Aihui Li

IL-1ra alleviates inflammatory hyperalgesia through preventing phosphorylation of NMDA receptor NR-1 subunit in rats

&NA; Although it has been shown that pro‐inflammatory cytokines such as interleukin‐1β (IL‐1β) facilitate perception of noxious inputs at the spinal level, the mechanisms have not been understood. This study determined the cell type that produces IL‐1β, the co‐localization of IL‐1 receptor type I (IL‐1RI) and Fos and NR1 in the spinal cord, and the effects of IL‐1 receptor antagonist (IL‐1ra) on NR1 phosphorylation and hyperalgesia in a rat model of inflammatory pain. Phosphorylation of NR1, an essential subunit of the NMDA receptor (NMDAR), is known to modulate NMDAR activity and facilitate pain. Hyperalgesia was induced by injecting complete Freund’s adjuvant (CFA, 0.08 ml, 40 μg Mycobacterium tuberculosis) into one hind paw of each rat. Paw withdrawal latency (PWL) was tested before CFA (−48 h) for baseline and 2 and 24 h after CFA to assess hyperalgesia. IL‐1ra was given (i.t.) 24 h before CFA to block the action of basal IL‐1β and 2 h prior to each of two PWL tests to block CFA‐induced IL‐1β. Spinal cords were removed for double immunostaining of IL‐1β/neuronal marker and IL‐1β/glial cell markers, IL‐1RI/Fos and IL‐1RI/NR1, and for Western blot to measure NR1 phosphorylation. The data showed that: (1) astrocytes produce IL‐1β, (2) IL‐1RI is localized in Fos‐ and NR1‐immunoreactive neurons within the spinal dorsal horn, and (3) IL‐1ra at 0.01 mg/rat significantly increased PWL (P < 0.05) and inhibited NR1 phosphorylation compared to saline control. The results suggest that spinal IL‐1β is produced by astrocytes and enhances NR1 phosphorylation to facilitate inflammatory pain.

Interleukin 1β facilitates bone cancer pain in rats by enhancing NMDA receptor NR-1 subunit phosphorylation

It has been shown that interleukin-1beta (IL-1beta) facilitates nociception during neuropathic and inflammatory pain, but its involvement in bone cancer pain and its mechanisms have not previously been established. This study is an investigation of IL-1beta spinal expression and the N-methyl-D-aspartate (NMDA) receptor (NMDAR) NR1 subunit phosphorylation during cancer pain, co-localization of IL-1 receptor type I (IL-1RI) and NMDAR in the spinal cord, and the effects of IL-1 receptor antagonist (IL-1ra) on NMDAR1 (NR1) phosphorylation and hyperalgesia in a rat model of bone cancer pain. Cancer was induced by injecting AT-3.1 prostate cancer cells into the tibia of the male Copenhagen rat. Phosphorylation of NR1, an essential subunit of the NMDAR, is known to modulate NMDAR activity and facilitate pain. Mechanical hyperalgesia, established by a decrease in paw withdrawal pressure threshold (PWPT), was measured at baseline and 2 h after IL-1ra treatment. IL-1ra was given (i.t.) daily for 7 days between days 13 and 19 after the cancer cell inoculation. Spinal cords were removed for Western blot to measure IL-1beta and NR1 phosphorylation and for double immunostaining of IL-1RI and NR1. The data showed that 1) spinal IL-1beta was up-regulated and NR1 phosphorylation was increased, 2) IL-1ra at 0.1 mg/rat significantly (P<0.05) inhibited mechanical hyperalgesia, increasing PWPT on day 14 from 71.1+/-3.1-85.3+/-4.6 g and on day 19 from 73.5.0+/-3.5-87.1+/-3.7 g, and inhibited NR1 phosphorylation compared with saline control, and 3) IL-1RI is localized in NR1-immunoreactive neurons within the spinal cord. The results suggest that spinal IL-1beta enhances NR1 phosphorylation to facilitate bone cancer pain.

Electroacupuncture suppresses hyperalgesia and spinal Fos expression by activating the descending inhibitory system.

Although electroacupuncture (EA) is widely used to treat pain, its mechanisms have not been completely understood. The present study investigated the descending inhibitory system involvement in EA action. Inflammatory pain was induced by injecting complete Freunds adjuvant subcutaneously into one hind paw of rats with dorsolateral funiculus lesions and sham-operated rats. EA treatment, 10 Hz at 3 mA, was given twice for 20 min each, once immediately post- and again 2 h post-Freunds adjuvant at GB 30, at the junction of the lateral 1/3 and medial 2/3 of the distance between the greater trochanter and sacral hiatus. For sham EA control, acupuncture needles were inserted bilaterally into GB 30 without electrical or manual stimulation. Paw withdrawal latency to a noxious thermal stimulus was measured at baseline and 20 min after EA treatment. Compared to sham EA, EA significantly (P<0.05, n=9) increased withdrawal latency of the inflamed hind paws in the sham-operated rats but not in those with dorsolateral funiculus lesions, indicating that lesioning blocked EA-produced anti-hyperalgesia. EA, compared to sham EA, also significantly inhibited Fos expression in laminae I-II of the spinal cord in the sham-operated rats (58.4+/-6.5 vs. 35.2+/-5.4 per section) but not in those with dorsolateral funiculus lesions. Further, EA activated serotonin- and catecholamine-containing neurons in the nucleus raphe magnus and locus coeruleus that project to the spinal cord. The results demonstrate that EA inhibits transmission of noxious messages and hyperalgesia by activating supraspinal neurons that project to the spinal cord.

Electroacupuncture attenuates bone cancer pain and inhibits spinal interleukin-1β expression in a rat model

BACKGROUND:Although pain affects the quality of life of cancer patients, current medical treatments are either ineffective or have side effects. In the present study we investigated the effect of electroacupuncture (EA) on cancer-induced hyperalgesia and expression of interleukin-1β (IL-1β), upregulation of which is related to the maintenance of persistent pain, in a rat model of bone cancer pain. METHODS:Cancer was induced by injecting AT-3.1 prostate cancer cells into the tibia of male Copenhagen rats. The resulting pain was treated with 10 Hz/2 mA/0.4 ms pulse EA for 30 min daily at the equivalent of the human acupoint GB30 (Huantiao) between Days 14 and 18 after cancer cell inoculation. For sham control, EA needles were inserted into GB30 without stimulation. Thermal hyperalgesia, a decrease in paw withdrawal latency to a noxious thermal stimulus, was measured at baseline and 20 min after EA treatment. IL-1β and its mRNA were respectively determined by immunohistochemistry and reverse transcription-polymerase chain reaction analysis. RESULTS:Thermal hyperalgesia developed between Days 12 and 18 after cancer cell inoculation. EA significantly (P < 0.05) attenuated this hyperalgesia, increasing paw withdrawal latency from 7.0 ± 0.3 s to 9.2 ± 0.4 s, and inhibited the upregulation of IL-1β and its mRNA compared to the sham control. Intrathecal injection of IL-1 receptor antagonist (IL-1ra, 0.1 mg/rat) also significantly inhibited cancer-induced thermal hyperalgesia. CONCLUSION:The data suggest that EA alleviates bone cancer pain, at least in part by suppressing IL-1β expression. The results support the clinical use of EA in the treatment of cancer pain.

Electroacupuncture inhibition of hyperalgesia in an inflammatory pain rat model: involvement of distinct spinal serotonin and norepinephrine receptor subtypes

BACKGROUND Although acupuncture analgesia is well documented, its mechanisms have not been thoroughly clarified. We previously showed that electroacupuncture (EA) activates supraspinal serotonin- and norepinephrine-containing neurones that project to the spinal cord. This study investigates the involvement of spinal alpha(2)-adrenoceptors (α2-ARs) and 5-hydroxytryptamine (serotonin) receptors (5-HTRs) in EA effects on an inflammatory pain rat model. METHODS Inflammatory hyperalgesia was induced by injecting complete Freunds adjuvant (CFA, 0.08 ml) into the plantar surface of one hind paw and assessed by paw withdrawal latency (PWL) to a noxious thermal stimulus. The selective α2a-AR antagonist BRL-44408, α2b-AR antagonist imiloxan hydrochloride, 5-HT2B receptor (5-HT2BR) antagonist SB204741, 5-HT3R antagonist LY278584, or 5-HT1AR antagonists NAN-190 hydrobromide, or WAY-100635 were intrathecally administered 20 min before EA or sham EA, which was given 2 h post-CFA at acupoint GB30. RESULTS EA significantly increased PWL compared with sham [7.20 (0.46) vs 5.20 (0.43) s]. Pretreatment with α2a-AR [5.35 (0.45) s] or 5-HT1AR [5.22 (0.38) s] antagonists blocked EA-produced anti-hyperalgesia; α2b-AR, 5-HT2BR, and 5-HT3R antagonist pretreatment did not. Sham plus these antagonists did not significantly change PWL compared with sham plus vehicle, indicating that the antagonists had little effect on PWL. Immunohistochemical staining demonstrated that α2a-ARs are on primary afferents and 5-HT1ARs are localized in N-methyl-d-aspartic acid (NMDA) subunit NR1-containing neurones in the spinal dorsal horn. CONCLUSIONS The data show that α2a-ARs and 5-HT1ARs are involved in the EA inhibition of inflammatory pain and that the NMDA receptors are involved in EA action.

Electroacupuncture activates corticotrophin-releasing hormone-containing neurons in the paraventricular nucleus of the hypothalammus to alleviate edema in a rat model of inflammation

BackgroundStudies show that electroacupuncture (EA) has beneficial effects in patients with inflammatory diseases. This study investigated the mechanisms of EA anti-inflammation, using a rat model of complete Freunds adjuvant (CFA)-induced hind paw inflammation and hyperalgesia.DesignFour experiments were conducted on male Sprague-Dawley rats (n = 6–7/per group). Inflammation was induced by injecting CFA into the plantar surface of one hind paw. Experiment 1 examined whether EA increases plasma adrenocorticotropic hormone (ACTH) levels. Experiments 2 and 3 studied the effects of the ACTH and corticotropin-releasing hormone (CRH) receptor antagonists, ACTH(11–24) and astressin, on the EA anti-edema. Experiment 4 determined whether EA activates CRH neurons in the paraventricular nucleus of the hypothalammus. EA treatment, 10 Hz at 3 mA and 0.1 ms pulse width, was given twice for 20 min each, once immediately post and again 2 hr post-CFA. Plasma ACTH levels, paw thickness, and paw withdrawal latency to a noxious thermal stimulus were measured 2 h and 5 h after the CFA.ResultsEA significantly increased ACTH levels 5 h (2 folds) after CFA compared to sham EA control, but EA alone in naive rats and CFA alone did not induce significant increases in ACTH. ACTH(11–24) and astressin blocked EA anti-edema but not EA anti-hyperalgesia. EA induced phosphorylation of NR1, an essential subunit of the N-methyl-D-aspartic acid (NMDA) receptor, in CRH-containing neurons of the paraventricular nucleus.ConclusionThe data demonstrate that EA activates CRH neurons to significantly increase plasma ACTH levels and suppress edema through CRH and ACTH receptors in a rat model of inflammation.

Electroacupuncture attenuates bone-cancer-induced hyperalgesia and inhibits spinal preprodynorphin expression in a rat model.

Cancer pain impairs the quality of life of cancer patients, but opioid intervention can cause significant side effects that further decrease quality of life. Although electroacupuncture (EA) has been used to treat cancer pain, its mechanisms are largely unknown. To examine its effects and underlying mechanisms on cancer pain, we injected AT‐3.1 prostate cancer cells into the tibia to induce bone cancer in the male Copenhagen rat. The resulting pain was treated with 10Hz/2mA/0.4ms pulse EA for 30min daily at the point equivalent to the human acupoint GB30 (Huantiao) between days 14 and 18 after the injection. For sham control, EA needles were inserted into GB30 without stimulation. Thermal hyperalgesia, a decrease in paw withdrawal latency (PWL) to a noxious thermal stimulus, and mechanical hyperalgesia, a decrease in paw withdrawal pressure threshold (PWPT), was measured at baseline and 20min after the EA treatment. Preprodynorphin mRNA and dynorphin were determined by RT‐PCR and immunohistochemistry, respectively. Thermal and mechanical hyperalgesia developed ipsilaterally between days 12 and 18 after cancer cell inoculation. EA significantly (P<0.05) attenuated this hyperalgesia, as shown by increased PWL and PWPT, and inhibited up‐regulation of preprodynorphin mRNA and dynorphin compared to sham control. Intrathecal injection of antiserum against dynorphin A (1–17) also significantly inhibited the cancer‐induced hyperalgesia.

Spinal interleukin-17 promotes thermal hyperalgesia and NMDA NR1 phosphorylation in an inflammatory pain rat model

Summary IL‐17 is up‐regulated in spinal astrocytes during inflammation and facilitates behavioral hyperalgesia through promoting phosphorylation of NR1, the essential subunit of the N‐methyl d‐aspartate receptor. ABSTRACT It is known that interleukin‐17 (IL‐17) is associated with autoimmune disorders and that peripheral IL‐17 plays a role in arthritis and neuropathic pain. The present study investigated the possibility of spinal cell expression of IL‐17 during inflammatory pain and possible IL‐17 involvement in such pain. Hyperalgesia was induced by injecting complete Freund adjuvant (CFA, 0.08 mL, 40 μg Mycobacterium tuberculosis) into one hind paw of the rat. Paw withdrawal latency (PWL) was tested before (−48 h) and 2 and 24 h after CFA injection to assess hyperalgesia. IL‐17 antibody (0.2–2 μg/rat) was given intrathecally (i.t.) 24 h before CFA to block the action of basal IL‐17 and 2 h before each of 2 PWL tests to block CFA‐induced IL‐17. I.t. recombinant IL‐17 (10–400 ng per rat) was administered to naive rats to determine its effects on PWL and phosphorylated NR1 (p‐NR1). p‐NR1 modulates N‐methyl‐d‐aspartate receptor (NMDAR) activity to facilitate pain. Spinal cords were removed for IL‐17 immunostaining, double immunostaining of IL‐17/cell markers and IL‐17 receptor A (IL‐17RA)/NR1, for Western blot testing of IL‐17, p‐NR1, IL‐17RA, and GFAP, for in situ IL‐17RA hybridization, and for real time polymerase chain reaction of IL‐17RA. The data reveal that IL‐17 is up‐regulated in activated and nonactivated astrocytes; that IL‐17RA is localized in NR1‐immunoreactive neurons and up‐regulated; and that IL‐17 antibody at 2 μg/rat significantly increased PWL (P < .05) and decreased p‐NR1 and IL‐17RA compared to control in CFA‐ and IL‐17‐injected rats. The results suggest that spinal IL‐17 is produced by astrocytes and enhances p‐NR1 to facilitate pain.

Corticosterone mediates electroacupuncture-produced anti-edema in a rat model of inflammation

BackgroundElectroacupuncture (EA) has been reported to produce anti-edema and anti-hyperalgesia effects on inflammatory disease. However, the mechanisms are not clear. The present study investigated the biochemical mechanisms of EA anti-inflammation in a rat model.MethodsThree experiments were conducted on male Sprague-Dawley rats (n = 7–8/per group). Inflammation was induced by injecting complete Freunds adjuvant (CFA) subcutaneously into the plantar surface of one hind paw. Experiment 1 measured plasma corticosterone (CORT) levels to see if EA regulates CORT secretion. Experiment 2 studied the effects of the adrenal gland on the therapeutic actions of EA using adrenalectomy (ADX) rats. Experiment 3 determined whether a prototypical glucocorticoid receptor antagonist, RU486, affects EA anti-edema. EA treatment, 10 Hz at 3 mA and 0.1 ms pulse width, was given twice, for 20 min each, once immediately after CFA administration and again 2 h post-CFA. Plasma CORT levels, paw thickness, indicative of the intensity of inflammation, and paw withdrawal latency (PWL) were measured 2 h and 5 h after the CFA injection.ResultsEA significantly increased plasma corticosterone levels 2 h (5 folds) and 5 h (10 folds) after CFA administration compared to sham EA control, but EA alone in naive rats and CFA alone did not induce significant increases in corticosterone. Adrenalectomy blocked EA-produced anti-edema, but not EA anti-hyperalgesia. RU486 (15 μl, 15 μg/μl), a prototypical glucocorticoid receptor antagonist, also prevented EA anti-edema.ConclusionThe data demonstrate that EA activates the adrenals to increase plasma corticosterone levels and suppress edema and suggest that EA effects differ in healthy subjects and in those with pathologies.

The effects of opioid receptor antagonists on electroacupuncture-produced anti-allodynia/hyperalgesia in rats with paclitaxel-evoked peripheral neuropathy

Research supports the effectiveness of acupuncture for conditions such as chronic low back and knee pain. In a five-patient pilot study the modality also improved the symptoms of chemotherapy-induced neuropathic pain. Using an established rat model of paclitaxel-induced peripheral neuropathy, we evaluated the effect of electroacupuncture (EA) on paclitaxel-induced hyperalgesia and allodynia that has not been studied in an animal model. We hypothesize that EA would relieve the paclitaxel-induced mechanical allodynia and hyperalgesia, which was assessed 30 min after EA using von Frey filaments. Beginning on day 13, the response frequency to von Frey filaments (4-15 g) was significantly increased in paclitaxel-injected rats compared to those injected with vehicle. EA at 10 Hz significantly (P<0.05) decreased response frequency at 4-15 g compared to sham EA; EA at 100 Hz only decreased response frequency at 15 g stimulation. Compared to sham EA plus vehicle, EA at 10 Hz plus either a μ, δ, or κ opioid receptor antagonist did not significantly decrease mechanical response frequency, indicating that all three antagonists blocked EA inhibition of allodynia and hyperalgesia. Since we previously demonstrated that μ and δ but not κ opioid receptors affect EA anti-hyperalgesia in an inflammatory pain model, these data show that EA inhibits pain through different opioid receptors under varying conditions. Our data indicate that EA at 10 Hz inhibits mechanical allodynia/hyperalgesia more potently than does EA at 100 Hz. Thus, EA significantly inhibits paclitaxel-induced allodynia/hyperalgesia through spinal opioid receptors, and EA may be a useful complementary treatment for neuropathic pain patients.