Studies are conducted in adult male and female Sprague Dawley rats (Envigo) in an AAALAC-accredited facility with approval from an Institutional Animal Care and Use Committee, and implemented in accordance with the Guide for the Care and Use of Laboratory Animals 8th Edition (National Research Council, 2011) with standards set by the National Institutes of Health.

Experimental procedure

Evaluation of cutaneous sensitivity of the hind paw to a mechanical stimulation using von Frey filaments in rodents is considered a surrogate for tactile allodynia seen in patients and continues to be a standard in the preclinical pain field. Several different methods have been implemented to measure tactile allodynia, one of the most common being the “up-down method” first described by Chaplan et al. in 1994. This technique aims to determine the 50% withdrawal threshold (i.e., stimulus that elicits a response 50% of the time that it is applied).

Semmes-Weinstein von Frey filaments (Stoelting, Wood Dale, IL, USA) with handle numbers 3.61, 3.84, 4.08, 4.31, 4.56, 4.74, 4.93, and 5.18 are used to measure mechanical allodynia in rats acclimated to individual chambers placed on a wire mesh floor. When the rat’s paw is flat on the mesh floor, the tip of the filament is applied perpendicularly to the plantar surface of the paw with enough force to cause a slight buckling against the paw. Testing starts with the 4.31 handle number filament, which is held until a positive response is noted (withdrawal) or for a maximum of 6-8 seconds. The force of the next filament depends on the response elicited by the previous one according to the up-down method (Dixon, 1980). The 50% withdrawal threshold is determined by counting 6 critical responses when the response threshold is first crossed (positive to negative or negative to positive) followed by 4 additional stimuli applications (up or down). The number of actual responses collected using this paradigm can vary from a minimum of 4 to a maximum of 9. This entire series of responses is recorded using the convention X = positive response; withdrawal and O = negative response; no withdrawal. The 50% withdrawal threshold (g) is calculated according to the formula provided in Chaplan et al., 1994.


Chaplan SR, Bach FW, Pogrel JW, Chung JM, and Yaksh TL (1994). Quantitative assessment of tactile allodynia in the rat paw. J Neurosci Methods, 53 (1): 55-63. PMID: 7990513 DOI: 10.1016/0165-0270(94)90144-9

Dixon WJ (1980). Efficient analysis of experimental observations. Ann Rev Pharmacol Toxicol, 20: 441-462. PMID: 7387124 DOI: 10.1146/annurev.pa.20.040180.002301

National Research Council (2011). Guide for the Care and Use of Laboratory Animals: Eighth Edition. Washington, DC: The National Academies Press

This work was conducted by PsychoGenics Inc. (Paramus, NJ) in collaboration with PSPP, NINDS, NIH under contract # 75N95019D00026