Change search
ReferencesLink to record
Permanent link

Direct link
The 'specific' tyrosine kinase inhibitor genistein inhibits the enzymic hydrolysis of anandamide: implications for anandamide uptake
Umeå University, Faculty of Medicine, Pharmacology and Clinical Neuroscience, Pharmacology. Farmakologi.
Umeå University, Faculty of Medicine, Pharmacology and Clinical Neuroscience, Pharmacology. Farmakologi.
2007 (English)In: British Journal of Pharmacology, ISSN 0007-1188, Vol. 150, no 7, 951-960 p.Article in journal (Refereed) Published
Abstract [en]

Background and Purpose: The cellular uptake of anandamide is reduced by inhibitors of fatty acid amide hydrolase (FAAH) and by agents disrupting endocytotic mechanisms. However, it is not clear if these events occur over the same time frame and if they occur to the same extent in different cells. We have therefore investigated the effects of such compounds in three cell lines of different origins using different assay incubation times and temperatures.

Experimental approach: FAAH activity and cellular uptake of anandamide was measured using anandamide, radio-labelled either in the ethanolamine or arachidonoyl part of the molecule.

Key results: The FAAH inhibitor URB597 inhibited the uptake of anandamide into C6 glioma, RBL2H3 basophilic leukaemia cells and P19 embryonic carcinoma cells at incubation time 4 min. However, a time-dependent and temperature-sensitive residual uptake remained after URB597 treatment. The combination of progesterone and nystatin reduced the uptake, but also decreased the amount of anandamide retained by the wells. Genistein inhibited anandamide uptake in a manner that was not additive to that of URB597. However, genistein was a potent competitive inhibitor of FAAH (Ki value 8 mM).

Conclusions and implications: The reduction of anandamide uptake by genistein can be explained by its ability to inhibit FAAH with a potency which overlaps that for inhibition of tyrosine kinase. The FAAH- resistant but time-dependent uptake of anandamide is seen in all three cell lines studied and is thus presumably a generally occurring process.

Place, publisher, year, edition, pages
2007. Vol. 150, no 7, 951-960 p.
Keyword [en]
Amidohydrolases/*antagonists & inhibitors/metabolism, Animals, Arachidonic Acids/*metabolism, Benzamides/pharmacology, Brain/enzymology, Carbamates/pharmacology, Cell Line; Tumor, Genistein/*pharmacology, Hydrolysis, Mice, Nystatin/pharmacology, Polyunsaturated Alkamides/*metabolism, Progesterone/pharmacology, Protein Kinase Inhibitors/pharmacology, Protein-Tyrosine Kinases/antagonists & inhibitors, Rats
URN: urn:nbn:se:umu:diva-5987DOI: 10.1038/sj.bjp.0707172PubMedID: 17325653OAI: diva2:145655
Available from: 2007-12-04 Created: 2007-12-04Bibliographically approved
In thesis
1. The cellular processing of the endocannabinoid anandamide and its pharmacological manipulation
Open this publication in new window or tab >>The cellular processing of the endocannabinoid anandamide and its pharmacological manipulation
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Anandamide (arachidonoyl ethanolamide, AEA) and 2-arachidonoyl glycerol (2-AG) exert most of their actions by binding to cannabinoid receptors. The effects of the endocannabinoids are short-lived due to rapid cellular accumulation and metabolism, for AEA, primarily by the enzymes fatty acid amide hydrolase (FAAH). This has led to the hypothesis that by inhibition of the cellular processing of AEA, beneficial effects in conditions such as pain and inflammation can be enhanced. The overall aim of the present thesis has been to examine the mechanisms involved in the cellular processing of AEA and how they can be influenced pharmacologically by both synthetic natural compounds.

Liposomes, artificial membranes, were used in paper I to study the membrane retention of AEA. The AEA retention mimicked the early properties of AEA accumulation, such as temperature-dependency and saturability.

In paper II, FAAH was blocked by a selective inhibitor, URB597, and reduced the accumulation of AEA into RBL2H3 basophilic leukaemia cells by approximately half. Treating intact cells with the tyrosine kinase inhibitor genistein, an isoflavone found in soy plants and known to disrupt caveolae-related endocytosis, reduced the AEA accumulation by half, but in combination with URB597 no further decrease was seen. Further on, the effects of genistein upon uptake were secondary to inhibition of FAAH. The ability to inhibit the accumulation and metabolism of AEA was shared by several flavonoids (shown in paper III). In paper IV, the isoflavone biochanin A and URB597 had effects in vivo, in a model of persistent pain, effects decreased by the cannabinoid receptor 1 antagonist AM251.

In paper VI, the cellular processing of the endocannabinoid metabolites following degradation was examined, a mechanism poorly understood. It was found that nitric oxide (NO) donors significantly increased the retention of tritium in cell membranes following incubation with either tritiated AEA or 2-AG. Further experiments revealed that the effect of NO donors mainly involves the arachidonate part of the molecules. Inhibition of FAAH completely reduced the effect of NO donors in cells with a large FAAH component, indicating that the effects were downstream of the enzyme.

These results suggest that the cellular processing of endocannabinoids can be affected in a manner of different ways by pharmacological manipulation in vitro and that naturally occurring flavonoid compounds can interact with the endocannabinoid system.

Place, publisher, year, edition, pages
Umeå: Farmakologi och klinisk neurovetenskap, 2009. 80 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1247
Endocannabinoid, anandamide, cellular processing, pain, flavonoids, fatty acid amide hydrolase
National Category
Pharmacology and Toxicology
Research subject
Medical Pharmacology
urn:nbn:se:umu:diva-22221 (URN)978-91-7264-732-9 (ISBN)
Farmakologi, 901 87, Umeå
Public defence
2009-05-22, Sal E04, byggnad 6E, Umeå universitetssjukhus, 901 89 Umeå, Umeå, 09:00 (English)
Available from: 2009-04-30 Created: 2009-04-28 Last updated: 2010-01-18Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Thors, Lina
By organisation
In the same journal
British Journal of Pharmacology

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 57 hits
ReferencesLink to record
Permanent link

Direct link