↵2-a Forebrain membrane AEA amidohydrolase was prepared as described under “Materials and Methods” and was incubated with DMSO vehicle, DAK or AEA for 1 hr at 37°C in a total volume of 2 ml. After the incubation, each incubate in its entirety was placed into prewetted dialysis tubing (molecular weight cut-off, 12,000–14,000) and dialyzed at 4°C with stirring against TME buffer containing 0.1% BSA for 18 hr. AEA amidohydrolase activity was determined in 1.5 ml of the pretreated dialysate or washed membranes using [¹⁴C]AEA as a substrate (final concentration, 11 μM). Data reported are the percent of the added [¹⁴C]AEA hydrolyzed to [¹⁴C]arachidonic acid during a 30-min incubation at 37°C.

↵2-b Rat brain membranes were prepared as outlined under “Materials and Methods” and were preincubated (final concentration 0.2 mg/ml) with either 1 μM DAK, 1 μM AEA or DMSO for 40 min at 37°C. The membranes were removed from the incubation mixture by centrifugation and were washed six additional times with buffer kept at 37°C. After the final wash, the membranes were incubated with 9 nCi of [¹⁴C]AEA labeled in the ethanolamine portion of the molecule at 37°C for 5 min (final concentration of AEA, 0.2 nM).