| Our peripheral sensory nerves detect an extensive array of somatosensory stimuli, including environmental temperature. Among different temperatures, the ability for us to detect and avoid cold temperature is critical for our survival. Previous studies show that cold sensing neurons play important roles in thermosensation, nociception (pain detection), thermoregulation, cool-induced analgesia, hypersensitivity to cold after inflammatory or neuropathic injury, and smooth muscle contraction (reflexes) in both the bladder and airway of the lungs. Psychophysical and electrophysiology data show that cold responses are mediated by both Adelta- and C-fibers which are localized in discrete peripheral receptive zones. Each of which is likely to provide one or more distinct cold sensations ranging from pleasantly cool to painfully aching, prickling, and burning cold.;With this diversity in the neuronal basis of cold sensations, it is remarkable that the majority of cold responses in vivo (at the molecular level) are dependent on the cold- and menthol-receptor, transient receptor potential melastatin 8 (TRPM8), a member of the TRP family of ion channels. TRPM8 is expressed in a subset of primary afferent nerve terminals which are distributed as free nerve endings in the peripheal system (i.e., skin and tooth). Genetic disruption of the Trpm8 gene in animal models lead to severe deficits in cool temperature discrimination, detection in noxious cold temperatures, nocifensive responses to cooling compounds, and injury-evoked hypersensitivity to cold. This suggests that TRPM8 serves as the primary transducer of diverse cold sensations.;In order to determine how TRPM8 plays an important role in such a critical, yet diverse function in cold signaling, we have established a line of bacterial artificial chromosome (BAC) transgenic mice in which expression of enhanced green fluorescent protein (eGFP) is driven by the TRPM8 transcriptional promoter (Trpm8GFP). This Trpm8GFP transgenic mouse enables us to label all the TRPM8-expressing neurons throughout the animal and provides an opportunity to trace not only cell somas but also axons and nerve endigs in both peripheral and central terminations. Based on tracer expression, we show that TRPM8 neurons bear the neurochemical hallmarks of both Adelta- and C-fibers, and presumptive nociceptors and non-nociceptors. Furthermore, TRPM8 axons diffusely innervate the skin and oral cavity, terminating in discrete peripheral zones that mediate distinct perceptions of innocuous cool, noxious cold, and first- and second-pain.;Furthermore, we investigated the time when animals begin to detect cold temperatures. In order to address this question, we generated a developmental time course of TRPM8 expression in dorsal root ganglia (DRG) to define when the expression begins in Trpm8GFP transgenic mice. We also examined when TRPM8 afferent nerves innervate the dorsal horn of the spinal cord and establish adult-like termination patterns. Lastly, in order to determine when the functional synaptic connections are formed developmentally, we looked for c-fos expression in postsynaptic neurons within the terminal zone at different postnatal stages. In addition, previous studies---including our own---reported that these cold sensing neurons are a diverse and heterogeneous cell population which is responsible for a wide arrayof cold perceptions. In our work, we determine when this heterogeneity is established developmentally. We analyzed Trpm8GFP expression in the DRG throughout different embryonic and postnatal stages and correlated TRPM8 expressio to the neurochemical phenotypes of other known markers. Specifically, we have examined Trpm8GFP expression with markers for specific fiber types, trophic factors, and somatosensory and nociceptive signaling. Our results indicated that functional synaptic connections are established by the second postnatal week and that this is alsothe time when heterogeneous phenotypes within TRPM8 neuronal population resemble that of adult mice.;In conclusion, we have established a developmental profile of the neurons that express TRPM8 as a means to understand how distinct cold neura circuits are established and when heterogeneity within this population is generated developmentally. Moreover, the heterogeneity within the TRPM8 neuron population and the fact that TRPM8 neurons terminate in topographically diverse site makes the peripheral cold circuitry cellularly and anatomically complex. Thus, this suggests that cold fibers, caused by the diverse neuronal context of TRPM8 expression, use a single molecular sensor to convey a wide range of cold sensations. (Abstract shortened by UMI.)... |
