Characterization of cold sensitivity and thermal preference using an operant orofacial assay

Abstract

Background: A hallmark of many orofacial pain disorders is cold sensitivity, but relative to heat-related pain, mechanisms of cold perception and the development of cold allodynia are not clearly understood. Molecular mediators of cold sensation such as TRPM8 have been recently identified and characterized using in vitro studies. In this study we characterized operant behavior with respect to individually presented cold stimuli (24, 10, 2, and -4 degrees C) and in a thermal preference task where rats chose between -4 and 48 degrees C stimulation. We also evaluated the effects of menthol, a TRPM8 agonist, on operant responses to cold stimulation (24, 10, and -4 degrees C). Male and female rats were trained to drink sweetened milk while pressing their shaved faces against a thermode. This presents a conflict paradigm between milk reward and thermal stimulation.

Results: We demonstrated that the cold stimulus response function was modest compared to heat. There was a significant effect of temperature on facial (stimulus) contacts, the ratio of licking contacts to stimulus contacts, and the stimulus duration/contact ratio. Males and females differed only in their facial contacts at 10 degrees C. In the preference task, males preferred 48 degrees C to -4 degrees C, despite the fact that 48 degrees C and -4 degrees C were equally painful as based on their reward/stimulus and duration/contact ratios. We were able to induce hypersensitivity to cold using menthol at 10 degrees C, but not at 24 or -4 degrees C.

Conclusion: Our results indicate a strong role for an affective component in processing of cold stimuli, more so than for heat, which is in concordance with human psychophysical findings. The induction of allodynia with menthol provides a model for cold allodynia. This study provides the basis for future studies involving orofacial pain and analgesics, and is translatable to the human experience.

Figure 1

Behavioral response to cold stimuli. Four outcome measures are shown: intake (A), facial contact events (B), licks (reward)/facial contact ratio (measures success) (C), and facial duration/contact (D) for cold temperatures (-4, 2, 10, and 24°C), expressed as a percent of baseline (37°C). + indicates significant increase, and * indicates a significant decrease (P < 0.05). (E) represents the change in the licks/facial contact (success ratio) over the 30-minute test period for each of the temperatures tested.

Figure 2

Behavioral response to thermal preference. Rats were first placed in the thermal preference testing box (A) with left and right thermodes set at 37°C to ensure that there was not a side preference (B). In two additional testing sessions, rats chose between -4 and 48°C (C) and their reward/stimulus (D) and stimulus duration/contact ratios were found to be no different for -4°C and 48°C, but significantly lower for -4°C and 48°C relative to 37°C (ratios calculated from temperature choice trials). * indicates significant decrease, P < 0.05.

Figure 3

Behavioral response to menthol and vehicle treatment. Stimulus contacts (A), reward/stimulus success ratio (B), and stimulus duration/contact (C) at 24, 10, and -4°C in the presence of menthol (solid lines) or vehicle (dotted lines), expressed as a percent change from untreated baseline temperatures. Data points indicate average change from baseline, + indicates significant increase, and * indicates a significant decrease. The change in reward/stimulus success ratio over time (D) is also given for menthol (blue), vehicle (orange), baseline at 10°C (green), and baseline at 37°C (pink).