Enzymatic formation of N-acylethanolamines from N-acylethanolamine plasmalogen through N-acylphosphatidylethanolamine-hydrolyzing phospholipase D-dependent and -independent pathways

Published Date
October 2011, Vol.1811(10):565–577, doi:10.1016/j.bbalip.2011.07.009

Author 

• Kazuhito Tsuboi ^a
• Yasuo Okamoto ^a,b
• Natsuki Ikematsu ^c
• Manami Inoue ^c
• Yoshibumi Shimizu ^c
• Toru Uyama ^a
• Jun Wang ^a,d
• Dale G. Deutsch ^e
• Matthew P. Burns ^e
• Nadine M. Ulloa ^e
• Akira Tokumura ^c
• Natsuo Ueda ^a,,

• aDepartment of Biochemistry, Kagawa University School of Medicine, 1750-1 Ikenobe, Miki, Kagawa 761-0793, Japan
• bDepartment of Physiology, Kanazawa University Graduate School of Medicine, Kanazawa, Ishikawa 920-8640, Japan
• cInstitute of Health Biosciences, University of Tokushima Graduate School, Tokushima 770-8505, Japan
• dDepartment of Anesthesiology, The First Affiliated Hospital, China Medical University, Shenyang 110001, China
• eDepartment of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York 11794, USA

Received 11 May 2011. Revised 28 June 2011. Accepted 14 July 2011. Available online 23 July 2011. 

Abstract

Bioactive N-acylethanolamines include anandamide (an endocannabinoid), N-palmitoylethanolamine (an anti-inflammatory), and N-oleoylethanolamine (an anorexic). In the brain, these molecules are formed from N-acylphosphatidylethanolamines (NAPEs) by a specific phospholipase D, called NAPE-PLD, or through NAPE-PLD-independent multi-step pathways, as illustrated in the current study employing NAPE-PLD-deficient mice. Although N-acylethanolamine plasmalogen (1-alkenyl-2-acyl-glycero-3-phospho(N-acyl)ethanolamine, pNAPE) is presumably a major class of N-acylethanolamine phospholipids in the brain, its enzymatic conversion to N-acylethanolamines is poorly understood. In the present study, we focused on the formation of N-acylethanolamines from pNAPEs. While recombinant NAPE-PLD catalyzed direct release of N-palmitoylethanolamine from N-palmitoylethanolamine plasmalogen, the same reaction occurred in the brain homogenate of NAPE-PLD-deficient mice, suggesting that this reaction occurs through both the NAPE-PLD-dependent and -independent pathways. Liquid chromatography-mass spectrometry revealed a remarkable accumulation of 1-alkenyl-2-hydroxy-glycero-3-phospho(N-acyl)ethanolamines (lyso pNAPEs) in the brain of NAPE-PLD-deficient mice. We also found that brain homogenate formed N-palmitoylethanolamine, N-oleoylethanolamine, and anandamide from their corresponding lyso pNAPEs by a Mg²⁺-dependent “lysophospholipase D”. Moreover, the brain levels of alkenyl-type lysophosphatidic acids, the other products from lyso pNAPEs by lysophospholipase D, also increased in NAPE-PLD-deficient mice. Glycerophosphodiesterase GDE1 can hydrolyze glycerophospho-N-acylethanolamines to N-acylethanolamines in the brain. In addition, we discovered that recombinant GDE1 has a weak activity to generate N-palmitoylethanolamine from its corresponding lyso pNAPE, suggesting that this enzyme is at least in part responsible for the lysophospholipase D activity. These results strongly suggest that brain tissue N-acylethanolamines, including anandamide, can be formed from N-acylated plasmalogen through an NAPE-PLD-independent pathway as well as by their direct release via NAPE-PLD.

Highlights

► N-Acylethanolamine (NAE) plasmalogens are precursors of NAEs in the brain. ► The phospholipase D-type enzyme NAPE-PLD converted NAE plasmalogen to NAE. ► Brain homogenates of NAPE-PLD-null mice also formed NAE from NAE plasmalogen. ► The homogenates could hydrolyze N-acylated plasmalogen lysophospholipids to NAEs. ► Thus NAPE-PLD-independent formation of NAEs from NAE plasmalogens was also shown.

Keywords

• N-acylethanolamine
• Anandamide
• Endocannabinoid
• NAPE-PLD
• Phospholipase
• Plasmalogen

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  Corresponding author at: Department of Biochemistry, Kagawa University School of Medicine, 1750-1 Ikenobe, Miki, Kagawa 761-0793, Japan. Tel.: + 81 87 891 2102; fax: + 81 87 891 2105.

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