Von Frey Test for Mechanical Allodynia

For the assessment of mechanical allodynia, we used Von Frey testing (Shields et al, 2003; Mitsi et al, 2015) with ascending forces expressed in grams (0.1–3.6 g; Electronic von Frey Anesthesiometer; IITC). Each filament was applied five times in a row against the lateral area of the paw. Hindpaw withdrawal or licking induced by the filament was defined as a positive allodynia response. A positive response in three out of five repetitive stimuli was defined as the allodynia threshold. Mice were habituated to the Von Frey apparatus for 30 min every day for 10 days, before the SNI surgeries. For the tolerance studies, mice were injected subcutaneously for 17 consecutive days with 3 mg/kg oxycodone and allodynia was assessed 1 h after the injection.