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Anandamide degradation and N-acylethanolamines level in wild-type and CB1 cannabinoid receptor knockout mice of
M Maccarrone1, M Attinà, M Bari
1Departments of Experimental Medicine and Biochemical Sciences, University of Rome 'Tor Vergata', Italy. Maccarrone@med.uniroma2.it
Abstract:
CD1 mice lacking the CB1 receptors (knockout, KO) were compared with wild-type littermates for their ability to degrade N-arachidonoylethanolamine (anandamide, AEA) through a membrane transporter (AMT) and a fatty acid amide hydrolase (FAAH). The regional distribution and age-dependence of AMT and FAAH activity were investigated. Anandamide membrane transporter and FAAH increased with age in knockout mice, whereas they showed minor changes in wild-type animals. Remarkably, they were higher in all brain areas of 6-month-old knockout versus wild-type mice, and even higher in 12-month-old animals. The molecular mass (approximately 67 kDa) and isoelectric point (approximately 7.6) of mouse brain FAAH were determined and the FAAH protein content was shown to parallel the enzyme activity. The kinetic constants of AMT and FAAH in the cortex of wild-type and knockout mice at different ages suggested that different amounts of the same proteins were expressed. The cortex and hippocampus of wild-type and knockout mice contained the following N-acylethanolamines: AEA (8% of total), 2-arachidonoylglycerol (5%), N-oleoylethanolamine (20%), N-palmitoylethanolamine (53%) and N-stearoylethanolamine (14%). These compounds were twice as abundant in the hippocampus as in the cortex. Minor differences were observed in AEA or 2-arachidonoylglycerol content in knockout versus wild-type mice, whereas the other compounds were lower in the hippocampus of knockout versus wild-type animals.
Insights
Mice lacking CB1 receptors show increased levels of anandamide transporters and FAAH enzymes with age. These changes are more pronounced in knockout mice, impacting N-acylethanolamine levels in the brain.
Area of Science:
- Neuroscience
- Biochemistry
- Pharmacology
Background:
- The endocannabinoid system, including CB1 receptors, plays a crucial role in regulating various physiological processes.
- N-arachidonoylethanolamine (anandamide, AEA) is a key endocannabinoid degraded by the anandamide membrane transporter (AMT) and fatty acid amide hydrolase (FAAH).
Purpose of the Study:
- To investigate the impact of CB1 receptor deficiency on AEA degradation pathways (AMT and FAAH).
- To examine the age-dependent changes and regional distribution of AMT and FAAH in CB1 receptor knockout (KO) mice compared to wild-type (WT) littermates.
Main Methods:
- Comparative analysis of AMT and FAAH activity and protein content in the brains of CB1 KO and WT mice at different ages.
- Biochemical characterization of mouse brain FAAH, including molecular mass and isoelectric point determination.
- Quantification of various N-acylethanolamines in the cortex and hippocampus of KO and WT mice.
Main Results:
- AMT and FAAH activity increased with age in KO mice, but showed minor changes in WT mice.
- Higher AMT and FAAH levels were observed in all brain areas of KO mice compared to WT mice, with further increases at 12 months of age.
- FAAH protein content paralleled enzyme activity, and kinetic analysis suggested differences in protein expression levels between KO and WT mice.
- While AEA and 2-arachidonoylglycerol levels showed minor differences, other N-acylethanolamines were lower in the hippocampus of KO mice compared to WT mice.
Conclusions:
- CB1 receptor deficiency leads to an age-dependent upregulation of AEA degradation enzymes (AMT and FAAH) in the mouse brain.
- These alterations in AEA metabolism may have implications for endocannabinoid signaling and function in the absence of CB1 receptors.
