Figure 1.
Comparison of different transgenic mice reveals test-site and stimulus-intensity specific mechanosensory responses.
Nav1.7Nav1.8 mice (blue columns, n = 7), Nav1.7Advill mice (red column, n = 9) and Nav1.7Wnt1 mice (green column, n = 9) mice show normal responses to von Frey hairs applied using either the up-down method (a) or the repeated measures method in comparison to littermate mice (white columns, n = 36). (b). Both Nav1.7Advill mice (n = 9) and Nav1.7Wnt1 mice (n = 9) show a behavioural deficit in response to the abdominal von Frey test in comparison to Nav1.7Nav1.8 mice (n = 7) and littermate mice (n = 36) (c). The abdomens of C57BL/6 (n = 12) mice are significantly more sensitive than the plantar surface of the hindpaw (d), which is loss if the abdomen is shaved (e). Shaving the abdominal hair attenuates the sensitivity to von Frey hair stimulation of Nav1.7Nav1.8 (n = 10) and littermate mice (n = 21) but has no effect of Nav1.7Advill (n = 7) or Nav1.7Wnt1 mice (n = 11) (f). Nav1.7Nav1.8 (n = 14), Nav1.7Advill (n = 8) Nav1.7Wnt1 (n = 9) show a significant increase withdrawal threshold in response to the Randall-Siletto test when applied to the tail but not the paw when compared to littermate (n = 26) mice (g). Nav1.8KO (light blue column, n = 11) and Nav1.9KO (turquoise column, n = 8) but not Nav1.3KO (yellow column, n = 6) show a significant increase withdrawal threshold in response to the Randall-Siletto test when applied to the tail when compared to littermate (n = 27) mice, however no difference is seen when applied to the paw (h). TRPA1 KO mice (pink columns, n = 8) show a behavioural deficit to Randall-Selitto test applied to the paw but not tail in comparison to littermate mice (white columns, n = 8) (i). Data analysed by two-way analysis of variance followed by a Bonferroni post-hoc test. Results are presented as mean ± S.E.M. ** P<0.01 and *** P<0.001 (individual points).
Figure 2.
The DRG innervating the hindpaw and tail consist of different ratios of neuronal subpopulations.
Example section of an L4 (a) and an S1 (b) DRG (N52: green, Nav1.8: red, scale bar = 250 µm). Overall percentage of estimated number of N52, Nav1.8 and double stained cells within L4 (n = 52), L5 (n = 43), L6 (n = 32), S1 (n = 18) and S2 (n = 17) DRG (c). Total estimated number of N52, Nav1.8 and double-stained cells within L4 (n = 52), L5 (n = 43), L6 (n = 32), S1 (n = 18) and S2 (n = 17) DRG (d). All data analysed by two-way analysis of variance followed by a Bonferroni post-hoc test. Results are presented as mean ± S.E.M. ** P<0.01 and *** P<0.001 (individual points).
Figure 3.
Comparison of different transgenic mice reveals stimulus-intensity specific responses to noxious thermal stimuli.
Behavioural responses of different Nav1.7 tissue-specific knockouts to the Hargreaves test applied to the hindpaw. (a) Nav1.7Nav1.8 mice (blue columns, n = 14), Nav1.7Advill mice (red column, n = 7) and Nav1.7Wnt1 mice (green column, n = 12) all show a behavioural deficit in response to the Hargreaves test at a heat ramp of 0.6°C.s−1 in comparison to littermate mice (white columns, n = 27), however only Nav1.7Advill and Nav1.7Wnt1 mice show a behavioural deficit in response to the Hargreaves test at a heat ramp of 2.0°C.s−1. (b) Nav1.8KO mice (light blue column, n = 6) and Nav1.9KO mice (turquoise column, n = 10) but not Nav1.3KO mice (orange column, n = 6) show a significantly increased withdrawal latency to the Hargreaves test at a heat ramp of 0.6°C.s−1 in comparison to littermate mice (white columns, n = 18), however this significant increase is lost the when the Hargreaves test is conducted using a heat ramp of 2.0°C.s−1. Data analysed by two-way analysis of variance followed by a Bonferroni post-hoc test. Results are presented as mean ± S.E.M. * P<0.05 and *** P<0.001 (individual points).
Figure 4.
Comparison of different transgenic mice reveals distinct mechanisms underlie responses to cooling and noxious cold stimuli.
(a) Nav1.7Advill mice (red columns, n = 8) avoid the 0°C test plate to the same extent as littermate controls (white columns, n = 8) in the thermal place preference test but show a behavioural deficit in the noxious cooling range (14–12°C). (b) Nav1.8-DTA mice (black columns, n = 6) avoid the cooling stimuli to the same extent as littermate controls (white columns, n = 6) in the thermal place preference test but show a trend indicating a behavioural deficit in response to 0°C. Data analysed by two-way analysis of variance followed by a Bonferroni post-hoc test. Results are presented as mean ± S.E.M. * P<0.05 (individual points).
Figure 5.
The affect of circadian rhythm on von Frey responses over a 24-hour period.
(a) Behavioural responses of C57BL/6 mice to the von Frey hairs applied to the hindpaw over a 24 h period. Measurements were taken every 4 hours starting at 07:00. (b) Behavioural responses of Nav1.8-DTA mice to the von Frey hairs applied to the hindpaw over a 24 h period. (a) Data analysed by two-way analysis of variance followed by a Bonferroni post-hoc test and (b) t-test. Results are presented as mean ± S.E.M. * P<0.05, *** P<0.001.