Lysophosphatidylethanolamine increases intracellular Ca2+ through LPA1 in PC-12 neuronal cells

https://doi.org/10.1016/j.bbrc.2015.04.042Get rights and content

Highlights

  • Lysophosphatidylethanolamine (LPE) increased intracellular Ca2+ concentration in PC-12 neuronal cells.

  • LPE-induced Ca2+ response was inhibited by LPA1 antagonist and other pharmacological tools.

  • LPE-induced Ca2+ response was mediated through LPA1-Gi/o protein-phospholipase C–IP3–Ca2+ rise-Ca2+ influx.

Abstract

G protein-coupled receptors (GPCRs) have been implicated in lysophosphatidylethanolamine (LPE)-induced increases in intracellular Ca2+ ([Ca2+]i), but in different cell types, this response may be dependent or independent of lysophosphatidic acid (LPA) GPCR. The effects of LPEs from Grifola frondosa on the neuronal differentiation and apoptosis of PC-12 neuronal cells have been previously reported. In the present study, the authors sought to identify the mechanism responsible for the effects of LPEs in PC-12 neuronal cells. LPE increase [Ca2+]i concentration-dependently in PC-12 neuronal cells, but this LPE-induced [Ca2+]i increase was less than that elicited by LPA. Studies using specific inhibitors showed that LPE-induced Ca2+ response was mediated via pertussis toxin-sensitive Gi/o proteins, edelfosine-sensitive phospholipase C, and 2-APB-sensitive IP3 receptor and by Ca2+ influx across the cell membrane, and that this did not involve the conversion of LPE to LPA. Furthermore, LPE- and LPA-induced responses were found to show homologous and heterologous desensitization in PC-12 cells. VPC32183 and Ki16425 (antagonists of LPA1 and LPA3) inhibited LPE-induced [Ca2+]i increases. Furthermore, AM-095 (a specific inhibitor of LPA1) inhibited LPE-induced Ca2+ response completely in PC-12 cells. These findings indicate LPE increases [Ca2+]i via a LPA1/Gi/o proteins/phospholipase C/IP3/Ca2+ rise/Ca2+ influx pathway in PC-12 neuronal cells.

Introduction

Lysophosphatidic acid (LPA) is a representative lyso-type intercellular mediator that acts through G protein-coupled receptors (GPCRs; LPA1-6) [1]. Another lysolipid, lysophosphatidylethanolamine (LPE) has been detected in human serum at concentrations of several hundreds of ng/ml [2], [3]. However, its action has not been much studied. In SK-OV3 and OVCAR-3 ovarian cancer cells, the effect of LPE on intracellular Ca2+ ([Ca2+]i) was suggested to be mediated through GPCRs, but not through GPCRs for LPA [4]. On the other hand, in MDA-MB-231 cells, LPE-induced [Ca2+]i increases were observed to be mediated via the GPCRs of LPA1 and CD97 [5], [6]. These findings show increases in [Ca2+]i by LPE may be either dependent or independent of LPA1 in different cells.

In a previous study, several types of LPEs isolated from Grifola frondosa were found to induce neuronal differentiation and suppress serum-deprivation induced apoptosis via MAPK activation in PC12 neuronal cells [7]. However, the involvements of GPCRs in these processes and the action mechanisms involved have not been elucidated. Therefore, in the present study, we investigated the signaling of LPE in PC-12 in neuronal cells and the mechanism involved.

Section snippets

Materials

1-Oleoyl-2-hydroxy-sn-glycero-3-phosphoethanolamine (18:1 LPE), 1-stearoyl-2-hydroxy-sn-glycero-3-phosphoethanolamine (18:0 LPE), 1-octadecyl-2-hydroxy-sn-glycero-3-phosphoethanolamine (ether-linked 18:0 LPE), 1-palmitoyl-2-hydroxy-sn-glycero-3-phosphoethanolamine (16:0 LPE), 1-myristoyl-2-hydroxy-sn-glycero-3-phosphoethanolamine (14:0 LPE), 1-oleoyl-2-hydroxy-sn-glycero-3-phosphate (sodium salt), and VPC32183 were purchased from Avanti Polar Lipids (Alabaster, AL, USA). Fura 2-AM, EGTA,

LPE increased [Ca2+]i in PC-12 neuronal cells

Previously, we observed LPE-induced increases of [Ca2+]i in MDA-MB-231 breast cancer cells and SK-OV3 ovarian cancer cells [4], [5]. In the present study, we treated cells with structurally different LPEs, that is oleoyl LPE (18:1 LPE), stearoyl LPE (18:0 LPE), octadecanyl LPE (ether-linked 18:0 LPE), palmitoyl LPE (16:0 LPE), and myristoyl LPE (14:0 LPE). As shown in Fig. 1-A and B, synthetic LPEs, such as, 18:1 LPE, 18:0 LPE, 18:0 ether-linked LPE, and 14:0 LPE, induced transient [Ca2+]i

Discussion

In the present study, LPE-induced [Ca2+]i increase was found to be mediated via LPA1 in PC-12 neuronal cells. Five results support this finding: 1) the heterologous desensitization exhibited by LPE- and LPA-induced [Ca2+]i increases, 2) the abrogation of LPE-induced response by the LPA1 and LPA 3 antagonists, Ki16425 and VPC32183, 3) the Gi/o-coupling character of LPA1 and the PTX-sensitivity of LPE-induced [Ca2+]i increase observed in PC-12 cells, 4) the complete inhibition of LPE-induced

Conflict of interest

None.

Acknowledgments

This research was supported by the Basic Science Research Program of the Korean National Research Foundation (NRF) funded by the Korean Ministry of Education, Science and Technology (NRF-2011-0021158) and by the Korean National Research Foundation funded by the Korean government (MSIP) (Grant no. 2009–0083538).

References (22)

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