Carrier-mediated transport and enzymatic hydrolysis of the endogenous cannabinoid 2-arachidonylglycerol.
The human astrocytoma cell line CCF-STTGI accumulates [3H]2-AG through an Na(+)- and energy-independent process, with a Km of 0.7 +/- 0.1 microM. Non-radioactive 2-AG, anandamide or the anandamide transport inhibitor 4-hydroxyphenyl arachidonamide inhibit [3H]2-AG uptake with half-maximal inhibitory concentrations (IC50) of 5.5 +/- 1.0 microM, 4.2 +/- 0.3 microM and 1.8 = 0.1 microM, respectively. A variety of lipid transport substrates and inhibitors interfere with neither [3H]2-AG nor [3H]anandamide uptake. These results suggest that 2-AG and anandamide are internalized in astrocytoma cells through a common carrier-mediated mechanism. After incubation with [3H]2-AG, radioactivity is recovered in phospholipids, monoacylglycerols (unmetabolized [3H]2-AG), free fatty acids ([3H]arachidonate) and, to a minor extent, diacylglycerols and triacylglycerols. Arachidonic acid (100 microM) and triacsin C (10 microM), an acyl-CoA synthetase inhibitor, prevent incorporation of [3H]arachidonic acid in phospholipids and significantly reduce [3H]2-AG transport. Thus, the driving force for 2-AG internalization may derive from the hydrolysis of 2-AG to arachidonate and the subsequent incorporation of this fatty acid into phospholipids.
Beltramo
M
M
Department of Pharmacology, University of California, Irvine 92697, USA.
Piomelli
D
D
D016718:Arachidonic Acid / Q000378:metabolism / Q000494:pharmacology; D001095:Arachidonic Acids / Q000378:metabolism / Q000494:pharmacology; D001254:Astrocytoma / Q000201:enzymology* / Q000473:pathology; D001667:Binding, Competitive / Q000187:drug effects; D001692:Biological Transport / Q000187:drug effects; D002121:Calcium Channel Blockers / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002855:Chromatography, Thin Layer; D004305:Dose-Response Relationship, Drug; D063388:Endocannabinoids; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005989:Glycerides / Q000378:metabolism* / Q000494:pharmacology; D005990:Glycerol / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007424:Intracellular Fluid / Q000187:drug effects / Q000201:enzymology; D050356:Lipid Metabolism; D008055:Lipids / Q000032:analysis; D018377:Neurotransmitter Agents / Q000378:metabolism*; D010743:Phospholipids / Q000032:analysis / Q000378:metabolism; D053284:Polyunsaturated Alkamides; D014226:Triazenes / Q000494:pharmacology; D014316:Tritium; D014407:Tumor Cells, Cultured
D001095:Arachidonic Acids; D002121:Calcium Channel Blockers; D002352:Carrier Proteins; D063388:Endocannabinoids; D004791:Enzyme Inhibitors; D005989:Glycerides; D008055:Lipids; D018377:Neurotransmitter Agents; D010743:Phospholipids; D053284:Polyunsaturated Alkamides; D014226:Triazenes; D014316:Tritium; D016718:Arachidonic Acid; C034613:triacsin C; C094503:glyceryl 2-arachidonate; D005990:Glycerol; C078814:anandamide; C107594:N-(4-hydroxyphenyl)arachidonylamide