Targeting Microglia Activation to Treat Neuropathic Pain Following Spinal Cord Injury
IP injection of rapamycin reduces neuropathic pain in mice following spinal cord injury.
Among the many cellular functions the mammalian target of rapamycin (mTOR) pathway is involved in, are neuroprotection in neurodegenerative disorders1,2 and neuroregeneration following traumatic brain injury (TBI) and spinal cord injury (SCI).3,4 Rapamycin inhibits the mTOR pathway by preventing phosphorylation of its targets, p70S6K and 4EBP1.5
Neuroprotective properties of rapamycin are conferred by its promotion of autophagy;1,2 rapamycin was also implicated in suppression of microglial activation and reduction of inflammation in the CNS.6 In addition, rapamycin administration was shown to lessen neural tissue damage and locomotor impairment following SCI in mice.7
Activation of glial cells and of p38 MAPK have been shown to be involved in development of chronic neuropathic pain (with burning, stabbing and electric shock-like symptoms) which affects up to 80% of SCI patients.8-10 Neuroprotective therapy in the (sub)acute phase following SCI, with anti-CD11d monoclonal antibody IV injection11, fibronectin injection into the spinal dorsal column12, or intrathecal injection of carbenoxolone13 led to reduces levels of neuropathic pain in animal studies.
In the present study, authors sought to investigate effects of rapamycin in SCI-related neuropathic pain using a spinal cord contusion injury model in mice.14 In these animals, rapamycin treatment led to reduced levels of phosphorylated p70S6K at the lesion site, compared to vehicle- and sham-treated controls, indicating inhibition of the mTOR pathway (p <.05). Locomotor recovery (including coordination, trunk stability, joint movements and stepping ability) was assessed with locomotor tests and measured using the Basso Mouse Scale (BMS) score in a blinded manner. Sham controls showed no locomotor impairment; mice treated with rapamycin had higher BMS scores from 7 to 42 days following SCI than vehicle-treated animals (p <.05).
Mechanical hypersensitivity was assessed using the von Frey method in which cutaneous sensitivity to an innocuous mechanical stimulation of the plantar hindpaws (poking with filaments) is evaluated by paw withdrawal threshold. Sham controls showed no change in withdrawal threshold, whereas rapamycin-treated animals had higher thresholds than vehicle-treated mice from 14-42 days following SCI, indicating an improvement in mechanical hypersensitivity (p <.05). Similarly, thermal sensitivity was improved by the rapamycin treatment (p <.05).
Immunohistochemical studies showed microglia activation in the superficial dorsal horn of vehicle-, but not rapamycin-treated or sham mice, as indicated by an increase in Iba-1 stained microglia 42 days after SCI. These microglia were hypertrophic in vehicle- but not treated mice, as indicated by thickening of processes and larger cell bodies. Activation of glial cells is thought to result in neuropathic pain through release of pro-inflammatory cytokines, neurotransmitters and reactive oxygen species.15 Vehicle-treated animals also had increased number and density of GFAP-stained cells (a marker of astrocytes) in the superficial dorsal horn, as well as increased phosphorylation of p38 MAPK, indicating glial cell activation, an essential factor in the development of neuropathic pain (p <.01).
Vehicle-treated mice showed increases in double immunostaining of phosphorylated p38 MAPK and Iba-1 in the lumbar spinal cord 42 days after injury, indicating rapamycin treatment was effective in reducing p38 MAPK activation and secondary neural tissue damage.
Overall, results from this study show that rapamycin is effective in inhibiting the mTOR pathway at the site of injury, in improving locomotor function, and in diminishing mechanical and thermal hypersensitivity. The treatment also shows significant improvement in suppressing microglial activation in the acute phase following injury and in attenuating development of SCI-related neuropathic pain in the chronic phase.
1.Malagelada C, Jin ZH, Jackson-lewis V, Przedborski S, Greene LA. Rapamycin protects against neuron death in in vitro and in vivo models of Parkinson's disease. J Neurosci. 2010;30(3):1166-75.
2.Ravikumar B, Vacher C, Berger Z, et al. Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease. Nat Genet. 2004;36(6):585-95.
3.Erlich S, Alexandrovich A, Shohami E, Pinkas-kramarski R. Rapamycin is a neuroprotective treatment for traumatic brain injury. Neurobiol Dis. 2007;26(1):86-93.
4.Kanno H, Ozawa H, Sekiguchi A, et al. The role of mTOR signaling pathway in spinal cord injury. Cell Cycle. 2012;11(17):3175-9.
5.Schmelzle T, Hall MN. TOR, a central controller of cell growth. Cell. 2000;103(2):253-62.
6.Dello russo C, Lisi L, Tringali G, Navarra P. Involvement of mTOR kinase in cytokine-dependent microglial activation and cell proliferation. Biochem Pharmacol. 2009;78(9):1242-51.
7.Tang P, Hou H, Zhang L, et al. Autophagy reduces neuronal damage and promotes locomotor recovery via inhibition of apoptosis after spinal cord injury in rats. Mol Neurobiol. 2014;49(1):276-87.
8.Detloff MR, Fisher LC, Mcgaughy V, Longbrake EE, Popovich PG, Basso DM. Remote activation of microglia and pro-inflammatory cytokines predict the onset and severity of below-level neuropathic pain after spinal cord injury in rats. Exp Neurol. 2008;212(2):337-47.
9.Crown ED, Gwak YS, Ye Z, Johnson KM, Hulsebosch CE. Activation of p38 MAP kinase is involved in central neuropathic pain following spinal cord injury. Exp Neurol. 2008;213(2):257-67.
10.Finnerup NB, Johannesen IL, Sindrup SH, Bach FW, Jensen TS. Pain and dysesthesia in patients with spinal cord injury: A postal survey. Spinal Cord. 2001;39(5):256-62.
11.Gris D, Marsh DR, Oatway MA, et al. Transient blockade of the CD11d/CD18 integrin reduces secondary damage after spinal cord injury, improving sensory, autonomic, and motor function. J Neurosci. 2004;24(16):4043-51.
12.Lin CY, Lee YS, Lin VW, Silver J. Fibronectin inhibits chronic pain development after spinal cord injury. J Neurotrauma. 2012;29(3):589-99.
13.Roh DH, Yoon SY, Seo HS, et al. Intrathecal injection of carbenoxolone, a gap junction decoupler, attenuates the induction of below-level neuropathic pain after spinal cord injury in rats. Exp Neurol. 2010;224(1):123-32.
14.Tateda S, Kanno H, Ozawa H, et al. Rapamycin suppresses microglial activation and reduces the development of neuropathic pain after spinal cord injury. J Orthop Res. 2016.
15.Martin DL. Synthesis and release of neuroactive substances by glial cells. Glia. 1992;5(2):81-94.