Fatty acid amide hydrolase (FAAH) regulates hypercapnia/ischemia‐induced increases in n‐acylethanolamines in mouse brain

Abstract

N-acylethanolamines (NAEs) are endogenous lipid ligands for several receptors including cannabinoid receptors and peroxisome proliferator-activated receptor-alpha (PPAR-α), which regulate numerous physiological functions. Fatty acid amide hydrolase (FAAH) is largely responsible for the degradation of NAEs. However, at high concentrations of ethanolamines and unesterified fatty acids, FAAH can also catalyze the reverse reaction, producing NAEs. Several brain insults such as ischemia and hypoxia increase brain unesterified fatty acids. Because FAAH can catalyze the synthesis of NAE, we aimed to test whether FAAH was necessary for CO -induced hypercapnia/ischemia increases in NAE. To test this, we examined levels of NAEs, 1- and 2-arachidonoylglycerols as well as their corresponding fatty acid precursors in wild-type and mice lacking FAAH (FAAH-KO) with three Kill methods: (i) head-focused, high-energy microwave irradiation (microwave), (ii) 5 min CO followed by microwave irradiation (CO + microwave), and (iii) 5 min CO only (CO ). Both CO -induced groups increased, to a similar extent, brain levels of unesterified oleic, arachidonic, and docosahexaenoic acid and 1- and 2-arachidonoylglycerols compared to the microwave group in both wild-type and FAAH-KO mice. Oleoylethanolamide (OEA), arachidonoylethanolamide (AEA), and docosahexaenoylethanolamide (DHEA) levels were about 8-, 7-, and 2.5-fold higher, respectively, in the FAAH-KO mice compared with the wild-type mice. Interestingly, the concentrations of OEA, AEA, and DHEA increased 2.5- to 4-fold in response to both CO -induced groups in wild-type mice, but DHEA increased only in the CO group in FAAH-KO mice. Our study demonstrates that FAAH is necessary for CO - induced increases in OEA and AEA but not DHEA. Targeting brain FAAH could impair the production of NAEs in response to brain injuries.

Tetracosahexaenoylethanolamide, a novel N-acylethanolamide, is elevated in ischemia and increases neuronal output

“…were increased in mouse brain during periods of ischemia/ hypercapnia (28), and that this elevation was exacerbated in FAAH-KO mice. Furthermore, DHEA was increased in ischemic/hypercapnic wild-type and FAAH-KO mice, with FAAH-KO mice exhibiting up to four times higher levels of DHEA in the brain compared with wild-type mice (28).…”

“…MS/MS was performed with a SCIEX QTrap5500 mass spectrometer (SCIEX, Framingham, MA) with an Agilent 1290 HPLC system (Agilent Technologies, Santa Clara, CA) equipped with a Phenomenex Kinetex XB-C18 column, 50.9 × 4.6 mm, 2.6 µm (Phenomenex) and set to a flow rate of 600 l/min (28). The ion mass transitions used were as follows: DHEA (m/z 372.3/62.0), DHEA-d4 (m/z 376.3/ 66.0), THEA (m/z 400.3/62).…”

“…FAAH-KO mice were obtained from Dr. Ben Cravatt, Department of Chemistry at Scripps Research Institute (La Jolla, CA). Brain samples were obtained from FAAH-KO and C57BL/6J wild-type mice as secondary analysis of our previously published study (28). Mice were housed in groups of four per cage with a 12:12 h light:dark cycle in a temperature-controlled (21°C) and humiditycontrolled (40%) environment, with ad libitum access to standard chow and water.…”

“…Total lipid extracts (TLEs) were obtained from frozen powdered whole brain samples according to a modified Folch method (30) and similarly to previous studies performed in our laboratory (28). Briefly, lipids were extracted with 2:1 chloroform:methanol containing a known amount of 2 H 8 (deuterated-6-8, D8)-arachidonic acid (20:4n-6) (Cambridge Isotope Laboratories, Inc., Tewksbury, MA) as an internal standard to determine THA concentration.…”

“…Total THA and unesterified THA were analyzed on an Agilent 5977A quadrupole mass spectrometer coupled to an Agilent 7890B gas chromatograph (Agilent Technologies, Mississauga, ON, Canada) in negative chemical ionization mode. The details of the GC-MS set up were described previously (28,32). In this study, THA was analyzed in selected ion monitoring mode using M-1 for parent ion identification with ion dwell times of 500 ms.…”