Thrb expression in whole cell lysates from LGs of rats of both groups was determined by Western blot analysis. After homogenization, protein was quantitated by the biuret dye test. Samples were treated with Laemmli buffer, and equal amounts of protein per sample (200 μg) were subjected to SDS-PAGE (10% Tris-acrylamide) in a miniature laboratory gel apparatus (Miniprotean; Bio-Rad Laboratories, Richmond, CA), in parallel with prestained protein standards and β-mercaptoethanol (Bio-Rad, Hercules, CA). Proteins were then electrotransferred from the gel to an enhanced chemiluminescence (ECL) nitrocellulose membrane (Hybond; Amersham, Buckinghamshire, UK) for 2 hours at 120 V in a miniature transfer apparatus (Miniprotean; Bio-Rad Laboratories). After blocking, the membranes were incubated overnight using rabbit polyclonal anti–Thrb or anti–tubulin (a cytoskeleton protein) antibodies (Santa Cruz Biotechnology, Santa Cruz, CA) at a concentration of 0.4 μg/μL in a buffer containing 3% bovine serum albumin (BSA) and were washed three times as described. Blots were then incubated with immunoperoxidase and developed by DAB (Amersham, Buckinghamshire, UK). Membranes were scanned, converted to digital files, and analyzed (Scion Image Analysis Software; Scion Corp., Frederick, MD).