contribute to neuropathic pain. Long non-coding RNAs (lncRNAs) are emerging as key regulators of gene expression. Here, a conserved lncRNA is reported, named DRG-specifically enriched lncRNA (DS-lncRNA) for its high expression in DRG neurons. Peripheral nerve injury downregulates DS-lncRNA in injured DRG due, in part, to silencing of POU domain, class 4, transcription factor 3, a transcription factor that interacts with the DS-lncRNA gene promoter. Rescuing DS-lncRNA downregulation blocks nerve injury-induced increases in the transcriptional cofactor RALY-triggered DRG Ehmt2 mRNA and its encoding G9a protein, reverses the G9a-controlled downregulation of opioid receptors and Kcna2 in injured DRG, and attenuates nerve injury-induced pain hypersensitivities in male mice. Conversely, DS-lncRNA downregulation increases RALY-triggered Ehmt2/G9a expression and correspondingly decreases opioid receptor and Kcna2 expression in DRG, leading to neuropathic pain symptoms in male mice in the absence of nerve injury. Mechanistically, downregulated DS-lncRNA promotes more binding of increased RALY to RNA polymerase II and the Ehmt2 gene promoter and enhances Ehmt2 transcription in injured DRG. Thus, downregulation of DS-lncRNA likely contributes to neuropathic pain by negatively regulating the expression of RALY-triggered Ehmt2/G9a, a key neuropathic pain player, in DRG neurons.
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