Animal Studies

Low back pain is a common cause of chronic pain and disability. It is modeled in rodents by chronically compressing the lumbar dorsal root ganglia (DRG) with small metal rods, resulting in ipsilateral mechanical and cold hypersensitivity, and hyperexcitability of sensory neurons. Sodium channels are implicated in this hyperexcitability, but the responsible isoforms are unknown. In this study, we used siRNA- mediated knockdown of the pore-forming Na1.6 and regulatory Naβ4 sodium channel isoforms that have been previously implicated in a different model of low back pain caused by locally inflaming the L5 DRG. Knockdown of either subunit markedly reduced spontaneous pain and mechanical and cold hypersensitivity induced by DRG compression, and reduced spontaneous activity and hyperexcitability of sensory neurons with action potentials <1.5 msec (predominately cells with myelinated axons, based on conduction velocities measured in a subset of cells) 4 days after DRG compression. These results were similar to those previously obtained in the DRG inflammation model and some neuropathic pain models, in which sensory neurons other than nociceptors seem to play key roles. The cytokine profiles induced by DRG compression and DRG inflammation were also very similar, with upregulation of several type 1 pro-inflammatory cytokines and downregulation of type 2 anti-inflammatory cytokines. Surprisingly, the cytokine profile was largely unaffected by Naβ4 knockdown in either model. The Na1.6 channel, and the Naβ4 subunit that can regulate Na1.6 to enhance repetitive firing, play key roles in both models of low back pain; targeting the abnormal spontaneous activity they generate may have therapeutic value.

Diabetic neuropathic pain (DNP), an early symptom ofdiabeticneuropathy, involves complex mechanisms. Long non-coding RNA (lncRNA) dysregulation contributes to the pathogenesis of various human diseases. Here, we investigated the genome-wide expression patterns of lncRNAs and genes in the spinal dorsal horn of mice with streptozotocin-induced DNP. Microarray analysis identified 1,481 differentially expressed (DE) lncRNAs and 1,096 DE mRNAs in DNP mice. Functional analysis showed that transforming growth factor-beta receptor binding was the most significant molecular function and retrograde endocannabinoid signaling was the most significant pathway of DE mRNAs. Calcium ion transport was the second most significant biological process of DE lncRNAs. Finally, we found 289 neighboring and 57 overlapping lncRNA-mRNA pairs, including ENSMUST00000150952-Mbp and AK081017-Usp15, which may be involved in DNP pathogenesis. Microarray data were validated through quantitative PCR of selected lncRNAs and mRNAs. These results suggest that aberrant expression of lncRNAs may contribute to the pathogenesis of DNP.

There is strong evidence that spinoparabrachial neurons in the superficial dorsal horn contribute to persistent pain states, and that the lateral parabrachial complex (PB) conveys relevant nociceptive information to higher structures. The role of PB itself in hyperalgesia and how it recruits descending facilitation has nevertheless received significantly less attention. The current study is a first step towards delineating the functional dynamics of PB and its link to descending control in acute and persistent inflammatory pain. In lightly anesthetized rats, we recorded behavioral withdrawal evoked by mechanical stimulation of the hindpaw, and simultaneously, activity of identified pain-modulating neurons, "ON-cells" and "OFF-cells," in the rostral ventromedial medulla (RVM). This was done before and after inactivation of PB, or to an inflamed paw (1 h, 1 d, or 5-6 d after intraplantar injection of Complete Freund's adjuvant, CFA). Inactivation of but not PB interfered with nociceptive input to RVM under basal conditions, as well as in acute inflammation. By contrast, blocking , but not , PB in established inflammation interfered with behavioral hyperalgesia and ON- and OFF-cell responses. Lesion of PB prior to CFA injection prevented this recruitment of PB in persistent inflammation. These experiments show that PB is required to initiate hyperalgesia, which is then maintained by PB, most likely in both cases via engagement of pain-modulating neurons of the RVM.The lateral parabrachial complex (PB) relays nociceptive information to brain circuits important for transmission and modulation of pain, but its specific role in persistent pain and engagement of descending control mechanisms has received relatively little attention. We show here that PB and to an inflammatory insult demonstrate different functions as inflammation persists, likely by engaging pain-facilitating neurons of the rostral ventromedial medulla. While the PB, the target of the major spinoparabrachial pathway, relays acute nociceptive information, the PB is recruited or unmasked in persistent inflammation to maintain hyperalgesia. These data point to plasticity in the PB itself or its direct and indirect connections with pain-modulating systems as central to development and maintenance of persistent pain.

Dysfunctions of genes transcription and translation in the nociceptive pathways play the critical role in development and maintenance of chronic pain. Circular RNAs (circRNAs) are emerging as new players in regulation of gene expression, but whether and how circRNAs are involved in chronic pain remains elusive. We showed here that complete Freund's adjuvant (CFA)-induced chronic inflammation pain significantly increased circRNA-Filip1l (filamin A interacting protein 1-like) expression in spinal neurons of mice. Blockage of this increase attenuated CFA-induced nociceptive behaviors, and overexpression of spinal circRNA-Filip1l in naïve mice mimicked the nociceptive behaviors as evidenced by decreased thermal and mechanical nociceptive threshold. Furthermore, we found that mature circRNA-Filip1l expression was negatively regulated by miRNA-1224 via binding and splicing of precursor of circRNA-Filip1l (pre-circRNA-Filip1l) in the Argonaute-2 (Ago2)-dependent manner. Increase of spinal circRNA-Filip1l expression resulted from the decrease of miRNA-1224 expression under chronic inflammation pain state. MiRNA-1224 knockdown or Ago2 overexpression induced nociceptive behaviors in naïve mice, which was prevented by the knockdown of spinal circRNA-Filip1l. Finally, we demonstrated that an ubiquitin protein ligase E3 component n-recognin 5 (Ubr5), validated as a target of circRNA-Filip1l, plays a pivotal role in regulation of nociception by spinal circRNA-Filip1l. These data suggest that miRNA-1224-mediated and Ago2-dependent modulation of spinal circRNA-Filip1l expression regulates nociception via targeting Ubr5, revealing a novel epigenetic mechanism of interaction between miRNA and circRNA in chronic inflammation pain.CircRNAs are emerging as new players in regulation of gene expression. Here, we found that the increase of circRNA-Filip1l mediated by miRNA-1224 in Ago2-dependent way in the spinal cord is involved in regulation of nociception via targeting Ubr5. Our study reveals a novel epigenetic mechanism of interaction between miRNA and circRNA in chronic inflammation pain.

Sensory neurons perceive environmental cues and are important of organismal survival. Peripheral sensory neurons interact intimately with glial cells. While the function of axonal ensheathment by glia is well studied, less is known about the functional significance of glial interaction with the somatodendritic compartment of neurons. Herein, we show that three distinct glia cell types differentially wrap around the axonal and somatodendritic surface of the polymodal dendritic arborization (da) neuron of the peripheral nervous system for detection of thermal, mechanical, and light stimuli. We find that glial cell-specific loss of the chromatin modifier gene dATRX in the subperineurial glial layer leads to selective elimination of somatodendritic glial ensheathment, thus allowing us to investigate the function of such ensheathment. We find that somatodendritic glial ensheathment regulates the morphology of the dendritic arbor, as well as the activity of the sensory neuron, in response to sensory stimuli. Additionally, glial ensheathment of the neuronal soma influences dendritic regeneration after injury.