Mouse model of touch-evoked itch (alloknesis)

Abstract

Lightly touching normal skin near a site of itch can elicit itch sensation, a phenomenon known as alloknesis. To investigate the neural mechanisms of alloknesis, we have developed an animal model. Low-threshold mechanical stimulation of the skin normally does not elicit any response in naive C57/BL6 mice. Following acute intradermal (i.d.) injection of histamine in the rostral back, mechanical stimulation 7 mm from the injection site elicited discrete hindlimb scratch bouts directed toward the stimulus. This began at 10 minutes and peaked 20-40 minutes post histamine injection, declining over the next hour. Histamine itself elicited bouts of scratching not associated with the mechanical stimulus, which ceased after 30 minutes. Histamine- and touch-evoked scratching was inhibited by the μ-opiate antagonist naltrexone. Touch-evoked scratching was observed following i.d. 5-HT (5-hydroxytryptamine), a protease-activated receptor (PAR)-4 agonist, and an MrgprC11 agonist BAM8-22, but not chloroquine or a PAR-2 agonist. The histamine H1 receptor antagonist terfenadine prevented scratching and alloknesis evoked by histamine, but not 5-HT, a PAR-4 agonist or an MrgprC11 agonist. In mice with experimental dry skin, there was a time-dependent increase in spontaneous and touch-evoked scratching. This animal model appears to be useful to investigate neural mechanisms of itch and alloknesis.

Fig. 1

Time course of scratching and alloknesis elicited by histamine. Bars: mean number of scratch bouts (right-hand y-axis) following histamine (□) or vehicle (■). Line graphs: mean alloknesis score (left-hand y-axis) following histamine ( ) or vehicle ( ). Error bars are S.E.M. (n = 6/ group). Following intradermal injection of histamine (271 nmol/10 μl; open bars and ) or saline (vehicle; filled bars and ) in the nape of the neck, the mouse was placed into the recording arena and videotaped for 30 min. At 5-min intervals, an innocuous von Frey filament (bending force 0.7 mN) was applied 3 times at separate sites oriented radially at a distance of 7 mm from the histamine injection site, and the presence or absence of a hindlimb scratch bout following each stimulus was noted (maximum alloknesis score = 3). The number of histamine- or vehicle-evoked hindlimb scratch bouts directed to the injection site (not associated with the von Frey stimulus) was counted after the experiment by reviewing the videotape.

Fig. 2

Effects of naltrexone on alloknesis and scratch evoked by histamine. A) Bar graph of the mean sum of alloknesis score over 30 to 60 min after histamine injection. The striped and black bars show, respectively, the sum of the alloknesis score evoked by saline and histamine (271 nmol/10 μl). The white and grey bars show the sum of score for alloknesis evoked by histamine when preceded by subcutaneous saline or by naltrexone (1 mg/kg), respectively. B) Same as in A for mean scratch bouts/ 30 min. Error bars are S.E.M. * p < 0.05, significant difference from saline group (n = 6/group). # p < 0.05, significant difference from histamine + saline group (n = 6/group).

Fig. 3

Scratching and alloknesis elicited by different pruritogens. A. Dose-response curve for scratch bouts (assessed over 30 min) elicited by id injection of pruritogens indicated in the legend. Error bars: SEM (n=6/group). B. Dose-response curve for alloknesis score elicited by the same pruritogens. C: time course of alloknesis for histamine (271 nmol/10 μl), PAR-2 agonist SLIGRL-NH2 (76 nmol/10 μl), PAR-4 agonist AYPGKF-NH2 (146 nmol/10 μl), and saline vehicle. D: time course of alloknesis for 5-HT (47 nmol/10 μl), chloroquine (193 nmol/10 μl), and BAM8-22 (50 nmol/10 μl).

Fig. 4

Effects of terfenadine on alloknesis and scratching evoked by four pruritogens. A. Bar graph of the mean alloknesis score during the time period 30–60 min after injection of the following: histamine (271 nmol/10 μl), 5-HT (47 nmol/10 μl), PAR-4 agonist AYPGKF-NH2 (146 nmol/10 μl), BAM8-22 (50 nmol/10 μl). Graph shows the alloknesis scores evoked by each pruritogen without (open bars) or following pretreatment with terfenadine (filled bars, 30 mg/kg), respectively. B. As in A for mean scratch bouts/ 30 min. Error bars are S.E.M. * p < 0.05, significant difference from control group (n = 6/group).

Fig. 5

Time-dependent changes in scratch bouts, TEWL and alloknesis score in dry skin mice. The dry skin was developed by treating with AEW twice a day for 12 days. A) TEWL was mesured on the day 0, 1, 3, 5, 8, 10 and 12. Black circle and white square show, respectively, AEW-treated and W-treated groups. B) As in A for Spontaneous scratching. C) As in A for Alloknesis score. Error bars are S.E.M. * p < 0.05, significant difference from day 0.