Involvement Of Spinal Glia In Inhibition Of Electroacupuncture On Hypersensitivity In Monoarthritic Rats

Arthritis, as an important medical problem, has been the subject of intense research. Pain is its predominant clinical feature, yet therapy is ineffective for many patients. Substantial evidence has indicated that electro-acupuncture (EA) is effective pain management that has been used in clinic.Complete Freund's adjuvant (CFA) has been utilized to induce rat arthritic immunopathological disease that displays many of the pathological features of human rheumatoid arthritis. Inflammation of the joint causes peripheral and central sensitization. The processes of sensitization are thought to be the basis of arthritic pain that appears as spontaneous pain (joints at rest), hyperalgesia (exaggerated pain response on noxious stimulation), and allodynia (pain on normally non-painful stimulation). It is well known now that spinal cord glia (microglia and astrocytes) importantly contribute to the development and maintenance of central sensitization. Robust glial activation has been observed on the lumbar spinal cord in various rodent models of chronic pain, including spinal nerve injury and peripheral nerve inflammation. The degree and time course of glial activation are correlated with pain facilitation. Activated spinal glia release a variety of algesic substances that enhance pain transmission, such as pro-inflammatory cytokines. Blocking the activation of spinal cord glia can block the development and maintenance of allodynia and hyperalgesia, suggesting that spinal glial activation is necessary for induction of exaggerated pain.Electroacupuncture (EA), an acupuncture therapy, has been used for several decades in the treatment of acute and chronic pain. In experimental animals,arthritis-induced hyperalgesia and allodynia successfully suppressed by repeated EA also. Several processes have been proposed to explain effects of EA, primarily those on pain. Recent published data have shown that healthy and pathological conditions respond differently to EA. Under conditions of inflammatory pain, EA induced strong and prolonged anti-hyperalgesia, suggesting the different mechanisms in EA analgesia between the physiological and pathological pain states. Whether the spinal glial activation is involved in EA analgesia in inflammatory pain model is unknown. Thus, the purpose of the present study is to examine whether the EA antagonizes activation of spinal glia, and whether disruption of spinal glial activation enhances antagonism of EA on CFA-induced mechanical allodynia and thermal hyperalgesia. The results are as follows:1, Time course of glial activation on the spinal cord in CFA-induced monoarthritis (MA) ratsFollowing unilateral intra-articular injection of CFA, mechanical allodynia, thermal hyperalgesia and joint inflammation developed at 4 hours and persisted for 10 days in the ipsilateral hindpaw. Microglia were markedly activated on both sides with the ipsilateral one more preferential from day 1, and maximal at day 3, while slightly decreased at day 10 post-CFA, characterized by immunostaining of OX42. Unlike microglial activation, a significant astrocytic activation occurred from day 3, and reached maximum by 10 days post-MA, characterized by potentiated immunostaining of glial fibrillary acidic protein (GFAP). And western blotting analysis confirmed above immunohistochemistry finding. This result suggests the involvement of spinal glia in MA-induced pain facilitation.2, Blockade of spinal glial activation inhibited MA-induced behavioral hypersensitivityIntrathecal injection of 1 nmol but not 0.1nmol FC (non-selective glial inhibitor) suppressed MA-induced edema, mechanical allodynia and thermal hyperalgesia, as well as spinal microglial and astrocytic activations.Intrathecal injection of 100 μg but not 20μg minocycline before MA inhibited spinal glial activation and delayed the development of mechanical allodynia and thermal hyperalgesia but not reversed established pain facilitation on day 3 post-CFA injection. These data indicate that microglia may be involved in the initiation of pain facilitation, whereas astrocytes may play an important role in the maintenance of exaggerated pain.3, CX3CR1 mediated the onset of spinal glial activation and behavioral hypersensitivityFollowing unilateral intra-articular injection of CFA, CX3CR1 (fractalkine receptor, predominantly express on spinal microglia) level significantly increased in the spinal cord, with a similar time course to microglia. Intrathecal injection of anti-CX3CR1 neutralizing antibody both delayed the development of behavioral hypersensitivity, and reversed established mechanical allodynia and thermal hyperalgesia. . These data provided new evidence for the contribution of endogenous fractalkine to the initiation and maintenance of inflammatory pain facilitation via activating spinal microglia.4, Dual regulation of EA on activation of spinal gliaEA delivered to the ipsilateral hindpaw at the "Huantiao"(GB30) and "Yanglingquan" (GB34) acupoints (100/2 Hz alternation, 1-2-3 mA, 30 min) was repeatedly given on day 1, 3, 5, and 7 post-MA or sham MA, and spinal astrocytic activition was examined. In NS-induced sham MA rats, EA produced moderate increase in GFAP expression on the spinal cord, compared to sham EA or normal rats. However, in CFA-induced MA rats, EA markedly suppressed MA-induced up-regulation of GFAP on the spinal dorsal horn, suggesting that EA can dually regulate spinal glial function.5, Disruption of glial function enhanced EA analgesia in MA ratsCFA-induced joint edema, mechanical allodynia and thermal hyperalgesia, as well as spinal glial activation were significantly suppressed by EA. Co-applicationof EA and low dose FC (0.1 nmol) or minocycline (20 g) completely reversed CFA-induced mechanical allodynia and thermal hyperalgesia. Although 0.1 nmol FC or 20μg minocycline alone did not alter the paw withdrawal threshold (PWT) to von Frey hairs and paw withdrawal latency (PWL) to thermal stimulus of the hindpaw ipsilateral to MA, EA-induced anti-allodynia and anti-hyperalgesia were significantly enhanced when they combined with EA. These data indicated that disruption of glial function significantly potentiated EA analgesia in MA rats.In conclusion, the present study demonstrated that1. Spinal microglia activation may contribute to the initiation of MA-induced pain facilitation, while astrocytes may play an important role in the maintenance of exaggerated pain.2. CX3CR1, an exclusive fractalkine receptor, which was dominantly expressed by spinal microglia, may take an essential part in spinal glial activation and inflammatory pain facilitation under conditions of inflammatory pain.3. EA regulated spinal glial activation dually that it increased glial activation in normal condition and acts opposed pathological states.4. Disruption of glial function may potentiate inhibition of EA on MA-induced thermal hyperalgia, as well as spinal glial activation.