Pain is ‘‘an unpleasant sensory and emotional experience associated with actual or potential tissue damage with a huge personal and economic impact, affecting more than 30% of people worldwide (Raja et al. 2020; Cohen et al. 2021). Frostbite is a cold, freezing thermal injury caused when tissues are exposed to temperatures below their freezing point leading to extensive digits or limb loss. These severe frostbite conditions are incurred mostly by high altitude mountaineers, soldiers both on & off the battlefield and homeless persons in extreme cold weather environments (Agrawal et al. 2020). Previous reports show that the United States armed forces had the highest incidence of cold injury in 2018–2019 (36.5 per 100,000 person-per year) whereas 149 British army personnel were found to be affected in 2002–2010, which indicate the severity of this debilitating disorder (Kuht et al. 2019; Gupta et al. 2021). It mainly affects the fingers/hands, toes/ feet, nose, cheeks, and ears because they are most vulnerable organs to the external environment. Dark skin coloration develops in the affected area, along with redness, warmth, hypersensitivity, chronic pain, and other complications (Persitz et al. 2022; Tarazi and Bitterman 2023).
Somatosensory neurons are pseudo-unipolar cells that sense alterations in external temperature are located in the dorsal root ganglia (DRG), alongside the spinal cord (Buijs and McNaughton 2020). The pain from cold temperatures starts with the conversion of thermal stimuli into electrical signals by molecular transducers in the plasma membrane of primary sensory nerve terminals. Some of the low and high threshold cold-sensitive nerve fibers detect noxious and innocuous cold temperatures (Lolignier et al. 2016). Neuroinflammation critically regulates the development and maintenance of chronic pain by generating specific inputs from dorsal root ganglion (DRG) which moves towards spinal cord and brain, ultimately leading to central sensitization (Beggs and Salter 2007; Zhang et al. 2008). Previous reports uncertain the role of microglial cells after peripheral nerve injury where multiple proinflammatory mediators causes the neuronal and nociceptors sensitization (Ji et al. 2014). IBA1 is a microglial marker which increases prominently during chronic pain condition and promotes the generation of different inflammatory markers such as TNF-α, IL-1β and IL-6 (Dayanand and Olivia E. Atherton1, Jennifer L. Tackett2, Emilio Ferrer1 2018). ICAM1 also contributes in this immunogenic and inflammatory responses leading to persistent pain development (Sagar et al. 2011; Peng et al. 2016). Microgliosis is the hallmark feature in all of the previously reported neuropathic pain models which causes the sensitization of different nociceptors such as TRPM8, TRPA1 and TRPV1 (Beggs and Salter 2006, 2007; Calvo and Bennett 2012). The intensity and duration of the stimuli are combined to form action potentials by voltage-gated ion channels where these sensitized nociceptors regulates mechanical and thermal pain hypersensitivity (Ahlgren et al. 1996). Among TRPs channels TRPM8 is mainly expressed in small diameter sensory neurons, C fiber thermo receptors, which terminate centrally in layer 1 of the spinal cord dorsal horn and peripherally in the epidermis where they sense altered temperatures (Ran et al. 2016). TRPA1 is a TRPM8-independent cold transducer, which is directly activated by cooling below 10oC and controls cold pain (MacDonald et al. 2020). Along with TRP channels and microgliosis NMDA receptors are upregulated during chronic pain condition regulates and contributes significantly in exacerbating the painful situation (Zhuo 2002; Petrenko et al. 2003; Chazot 2004). This highlights the involvement of these targets in development and progression of chronic pain during frostbite condition.
Unavailability of suitable preclinical model mimicking the actual frostbite induced chronic pain conditions has affected the drug discovery process to huge extent as these models are an important part of research because they help to understand the pathophysiology of a disease and provide excellent druggable targets. So, we have developed a novel model for frostbite induced chronic pain using deep frozen magnets showing different types of validity (face, predictive and constructive) which are part of standard paradigm. Further with the help of behavioral studies, we have showed the time course of mechanical, thermal and cold pain progression. Finally, molecular analysis unveiled the role of different oxido-nitrosative stress markers, microglial cells, proinflammatory cytokines, nociceptors and neuropeptides in progression and maintenance of frostbite induced pain. This novel in vivo rat model may allow quantification of pain parameters like allodynia and hyperalgesia in frostbite induced chronic pain and has the potential of being utilized to screen future treatment modalities.