Intrathecal α2-Adrenergic Agonists Stimulate Acetylcholine and Norepinephrine Release from the Spinal Cord Dorsal Horn in Sheep

Background Intrathecal injection of clonidine and dexmedetomidine produce behavioral analgesia by an alpha 2-adrenergic mechanism. Functional and anatomic studies suggest that this analgesia is mediated by cholinergic activation. This hypothesis was directly tested by measuring extracellular acetylcholine concentrations in spinal cord interstitial fluid by means of microdialysis after intrathecal injection of these alpha 2-adrenergic agonists in sheep. Methods Twelve sheep with chronically implanted thoracic intrathecal catheters were anesthetized with halothane. Multiple 200-micron-diameter dialysis fibers were inserted surgically at a mid-thoracic level through the dorsal horn and perfused with artificial cerebrospinal fluid. After baseline sampling, either clonidine (100 micrograms), dexmedetomidine (100 micrograms), or saline were injected intrathecally. Microdialysis samples were analyzed by high-pressure liquid chromatography for acetylcholine and norepinephrine. Results Both alpha 2-adrenergic agonists increased acetylcholine in microdialysate, whereas intrathecal saline had no effect. Analysis of the raw data showed that all groups differed significantly, with greater levels of acetylcholine following administration of dexmedetomidine than clonidine or saline. Unexpectedly, intrathecal clonidine also increased microdialysate norepinephrine levels. Conclusions These data are consistent with previous experiments measuring acetylcholine concentrations in cerebrospinal fluid and support analgesia from alpha 2-adrenergic agonists mediated in part by cholinergic activation. In addition, the increase in norepinephrine concentrations after intrathecal administration of clonidine suggest stimulation of norepinephrine release by this agent.

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Spinal Cord Glycine Transporter 2 Mediates Bilateral ST35 Acupoints Sensitization in Rats with Knee Osteoarthritis

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2019/7493286 ◽

2019 ◽

Vol 2019 ◽

pp. 1-17 ◽

Cited By ~ 1

Author(s):

Fuhai Bai ◽

Yongyuan Ma ◽

Haiyun Guo ◽

Yuheng Li ◽

Feifei Xu ◽

...

Keyword(s):

Spinal Cord ◽

Mast Cell ◽

Knee Osteoarthritis ◽

Dorsal Horn ◽

Intrathecal Administration ◽

Mast Cell Degranulation ◽

Spinal Cord Dorsal Horn ◽

Glycine Transporter ◽

Mechanical Threshold ◽

Glycine Transporter 2

The concept of “acupoint sensitization” refers to the functional status of acupoint switches from silent to active under pathological conditions. In clinic, acupoint sensitization provides important guidance for acupoints selection in different diseases. However, the mechanism behind this phenomenon remains unclear. We generated a model of knee osteoarthritis (KOA) by intra-articular injection of monosodium iodoacetate (MIA) into the left knee of rats. The paw withdrawal mechanical threshold (PWMT) and the total number of mast cells as well as mast cell degranulation rate (MCDR) of acupoint tissue were used to test whether the acupoints were sensitized. The results showed that KOA resulted in a reduced mechanical threshold and elevated total number of mast cell as well as mast cell degranulation rate at bilateral ST35 (Dubi) but not GB37 (Guangming) or nonacupoint area. The acupoint sensitization was accompanied by upregulation of glycine transporter 2 (GlyT2) and reduction of extracellular glycine levels in the bilateral dorsal horns of the spinal cord at L3-5. Selective inhibition of GlyT2 or intrathecal administration of glycine attenuated ST35 acupoint sensitization. The sensitization of bilateral ST35 was blocked after intraspinal GlyT2 short hairpin (sh) RNA (GlyT2-shRNA) microinjection to specifically downregulate GlyT2 expression in the left side (ipsilateral) L3-5 spinal cord dorsal horn before MIA injection. Moreover, electroacupuncture (EA) stimulation at ST35 ameliorated articular pathological lesions and improved KOA-related pain behaviors. GlyT2-shRNA injection reversed EA-induced pain relief but not EA-induced reduction of joint lesions. Overall, this study demonstrated that spinal GlyT2, especially elevated GlyT2 expression in the ipsilateral dorsal horn of the spinal cord, is a crucial mediator of ST35 acupoint sensitization in KOA rats.

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Antinociceptive and Hemodynamic Effects of a Novel α2-Adrenergic Agonist, MPV-2426, in Sheep

Anesthesiology ◽

10.1097/00000542-199911000-00036 ◽

1999 ◽

Vol 91 (5) ◽

pp. 1425-1425 ◽

Cited By ~ 18

Author(s):

James C. Eisenach ◽

Patricia Lavand'homme ◽

Chuanyao Tong ◽

Jen-Kun Cheng ◽

Hui-Lin Pan ◽

...

Keyword(s):

Spinal Cord ◽

Blood Flow ◽

Cerebrospinal Fluid ◽

Cord Blood ◽

Intrathecal Injection ◽

Adrenergic Agonists ◽

Hemodynamic Effects ◽

Adrenergic Agonist ◽

Spinal Cord Blood Flow ◽

Arterial Blood

Background alpha2-Adrenergic agonists produce analgesia primarily by a spinal action and hypotension and bradycardia by actions at several sites. Clonidine is approved for epidural use in the treatment of neuropathic pain, but its wider application is limited by hemodynamic side effects. This study determined the antinociceptive and hemodynamic effects of a novel alpha2-adrenergic agonist, MPV-2426, in sheep. Methods Forty sheep of mixed Western breeds with indwelling catheters were studied. In separate studies, antinociception to a mechanical stimulus, hemodynamic effects, arterial blood gas tensions, cerebrospinal fluid pharmacokinetics, and spinal cord blood flow was determined after epidural, intrathecal, and intravenous injection of MPV-2426. Results MPV-2426 produced antinociception with greater potency intrathecally (ED50 = 49 microg) than epidurally (ED50 = 202 microg), whereas intravenous administration had no effect. Intrathecal injection, in doses up to three times the ED95, failed to decrease systemic or central arterial blood pressures or heart rate, whereas larger doses, regardless of route, increased systemic arterial pressure. Bioavailability in cerebrospinal fluid was 7% after epidural administration and 0.17% after intravenous administration. Intrathecal MPV-2426, in an ED95 dose and three times this dose, produced a dose-independent reduction in thoracic and lumbar spinal cord blood flow. Conclusions MPV-2426 shares many characteristics of other alpha2-adrenergic agonists examined in sheep, but differs from clonidine and dexmedetomidine by lack of antinociception and minimal reduction in oxygen partial pressure after large intravenous and epidural injections. No hemodynamic depression was observed after intrathecal injection at antinociceptive doses. These results suggest this compound may be an effective spinal analgesic in humans with less hypotension than clonidine, although its relative potency to cause sedation was not tested in this study.

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Comparison of the Effects of Treatment with Intrathecal Lidocaine Given before and after Formalin on Both Nociception and Fos Expression in the Spinal Cord Dorsal Horn

Anesthesiology ◽

10.1097/00000542-199801000-00023 ◽

1998 ◽

Vol 88 (1) ◽

pp. 157-164 ◽

Cited By ~ 38

Author(s):

Kiran Yashpal ◽

Patrick Mason ◽

John E. McKenna ◽

Sushil K. Sharma ◽

James L. Henry ◽

...

Keyword(s):

Spinal Cord ◽

Dorsal Horn ◽

Intrathecal Injection ◽

Post Treatment ◽

Spinal Cord Dorsal Horn ◽

Formalin Injection ◽

Second Phase ◽

Nociceptive Responses ◽

Spinal Cord Dorsal ◽

Fos Expression

Background It has been proposed that the measure of noxious stimulus-induced Fos (the protein product of the immediate early gene c-fos) expression in the spinal cord dorsal horn of laboratory animals may provide an estimate of the potential of specific treatments to produce preemptive analgesia. The present study examined this hypothesis by comparing the effects of intrathecal lidocaine given before and after hindpaw formalin injection on persistent nociceptive responses and Fos expression in spinal cord dorsal horn of rats. Methods Formalin-induced nociception and Fos expression in the spinal cord, in response to a 50-microl injection of 2.5% formalin into the hind paw, were assessed in rats given an intrathecal injection of 50 microl 2% lidocaine by lumbar puncture between the L5 and L6 vertebrae, either 3 min before (pretreatment) or 5 min after (post-treatment) formalin injection. Results Pain behaviors (hindpaw licking, elevation, and favoring) in the second phase of the formalin test were significantly reduced by pretreatment, but were unaffected by post-treatment. The number of immunocytochemically stained Fos-positive cells and the immunoprecipitation of the Fos antibodies were reduced by pretreatment, and were also reduced, to a lesser extent, by post-treatment. Conclusions The finding that persistent nociceptive behaviors and Fos expression were suppressed by intrathecal lidocaine pretreatment suggests that nociception in the second phase of the formalin test depends on increases in central hyperexcitability generated during the first phase. On the other hand, the finding that the intrathecal injection of lidocaine after formalin treatment reduced Fos expression but not nociceptive responses indicates an uncoupling of the behavioral and Fos protein responses to formalin and suggests that changes in Fos expression may not be a good predictor of the ability of agents to produce preemptive analgesia.

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Intravenous Opioids Stimulate Norepinephrine and Acetylcholine Release in Spinal Cord Dorsal Horn

Anesthesiology ◽

10.1097/00000542-199601000-00017 ◽

1996 ◽

Vol 84 (1) ◽

pp. 143-154 ◽

Cited By ~ 65

Author(s):

Herve Bouaziz ◽

Chuanyao Tong ◽

Young Yoon ◽

David D. Hood ◽

James C. Eisenach

Keyword(s):

Spinal Cord ◽

Cerebrospinal Fluid ◽

Dorsal Horn ◽

Acetylcholine Release ◽

Ventral Horn ◽

Spinal Cord Dorsal Horn ◽

Cervical Cord ◽

Neural Pathways ◽

Functional Studies ◽

Intravenous Morphine

Background Opioids produce analgesia by direct effects as well as by activating neural pathways that release nonopioid transmitters. This study tested whether systematically administered opioids activate descending spinal noradrenergic and cholinergic pathways. Methods The effect of intravenous morphine on cerebrospinal fluid and dorsal horn microdialysate concentrations of norepinephrine and acetylcholine was examined in 20 sheep. Animals received either intravenous morphine or fentanyl alone, or morphine plus intravenous naloxone or intrathecal idazoxan. Results Intravenous morphine (0, 0.5, 1 mg/kg, intravenous) produced dose-dependent increases in cerebrospinal fluid norepinephrine and acetylcholine, but not epinephrine or dopamine. Morphine's effect was blocked by intravenous naloxone and by intrathecal idazoxan. In microdialysis experiments, intravenous morphine increased the concentration of norepinephrine and acetylcholine, but not epinephrine or dopamine, in the dorsal horn. In contrast, intravenous morphine exerted no effect on any of these monoamines in the ventral horn. Intravenous naloxone and cervical cord transection each blocked morphine's effect on dorsal horn norepinephrine. Conclusions These results support functional studies that indicate that systematically administered opioids cause spinal norepinephrine and acetylcholine release by a naloxone-sensitive mechanism. Idazoxan blockade of morphine's effects on cerebrospinal fluid norepinephrine was unexpected, and suggests that both norepinephrine and acetylcholine release in the spinal cord may be regulated by alpha 2-adrenoceptors. Microdialysis experiments suggest increased norepinephrine and acetylcholine levels in cerebrospinal fluid resulted from intravenous morphine-induced activation of bulbospinal pathways.

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Electroacupuncture Inhibits the Activity of Astrocytes in Spinal Cord in Rats with Visceral Hypersensitivity by Inhibiting P2Y1 Receptor-Mediated MAPK/ERK Signaling Pathway

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2020/4956179 ◽

2020 ◽

Vol 2020 ◽

pp. 1-12 ◽

Cited By ~ 1

Author(s):

Jingming Zhao ◽

Hui Li ◽

Chong Shi ◽

Tiezheng Yang ◽

Baoshi Xu

Keyword(s):

Spinal Cord ◽

Acetic Acid ◽

Dorsal Horn ◽

Molecular Mechanisms ◽

Intrathecal Injection ◽

Inhibitory Effect ◽

Visceral Hypersensitivity ◽

Spinal Cord Dorsal Horn ◽

Male Rats ◽

Spinal Cord Dorsal

Background. Irritable bowel syndrome (IBS) is a chronic functional bowel disease characterized by abdominal pain and changes in bowel habits in the absence of organic disease. Electroacupuncture (EA) has been shown to alleviate visceral hypersensitivity (VH) in IBS rat models by inhibiting the activation of astrocytes in the spinal cord. However, the underlying molecular mechanisms mediated by P2Y1 receptor of this effect of electroacupuncture remain unclear. Aim. To explore whether EA inhibits the activity of astrocytes in the spinal cord dorsal horn of rat with visceral hypersensitivity by inhibiting P2Y1 receptor and its downstream mitogen activated protein kinase/extracellular regulated kinase 1 (MAPK/ERK) pathway. Methods. Ten-day-old Sprague-Dawley (SD) male rats were given an intracolonic injection of 0.2 ml of 0.5% acetic acid (AA) to establish a visceral hypersensitivity model. EA was performed at Zusanli (ST 36) and Shangjuxu (ST 37) at 100 Hz for 1.05 s and 2 Hz for 2.85 s alternately, pulse width for 0.1 ms, 1 mA, 30 min/d, once a day, for 1 week. Cytokines IL-6, IL-1β, and TNF-α were analyzed by ELISA. The expressions of the P2Y1 receptor and pERK1/2 were analyzed by Western Blot and real-time PCR in the model and EA treated animals to explore the molecular mechanism of EA in inhibiting the activity of spinal cord dorsal horn (L6-S2 segment) astrocytes in rats with IBS visceral hypersensitivity. Results. EA significantly reduced the behavioral abdominal withdrawal reflex score (AWRs) of IBS rats with visceral hypersensitivity induced by AA. For comparison, intrathecal injection of astrocytes activity inhibitor fluorocitrate (FCA) also reduced visceral hypersensitivity in IBS rats. EA at Zusanli and Shangjuxu inhibited the mRNA and protein expression of the glial fibrillary acidic protein (GFAP) and in rat spinal cord and reduced the release of inflammatory cytokines IL-6, IL-1, and TNF-α from astrocytes. EA also inhibited acetic acid-induced expression of the P2Y1 receptor in the spinal cord. Adenosine 5′-[β-thio]diphosphate trilithium salt (ADP), a selective agonist of the P2Y1 receptor, reversed the inhibitory effect of EA on visceral hypersensitivity. ADP also overrode the downregulation of GFAP by EA. Conversely, MRS2179 (MRS), a selective antagonist of the P2Y1 receptor, inhibited visceral hypersensitivity suggesting that EA negatively regulated the P2Y1 receptor in astrocytes. Acetic acid also upregulated the expression of pERK1/2 protein and mRNA in the spinal cord of rats with visceral hypersensitivity, which was inhibited by EA and the inhibitory effect of EA on pERK1/2 was reversed by ADP. We also found that SCH772984 (SCH), an ERK1/2 inhibitor (10 μg, 10 μl), reduced the AWRs. Compared to the SCH group, AWR scores in SCH + EA group were decreased. The application of P2Y1 agonists failed to increase the AWR scores after the intrathecal injection of SCH. GFAP level in the spinal cord in the SCH group was significantly reduced when compared to the Model group. The GFAP expression was further reduced in the SCH + EA group. Conclusion. EA inhibited astrocyte activity in the spinal cord dorsal horn of rat with IBS visceral hypersensitivity by inhibiting the P2Y1 receptor and its downstream, PKC, and MAPK/ERK1/2 pathways.

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