2.3. Palmitic acid and drug treatments

Cultured neurons were treated with 0, 50, 100, 200 or 500 μmol/L palmitic acid (PA) (P5585; Sigma‐Aldrich) and 1 mmol/L l‐carnitine (LC) (C0158; Sigma‐Aldrich) for 8 hours to establish the appropriate concentration of PA. Neuronal viability was measured as described below. A 100 μmol/L PA treatment was chosen for further experiments assessing KB production. Briefly, LC was dissolved in PBS (1 mol/L). PA stock solutions of 200 mmol/L were prepared in 100% ethanol. Working solutions of 6 mmol/L PA were generated by incubating the PA in PBS containing 10% endotoxin‐free and fatty acid‐free bovine serum albumin at 37°C for 60 minutes. This solution was then added to the media and incubated for 8 hours before KB measurements.

All drugs were dissolved in DMSO. Neurons were incubated with 10 μmol/L pioglitazone (M3283; Abmole, Houston, TX, USA), 10 μmol/L GW9662 (M2748; Abmole) or 5 μmol/L apoptozole (M7575; Abmole) for 24 hours before further experiments. Neurons treated with an equal amount of DMSO served as controls.