Min Yan1,2* Abstract PI3K Contributed to Modulation of Spinal Nociceptive Information Related to ephrinBs/EphBs Li-Na Yu1., Xue-Long Zhou2., Jing Yu 1 , Hao Huang1, Li-Shan Jiang2, Feng-Jiang Zhang1, Jun-Li Cao2, 1 Department of Anesthesiology, The Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, People’s Republic of China, 2 Jiangsu Province Key Laboratory of Anesthesiology and Center for Pain Research and Treatment, Xuzhou Medical College, Xuzhou, Jiangsu, People’s Republic of China There is accumulating evidence to implicate the importance of EphBs receptors and ephrinBs ligands were involved in modulation of spinal nociceptive information. However, the downstream mechanisms that control this process are not well understood. In the present study, we investigated whether phosphatidylinositol 3-kinase (PI3K), as the downstream effectors, participates in modulation of spinal nociceptive information related to ephrinBs/EphBs. Intrathecal injection of ephrinB1-Fc produced a dose- and time-dependent thermal and mechanical hyperalgesia, accompanied by the increase of spinal PI3K-p110c, phosphorylation of AKT (p-AKT) and c-Fos expression. Pre-treatment with PI3K inhibitor wortmannin or LY294002 prevented activation of spinal AKT induced by ephrinB1-Fc. Inhibition of spinal PI3K signaling dose-dependently prevented and reversed pain behaviors and spinal c-Fos protein expression induced by intrathecal injection of ephrinB1-Fc. Inhibition of EphBs receptors by intrathecal injection of EphB1-Fc reduced formalin-induced inflammation and chronic constrictive injury-induced neuropathic pain behaviors accompanied by decreased expression of spinal PI3K,p-AKT and c- Fos protein. Furthermore, pre-treatment with PI3K inhibitor wortmannin or LY294002 prevented ephrinB1-Fc-induced ERK activation in spinal. These data demonstrated that PI3K and PI3K crosstalk to ERK signaling contributed to modulation of spinal nociceptive information related to ephrinBs/EphBs. Citation: Yu L-N, Zhou X-L, Yu J, Huang H, Jiang L-S, et al. (2012) PI3K Contributed to Modulation of Spinal Nociceptive Information Related to ephrinBs/ EphBs. PLoS ONE 7(8): e40930. doi:10.1371/journal.pone.0040930 Editor: Mark L. Baccei, University of Cincinnatti, United States of America Received December 30, 2011; Accepted June 15, 2012; Published August 3, 2012 Copyright: 2012 Yu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by a grant from the medicine health project of Zhejiang province (No. 2008A088). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: yanminnina@hotmail.com . These authors contributed equally to this work. Introduction Central sensitization, an activity-dependent functional plasticity in spinal cord neurons, is one of the main causes of behavior hyperalgesia under pathologic conditions and has been under intensive investigation [1,2,3]. Activation of postsynaptic mem- brane receptors or ion channels, intracellular kinase cascades, and intranuclear gene expression contributes to the induction, development, and maintenance of central sensitization. Eph receptor tyrosine kinases and their membranebound ligands, ephrins, which are the largest family of receptor tyrosine kinase(RTK)system, are involved in diverse aspects of develop- ment, such as tissue patterning, angiogenesis, axon guidance, and synapse formation [4,5,6,7]. Recent advances indicate that Eph receptors and ephrin ligands are present in the adult brain and peripheral tissue and play a critical role in modulating multiple aspects of physiology and pathophysiology(e.g., activity-dependent synaptic plasticity, regulation of pain threshold, epileptogenesis, inflammation response, and excitotoxic neuronal death) [8,9,10,11]. Interestingly, several Eph receptors and ephrin ligands are also expressed in the adult rat spinal cord and the dorsal root ganglion [12,13,14]. Bundesen et al. [12] reported that EphB2 receptor was present in the laminae I–III of the dorsal horn and on small- and medium-sized dorsal root ganglion neurons but not on large-diameter neurons, two important sites for modulation of nociceptive information. Recently, some studies also demonstrated that activation of spinal ephrinBs/EphBs system played a critical role in the development and maintenance of chronic pain after peripheral nerve injury [15,16,17].These studies indicated that Ephrin/Eph system may be involved in physiologic and pathologic pain modulation in the spinal cord level. However, the downstream mechanisms that control this process are not well understood. PI3K, which phosphorylates the D3 position of the inositol ring of phosphoinositides and thereby generates intracellular signaling molecules, has been demonstrated to be essential for a plethora of physiological and pathological processes [18,19,20]. It is well established that activation of PI3K signaling is involved in the modulation of nociceptive information and central sensitization produced by intense noxious stimuli [21,22,23,24,25,26,27]. Importantly, PI3K is proposed to mediate central sensitization and hyperalgesia induced by activation of central RTK system NGF/TrkA, BDNF/TrkB and G-CSF/G-CSFR signaling [22,23,25]. Moreover, several lines of evidence have shown that regulation of PI3K pathway is associated with EphBs receptors activation in other study fields, such as intraplantar injection of ephrinB1-Fc induced pain [28]and retinal endothelial cell [29] and microvascular endothelial cell migration and proliferation PLoS ONE | www.plosone.org 1 August 2012 | Volume 7 | Issue 8 | e40930