Mechanical hypersensitivity
The development of mechanical allodynia of the hind paw ipsilateral to the MIA-injected knee (indicating referred pain) was assessed by measuring paw withdrawal thresholds (PWT, expressed in grams) using an electronic von Frey anesthesiometer (IITC Life Science, Woodland Hills, CA). Animals were placed in individual plastic cages with a metal mesh floor which allowed access to the plantar surface of each hind paw. After the habituation period, a semi-flexible plastic von Frey filament was applied on the plantar surface of the ipsilateral hind paw with increasing force until a paw withdrawal response was elicited. The force at which this reaction occurs is automatically recorded by the device. Six to eight PWT measurements were taken at each time point and the mean value was used for further calculations.