Dose-dependent effects of icilin on thermal preference in the hindpaw and face of rats

Abstract

Icilin induces wet dog shakes (WDS) in rodents when injected systemically and activates the cold receptor TRPM8 and putative cold receptor TRPA1. It is assumed that WDS reflect an enhanced cold sensitivity; however, none have examined the relationship between WDS and cutaneous cold sensitivity following systemic icilin. In this study, we sought to characterize the effect of systemic and central icilin administration on WDS and thermal preference with either hindpaw or facial stimulation. It was found that a low dose of icilin (.025 mg), which transiently elevated WDS, decreased preference for cold with hindpaw stimulation (15 and 45 degrees C) when administered ip or it. Intracisternal administration of this dose produced similar results for facial stimulation (10 and 48 degrees C), but had no effect when administered ip. In contrast a high dose of icilin (.25 mg), which persistently elevated WDS, strongly increased preference for cold with hindpaw stimulation and had no effect on thermal preference with facial stimulation. These findings indicate that at the low concentration, systemic and central icilin enhances cold sensitivity, likely via TRPM8 and TRPA1 activation. In contrast, systemic icilin at the high concentration produces peripheral and/or central effects that diminish cold sensitivity, while WDS is maintained at a persistent rate.

Perspective: Icilin is a unique compound that produces dissociable effects on an innate behavior (WDS) and on operant behaviors related to thermal perception. This compound could help clarify the relationship between peripheral cold transduction and the central induction of thermogenesis and nocifensive behaviors, as well as alterations that produce pathological pain.

Figure 1

Illustration of intracisternal (i.c.m.) injection. Picture shows the flow of liquid, depicted in white, delivered by blind intracisternal injection (photo courtesy of Dr. Andrew Mannes, NIDCR, and the NIH Mouse Imaging Facility (MIF), NIH, Bethesda, MD). White arrow indicates needle path.

Figure 2

Effects of two doses of icilin and DMSO on thermal preference with hindpaw stimulation. For each graph, the cumulative difference in occupancy duration for the two compartments (hot - cold) is shown over successive response pairs (error bars = one standard deviation). Over 25 baseline sessions, the animals spent more time on the cold side during each response pair, resulting in a cumulative cold preference of 150-200 sec. in 900 sec. trials. DMSO had no significant effect on thermal preference compared to baseline (F₁₄ = 0.71, p = 0.75). (A) There was a delayed but significant reversal toward heat preference following 0.025 mg i.p. icilin compared to i.p. DMSO (F₁₄ = 8.23, p < 0.001) and compared to baseline (F₁₄ = 8.49, p < 0.001). In contrast, cold preference was significantly increased following 0.25mg i.p. icilin compared to DMSO (F₁₄ = 3.46, p < 0.001). (B) Similar to i.p. administration, i.t. administration of 0.025mg icilin progressively attenuated cold preference compared to baseline (F₁₄ = 3.27, p < 0.001). Significant interactions determined by repeated-measures ANOVA.

Figure 3

Effect of two doses of icilin and DMSO (i.c.m.) on thermal preference with facial stimulation at 10 and 48°C. The number of licks achieved in contact with either the 10°C thermode (white bars) or the 48°C thermode (black bars) are shown for baseline (n = 12) and following DMSO (n = 6), 0.025mg (n = 8), or 0.25mg icilin (n = 6). There was a significant effect of treatment on licking at 10°C (Kruskal-Wallis test, chi squared = 16.55, p = 0.001) and 48°C (chi squared = 20.93, p < 0.001). Post-hoc comparisons between treatments were made using the Mann-Whitney U test. * indicates a significant effect of low dose icilin treatment as compared to all other treatments (baseline, DMSO, and high dose icilin). + indicates a significant difference between DMSO and baseline only. For all statistical tests p < 0.05. Data are represented as mean ± SEM.