Pitfalls and solutions in assaying anandamide transport in cells.
Nonspecific binding of anandamide to plastic exhibits many features that could be mistaken as biological processes, thereby representing an important source of conflicting data on the uptake and release of this lipophilic substance. Herein, we propose an improved method to assay anandamide transport, by using glass slides (i.e., coverslips) as physical support to grow cells. Although the results obtained using plastic do not differ significantly from those obtained using glass, the new procedure has the advantage of being faster, simpler, and more accurate. In fact, the lack of aspecific adsorption of anandamide to the glass surface yields a lower background and a higher precision and accuracy in determining transport kinetics, especially for the export process. Remarkably, the kinetic parameters of anandamide uptake obtained with the old and the new procedures may be similar or different depending on the cell type, thus demonstrating the complexity of the interference of plastic on the transport process. In addition, the novel procedure is particularly suitable for visualization and measurement of anandamide transport in intact cells by using a biotinylated derivative in confocal fluorescence microscopy.
Oddi
Sergio
S
Department of Biomedical Sciences, University of Teramo, 64100 Teramo, Italy.
Fezza
Filomena
F
Catanzaro
Giuseppina
G
De Simone
Chiara
C
Pucci
Mariangela
M
Piomelli
Daniele
D
Finazzi-Agrò
Alessandro
A
Maccarrone
Mauro
M
D000327:Adsorption; D001095:Arachidonic Acids / Q000737:chemistry* / Q000378:metabolism*; D016477:Artifacts; D001692:Biological Transport; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D063388:Endocannabinoids; D005898:Glass / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D010969:Plastics / Q000737:chemistry; D053284:Polyunsaturated Alkamides / Q000737:chemistry* / Q000378:metabolism*; D017640:Silicates / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013499:Surface Properties
D001095:Arachidonic Acids; D063388:Endocannabinoids; D010969:Plastics; D053284:Polyunsaturated Alkamides; D017640:Silicates; C078814:anandamide
Br J Pharmacol. 2002 Nov;137(5):589-96
12381672
Cell Mol Life Sci. 2005 Feb;62(3):386-95
15723173
J Biol Chem. 2003 Sep 5;278(36):33896-903
12815050
Eur J Pharm Biopharm. 2006 Nov;64(3):369-78
16905298
Science. 1992 Dec 18;258(5090):1946-9
1470919
Prostaglandins Other Lipid Mediat. 2005 Sep;77(1-4):111-22
16099396
Proc Natl Acad Sci U S A. 2001 Jul 31;98(16):9371-6
11470906
Eur J Pharmacol. 2004 May 10;492(1):1-11
15145699
Brain Res Mol Brain Res. 1999 Feb 5;64(2):193-8
9931487
J Biol Chem. 2004 Oct 1;279(40):41991-7
15292270
PPAR Res. 2007;2007:23513
18288264
Life Sci. 2005 Aug 19;77(14):1584-604
15979096
Pharmacol Biochem Behav. 2005 Jun;81(2):248-62
15935456
J Biol Chem. 2005 Apr 1;280(13):12212-20
15657045
Proc Natl Acad Sci U S A. 2005 Dec 6;102(49):17852-7
16314570
J Pharmacol Exp Ther. 2000 Mar;292(3):960-7
10688610
Eur J Pharm Sci. 2004 Jun;22(2-3):181-9
15158903
Prog Neurobiol. 2007 Apr;81(5-6):349-79
17276576
Br J Pharmacol. 2001 Aug;133(8):1263-75
11498512
Chem Biol. 2009 Jun 26;16(6):624-32
19481477
J Neurochem. 1997 Aug;69(2):631-8
9231721
J Pharmacol Exp Ther. 2004 Nov;311(2):683-90
15284281
Mol Interv. 2006 Jun;6(3):149-61
16809476
Biochem J. 2004 May 15;380(Pt 1):265-72
14969584
Br J Pharmacol. 2006 Sep;149(1):73-81
16865094
Biochemistry. 2004 Jun 29;43(25):8184-90
15209515
Neuro Endocrinol Lett. 2004 Feb-Apr;25(1-2):14-23
15159677
Pharmacol Ther. 2007 Apr;114(1):13-33
17349697
Proc Natl Acad Sci U S A. 2003 Apr 1;100(7):4269-74
12655057
Curr Med Chem. 2007;14(25):2702-15
17979719
Nat Rev Drug Discov. 2008 May;7(5):438-55
18446159
Eur J Pharmacol. 2005 Jul 11;517(3):174-81
15987634
Cell Mol Life Sci. 2007 Jan;64(2):219-29
17187172
Trends Pharmacol Sci. 2007 Apr;28(4):180-7
17350694
Nat Rev Neurosci. 2003 Nov;4(11):873-84
14595399
Pharmacol Ther. 2004 Nov;104(2):117-35
15518883
Trends Pharmacol Sci. 2004 Feb;25(2):59-61
15106622
Biochem Soc Trans. 2006 Dec;34(Pt 6):1095-7
17073758
Nature. 1994 Dec 15;372(6507):686-91
7990962
Proc Natl Acad Sci U S A. 2008 Feb 19;105(7):2699-704
18263732
PLoS One. 2010 Apr 22;5(4):e10239
20422025
Cell Mol Life Sci. 2008 Mar;65(5):840-50
18213445
J Biol Chem. 2006 Apr 7;281(14):9066-75
16461355
Blood. 2002 Dec 1;100(12):4040-8
12393387
J Lipid Res. 2008 Jun;49(6):1216-23
18316795
Vitam Horm. 2009;81:25-53
19647107
Eur J Biochem. 2003 Sep;270(17):3498-506
12919314
Proc Natl Acad Sci U S A. 2009 Apr 14;106(15):6375-80
19307565
J Biol Chem. 2001 Mar 9;276(10):6967-73
11118429
Proc Natl Acad Sci U S A. 2004 Jun 8;101(23):8756-61
15138300