a-Synuclein gene ablation increases docosahexaenoic acid incorporation and turnover in brain phospholipids Mikhail Y. Golovko,* Thad A. Rosenberger,* Søren Feddersen, Nils J. Færgeman and Eric J. Murphy* , à *Department of Pharmacology, Physiology, and Therapeutics, University of North Dakota, Grand Forks, North Dakota, USA  Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark àDepartment of Chemistry, University of North Dakota, Grand Forks, North Dakota, USA Abstract Previously, we demonstrated that ablation of a-synuclein (Snca) reduces arachidonate (20:4n-6) turnover in brain phospholipids through modulation of an endoplasmic reticu- lum-localized acyl-CoA synthetase (Acsl). The effect of Snca ablation on docosahexaenoic acid (22:6n-3) metabolism is unknown. In the present study, we examined the effect of Snca gene ablation on brain 22:6n-3 metabolism. We deter- mined 22:6n-3 uptake and incorporation into brain phospho- lipids by infusing awake, wild-type and Snca )/) mice with [1- 14 C]22:6n-3 using steady-state kinetic modeling. In addi- tion, because Snca modulates 20:4n-6-CoA formation, we assessed microsomal Acsl activity using 22:6n-3 as a sub- strate. Although Snca gene ablation does not affect brain 22:6n-3 uptake, brain 22:6n-3-CoA mass was elevated 1.5- fold in the absence of Snca. This is consistent with the 1.6- to 2.2-fold increase in the incorporation rate and turnover in ethanolamine glycerophospholipid, phosphatidylserine, and phosphatidylinositol pools. Increased 22:6n-3-CoA mass was not the result of altered Acsl activity, which was unaffected by the absence of Snca. While Snca bound 22:6n-3, K d = 1.0 ± 0.5 lmol/L, it did not bind 22:6n-3-CoA. These effects of Snca gene deletion on 22:6n-3 brain metabolism are opposite to what we reported previously for brain 20:4n-6 metabolism and are likely compensatory for the decreased 20:4n-6 metabolism in brains of Snca )/) mice. Keywords: acyl-coenzyme A synthetase, a-synuclein, brain, docosahexaenoic acid, kinetics, phospholipid. J. Neurochem. (2007) 101, 201–211. a-Synuclein (Snca) is a 140-amino acid-soluble protein expressed in the nervous system and accounts for 0.1–1% of brain cytosolic protein (Shibayama-Imazu et al. 1993; Iwai et al. 1995). It is widely distributed in neurons (Maroteaux et al. 1988; Jakes et al. 1994; Jo et al. 2000; McLean et al. 2000), astrocytes (Mori et al. 2002; Castagnet et al. 2005), oligodendroglia (Richter-Landsberg et al. 2000; Mori et al. 2002), and microglia (Papadopoulos et al. 2006). Snca over- expression and mutations are associated with familial Parkinson disease (Polymeropoulos et al. 1997; Kruger et al. 1998; Singleton et al. 2003; Zarranz et al. 2004), and aggregates containing Snca are hallmark of a number of neurodegenerative disorders (Iseki et al. 1998; Spillantini et al. 1998; Takeda et al. 1998; Trojanowski et al. 1998; Lippa et al. 1999). Despite the close association with neurodegenerative diseases, the physiological function of Snca is poorly defined. Snca may have a number of diverse roles in the nervous system, including regulation of synaptic vesicle mobiliza- tion (Murphy et al. 2000a; Cabin et al. 2002; Dalfo ´ et al. 2004), chaperone activity (Ostrerova et al. 1999; Souza et al. 2000), modulation of dopamine transporter (Lee et al. Received July 14, 2006; revised manuscript received September 9, 2006; accepted September 29, 2006. Address correspondence and reprint requests to Eric J. Murphy, Department of Pharmacology, Physiology, and Therapeutics, School of Medicine and Health Sciences, University of North Dakota, 501 N. Columbia Rd, Grand Forks, ND 58202-9037, USA. E-mail: emurphy@medicine.nodak.edu Abbreviations used: 16:0, palmitic acid; 20:4n-6, arachidonic acid; 22:6n-3, docosahexaenoic acid; [22:6n-3-CoA], mass of 22:6n-3-coen- zyme A in tissue; Acsl, acyl-coenzyme A synthetase; Cer PCho, sphingomyelin; ChoGpl, choline glycerophospholipids; CSP-a, cysteine- string protein-a; ER, endoplasmic reticulum; EtnGpl, ethanolamine glycerophospholipids; FABP, fatty acid binding protein; J FA,i , net rate of incorporation from 22:6n-3-CoA into individual phospholipid; PLA 2 , phospholipase A 2 ; PLD, phospholipase D; PtdIns, phosphatidylinositol; PtdSer, phosphatidylserine; PUFA, polyunsaturated fatty acids; Snca, a-synuclein; TLC, thin-layer chromatography; WT, wild type. Journal of Neurochemistry , 2007, 101, 201–211 doi:10.1111/j.1471-4159.2006.04357.x Ó 2007 The Authors Journal Compilation Ó 2007 International Society for Neurochemistry, J. Neurochem. (2007) 101, 201–211 201