PMID- 27245842 OWN - NLM STAT- MEDLINE DCOM- 20171010 LR - 20181113 IS - 1742-2094 (Electronic) IS - 1742-2094 (Linking) VI - 13 IP - 1 DP - 2016 May 31 TI - STIMs and Orai1 regulate cytokine production in spinal astrocytes. PG - 126 LID - 10.1186/s12974-016-0594-7 [doi] LID - 126 AB - BACKGROUND: Our previous study demonstrated that a store-operated calcium channel (SOCC) inhibitor (YM-58483) has central analgesic effects. However, the cellular and molecular mechanisms of such effects remain to be determined. It is well-known that glial cells play important roles in central sensitization. SOC entry (SOCE) has been implicated in many cell types including cortical astrocytes. However, the role of the SOCC family in the function of astrocytes has not been determined. Here, we thoroughly investigated the expression and the functional significance of SOCCs in spinal astrocytes. METHODS: Primary cultured astrocytes were prepared from neonatal (P2-P3) CD1 mice. Expressions of mRNAs and proteins were respectively assessed by real-time PCR and Western blot analysis. SOCE was measured using a calcium imaging system. Live-cell STIM1 translocation was detected using a confocal microscope. Cytokine levels were measured by the enzyme-linked immunosorbent assay. RESULTS: We found that the SOCC family is expressed in spinal astrocytes and that depletion of calcium stores from the endoplasmic reticulum by cyclopiazonic acid (CPA) resulted in a large sustained calcium entry, which was blocked by SOCC inhibitors. Using the siRNA knockdown approach, we identified STIM1 and Orai1 as primary components of SOCCs in spinal astrocytes. We also observed thapsigargin (TG)- or CPA-induced puncta formation of STIM1 and Orai1. In addition, activation of SOCCs remarkably promoted TNF-alpha and IL-6 production in spinal astrocytes, which were greatly attenuated by knockdown of STIM1 or Orai1. Importantly, knockdown of STIM2 and Orai1 dramatically decreased lipopolysaccharide-induced TNF-alpha and IL-6 production without changing cell viability. CONCLUSIONS: This study presents the first evidence that STIM1, STIM2, and Orai1 mediate SOCE and are involved in cytokine production in spinal astrocytes. Our findings provide the basis for future assessment of SOCCs in pain and other central nervous system disorders associated with abnormal astrocyte activities. FAU - Gao, Xinghua AU - Gao X AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. AD - Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, Nanjing, China. FAU - Xia, Jingsheng AU - Xia J AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. FAU - Munoz, Frances M AU - Munoz FM AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. FAU - Manners, Melissa T AU - Manners MT AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. FAU - Pan, Rong AU - Pan R AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. FAU - Meucci, Olimpia AU - Meucci O AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. FAU - Dai, Yue AU - Dai Y AD - Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, Nanjing, China. FAU - Hu, Huijuan AU - Hu H AD - Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA, 19102, USA. hhu@drexelmed.edu. LA - eng GR - R01 NS087033/NS/NINDS NIH HHS/United States GR - R21 NS077330/NS/NINDS NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural DEP - 20160531 PL - England TA - J Neuroinflammation JT - Journal of neuroinflammation JID - 101222974 RN - 0 (4-methyl-4'-(3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl)-1,2,3-thiadiazole-5-carboxanilide) RN - 0 (Anilides) RN - 0 (Cytokines) RN - 0 (ORAI1 Protein) RN - 0 (Orai1 protein, mouse) RN - 0 (Stim1 protein, mouse) RN - 0 (Stim2 protein, mouse) RN - 0 (Stromal Interaction Molecule 1) RN - 0 (Stromal Interaction Molecule 2) RN - 0 (Thiadiazoles) SB - IM MH - Anilides/pharmacology MH - Animals MH - Animals, Newborn MH - Astrocytes/drug effects/*metabolism MH - Cell Survival/drug effects/physiology MH - Cells, Cultured MH - Cytokines/*biosynthesis MH - Female MH - Mice MH - ORAI1 Protein/antagonists & inhibitors/*physiology MH - Pregnancy MH - Spinal Cord/drug effects/*metabolism MH - Stromal Interaction Molecule 1/antagonists & inhibitors/*physiology MH - Stromal Interaction Molecule 2/antagonists & inhibitors/*physiology MH - Thiadiazoles/pharmacology PMC - PMC4886427 OTO - NOTNLM OT - Astrocytes OT - Cytokine OT - Orai1 OT - STIM1 OT - Store-operated calcium channels OT - The spinal cord EDAT- 2016/06/02 06:00 MHDA- 2017/10/11 06:00 PMCR- 2016/05/31 CRDT- 2016/06/02 06:00 PHST- 2016/03/16 00:00 [received] PHST- 2016/05/23 00:00 [accepted] PHST- 2016/06/02 06:00 [entrez] PHST- 2016/06/02 06:00 [pubmed] PHST- 2017/10/11 06:00 [medline] PHST- 2016/05/31 00:00 [pmc-release] AID - 10.1186/s12974-016-0594-7 [pii] AID - 594 [pii] AID - 10.1186/s12974-016-0594-7 [doi] PST - epublish SO - J Neuroinflammation. 2016 May 31;13(1):126. doi: 10.1186/s12974-016-0594-7.