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Down-regulation of Stargazin inhibits the enhanced surface delivery of ?-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor GluR1 subunit in rat dorsal horn and ameliorates postoperative pain.

ABSTRACT: Stargazin is the first transmembrane protein known to regulate synaptic targeting of ?-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors. However, it is unclear whether regulation of the surface delivery of spinal AMPA receptor subunits by stargazin contributes to postoperative pain development.Western blot analysis was used to examine changes in the surface delivery of AMPA receptor subunits, GluR1 and GluR2, in rat dorsal horn. The interaction between stargazin and GluR1 and GluR2 was examined by coimmunoprecipitation. Expression of stargazin was suppressed by intrathecal administration of small interfering RNA311.Membrane-bound GluR1, but not GluR2, in ipsilateral dorsal horn was increased at 3 h (1.49 ± 0.15-fold of ?-tubulin, mean ± SEM) and 1 day (1.03 ± 0.25) after incision, as compared with that in control rats (naive, 0.63 ± 0.23, P < 0.05, n = 6 per group). The amount of GluR1 coimmunoprecipitated with stargazin was greater at 3 h after incision (1.48 ± 0.31-fold of input) than that in control animals (0.45 ± 0.24, P < 0.05, n = 6 per group). Importantly, the increase in membrane GluR1 at 3 h after incision was normalized to near control level (0.72 ± 0.20-fold of ?-tubulin) by pretreatment with intrathecal stargazin small interfering RNA311 (0.87 ± 0.09), but not scrambled small interfering RNA (1.48 ± 0.24) or vehicle (1.25 ± 0.13, P < 0.05, n = 6 per group). Stargazin small interfering RNA311 pretreatment prevented the increase in stargazin-GluR1 interaction and decreased postoperative pain after incision.This study suggests a critical role of stargazin-mediated surface delivery of GluR1 subunit in the development of postoperative pain. A better therapeutic strategy for postoperative pain may involve selectively down-regulating spinal stargazin to inhibit synaptic targeting of GluR1 subunit.

SUBMITTER: Guo R

PROVIDER: S-EPMC4165695 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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Down-regulation of Stargazin inhibits the enhanced surface delivery of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor GluR1 subunit in rat dorsal horn and ameliorates postoperative pain.

Guo Ruijuan R Zhao Yujie Y Zhang Meijuan M Wang Yue Y Shi Rong R Liu Yang Y Xu Jie J Wu Anshi A Yue Yun Y Wu Jing J Guan Yun Y Wang Yun Y

Anesthesiology 20140901 3

<h4>Background</h4>Stargazin is the first transmembrane protein known to regulate synaptic targeting of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors. However, it is unclear whether regulation of the surface delivery of spinal AMPA receptor subunits by stargazin contributes to postoperative pain development.<h4>Methods</h4>Western blot analysis was used to examine changes in the surface delivery of AMPA receptor subunits, GluR1 and GluR2, in rat dorsal horn. The interaction ...[more]

PMID: 25093662

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Project description:Vesicular glutamate transporter-2 (VGluT2) mediates the uptake of glutamate into synaptic vesicles in neurons. Spinal cord dorsal horn interneurons are highly heterogeneous and molecularly diverse. The functional significance of VGluT2-expressing dorsal horn neurons in physiological and pathological pain conditions has not been explicitly demonstrated. Designer receptors exclusively activated by designer drugs (DREADDs) are a powerful chemogenetic tool to reversibly control neuronal excitability and behavior. Here, we used transgenic mice with Cre recombinase expression driven by the VGluT2 promoter, combined with the chemogenetic approach, to determine the contribution of VGluT2-expressing dorsal horn neurons to nociceptive regulation. Adeno-associated viral vectors expressing double-floxed Cre-dependent G?q-coupled human M3 muscarinic receptor DREADD (hM3D)-mCherry or G?i-coupled ?-opioid receptor DREADD (KORD)-IRES-mCitrine were microinjected into the superficial spinal dorsal horn of VGluT2-Cre mice. Immunofluorescence labeling showed that VGluT2 was predominantly expressed in lamina II excitatory interneurons. Activation of excitatory hM3D in VGluT2-expressing neurons with clozapine N-oxide caused a profound increase in neuronal firing and synaptic glutamate release. Conversely, activation of inhibitory KORD in VGluT2-expressing neurons with salvinorin B markedly inhibited neuronal activity and synaptic glutamate release. In addition, chemogenetic stimulation of VGluT2-expressing neurons increased mechanical and thermal sensitivities in naive mice, whereas chemogenetic silencing of VGluT2-expressing neurons reversed pain hypersensitivity induced by tissue inflammation and peripheral nerve injury. These findings indicate that VGluT2-expressing excitatory neurons play a crucial role in mediating nociceptive transmission in the spinal dorsal horn. Targeting glutamatergic dorsal horn neurons with inhibitory DREADDs may be a new strategy for treating inflammatory pain and neuropathic pain.

Dataset Information

Down-regulation of Stargazin inhibits the enhanced surface delivery of ?-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor GluR1 subunit in rat dorsal horn and ameliorates postoperative pain.

Publications

Down-regulation of Stargazin inhibits the enhanced surface delivery of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor GluR1 subunit in rat dorsal horn and ameliorates postoperative pain.

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