Recent advances in cancer chemotherapy have significantly increased the survival rate and time of cancer patients, although the use of these drugs is also associated with an increase in morbidity related to the development of painful peripheral neuropathy [18–20]. Oxaliplatin, a third generation platinum derivative, is one of the most effective chemotherapeutic drugs used to treat advanced colorectal cancer [21–25]. Unfortunately, oxaliplatin therapy is associated with significant side effects such as neurotoxicity, which is one of the most prevalent and dose-limiting effects [26–28], occurs in greater than 65% of patients and includes mechanical allodynia and hypersensitivity to cold [29–31]. Oxaliplatin-induced peripheral neuropathy (OIPN) occurs in two forms, acute and chronic. The onset of acute OIPN occurs either during or within 1-2 days after drug infusion, resolves between cycles and recurs with subsequent infusions [25, 27]. Chronic OIPN develops as a cumulative, dose-dependent effect [22, 27, 32, 33] in patients that receive more than 540 mg/m2, regardless of the dosing regimen . The symptoms of chronic OIPN can be debilitating, persistent and respond poorly to currently available therapeutics. Thus, it is critical that we gain a better understanding of the mechanisms underlying the development and persistence of chronic OIPN, which could lead to the development of new treatment modalities to improve symptom management.
The majority of studies examining the neurotoxic effects of oxaliplatin treatment have been done after treatment in the rat [7, 11, 13, 34], while the majority of studies examining the antineoplastic efficacy of oxaliplatin have been done in mouse. Recently, though, several studies have been done to examine oxaliplatin-induced neurotoxicity in the mouse [14, 15]. However, the mouse studies have not closely examined the neurotoxic effects of chronic oxaliplatin treatment on the status of peripheral nerves and spinal dorsal horn neuronal activity. In this study, we used a mouse model of oxaliplatin-induced neurotoxicity, based on our preliminary studies in the rat [7, 35], to investigate the effects of chronic oxaliplatin treatment on nocifensive behavior, peripheral nerve function and wide dynamic range neuron activity in the spinal dorsal horn. The dose of oxaliplatin and the administration schedule were chosen based on previous work in the rat  and preliminary pilot studies in the mouse. The mice received 3.5 mg/kg/iv twice weekly for four weeks, which is equivalent to 130 mg/m2 per administration and a cumulative dose of 1080 mg/m2 after 8 cycles. This dose of oxaliplatin and number of cycles has been found to cause neuropathy symptoms in patients [23, 33, 36]. The oxaliplatin treatment did not affect the general health and functioning of the mice, as assessed by appearance and activity, though the mice did lose weight during the course of the study. Although the weight loss was significant, it was tolerable and consistent with similar rat models . Moreover, the association between the extent of weight loss and peripheral neuropathy has been ruled out, at least in the rat . However, oxaliplatin treatment did produce significant mechanical and cold allodynia that persisted for the four week duration of the study, similar to symptoms reported by patients [1, 9] and to what has been found in previous studies in the rat [12, 13, 11, 38, 39] and mouse [14, 15, 40]. Thus, this mouse model of oxaliplatin-induced neurotoxicity is representative of the human clinical condition and can be a useful tool to study the mechanisms that underlie the development and persistence of chronic OIPN.
Oxaliplatin-induced neuropathy has been associated with damage to the peripheral sensory neurons [7, 41], which leads to alterations in peripheral nerve function [31, 42]. In this study, we found that chronic oxaliplatin treatment induced a decrease in conduction velocities in the caudal and digital nerves that was associated with a concomitant decrease in caudal action potential amplitude. These results are similar to the findings of studies that were done in the rat [7, 43]. Nucleolar morphology changes were also seen in this study that are similar to those shown previously . The mechanisms of these decreases remain unclear and require further study; however, as suggested by Jamieson and colleagues, one possibility is that oxaliplatin induces a decrease in phosphorylated neurofilaments in DRG neurons with a concomitant alterations in sensory axons . Following the finding that oxaliplatin induced a decrease in NCV and action potential amplitude; we examined the morphology of DRG neurons and sciatic nerve at the light and electron microscope levels. Our findings that the diameter of DRG cell bodies is reduced after chronic oxaliplatin treatment is also similar to findings in the rat [7, 34, 43, 45, 46], which are suggestive of neuronal atrophy that could be related to a decrease in phosphorylated neurofilament . Finally, our morphological examination of DRGs and sciatic nerves revealed that many of the neurons were multinucleolated, that the nucleoli were eccentric and that rare nerve fibers presented mild signs of axonopathy. The morphometrical analysis showed the presence of somatic and nucleolar atrophy of DRG neurons. Altered DRG neuron morphology and morphometry have also been found in the rat following oxaliplatin treatment [44, 47]. The underlying mechanisms of this phenomenon are unclear; although one study found that paclitaxel treatment induces nucleolar enlargement that can inhibit the neurotoxic effects of oxaliplatin .
Despite these decreases in NCV, the presence of mechanical and cold allodynia suggests an increase in peripheral nerve sensitivity [41, 48]. The exact mechanism of oxaliplatin-induced hyperexcitability is unclear. One theory is that a metabolite of oxaliplatin, oxalate, may alter the functional properties of voltage-gated sodium channels, which are intrinsic to action potential generation, resulting in a prolonged open state of the channels and hyperexcitability of sensory neurons [49–51]. One possible mechanism underlying the disruption of voltage-gated sodium channel function is the calcium chelating effect of oxalate, which inhibits intracellular calcium-dependent mechanisms [13, 50]. A second theory to explain oxaliplatin-induced hyperexcitability, with regard to cold allodynia, is that the sensitivity and expression levels of transient receptor potential melastatin 8 (TRPM8) and transient receptor potential ankyrin 1 (TRPA1) are increased after oxaliplatin treatment [15, 52]. TRPM8 is only expressed in the DRG and responds to innocuous cool and noxious cold (<15C°) temperatures [53–55]. Anand et al.  found that oxaliplatin treatment increased the icilin response in cultured DRG neurons, suggesting that both the TRPM8 and TRPA1 channels are affected and Gauchan et al.  showed that blocking TRPM8 function by administering capsazepine inhibited the induction of cold allodynia after oxaliplatin treatment.
Given that the mice developed mechanical and cold allodynia after oxaliplatin treatment, our final area of inquiry was to examine the effects of oxaliplatin on the activity of wide dynamic range neurons in the spinal dorsal horn. A variety of neuropathic pain models exhibit increased evoked wide dynamic range (WDR) neuron activity after nerve injury [56–61]. In this study, mice that received chronic oxaliplatin treatment had a significant increase in wide dynamic range neuron activity compared to naive mice. Our findings are similar to those seen in other drug-induced neuropathy models [56, 62] and may reflect physiological changes in the SDH as well as in primary afferents [58, 63]. The oxaliplatin-induced increase in WDR neuron activity may be due to increased neurotransmitter release in the SDH [64–69], resulting in altered synaptic transmission [68, 70] due to increased neurotransmitter levels. Increased levels of neurotransmitters, such as brain-derived neurotrophic factor (BDNF) can activate calcium permeable 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl) propanoic acid (AMPA) channels, which increase excitatory synaptic activity and miniature excitatory post-synaptic currents (mEPSCs) in the SDH [68, 70, 71]. Further, increased neurotransmitter levels in the SDH can lead to a decreased frequency of inhibitory synaptic events, thus allowing for more excitatory activity . The increase in WDR neuron activity is likely due to interactions between activated receptors and other signaling molecules [73–76], which results in longer lasting changes in SDH neurons such as what occurs in central sensitization .
It is well-documented that chemotherapeutic agents are toxic to the peripheral nervous system, though their mechanisms of action differ by class. However, many of the physiological mechanisms that underlie the development of chemotherapy-induced neurotoxicity remain unclear, regardless of the class of the drug. The platinum compounds, such as oxaliplatin, target the cell bodies of the neurons located in the DRG and disrupt the function of DNA [7, 77]. In addition to the mechanisms discussed above, oxaliplatin may induce neurotoxicity by increasing p53 and p38 activity [78, 79]. The taxanes [80, 81], vinca alkaloids [82, 83] and epothilones  all seem to exert their neurotoxic effects by inhibiting tubulin function and disrupting axonal transport and intracellular signaling processes. The symptoms associated with peripheral neurotoxicity vary by drug class but all have a significant deleterious effect on the quality of life of cancer patients that can become dose-limiting. Thus, it is important that research continues to increase the understanding of the mechanisms underlying the development of chemotherapy-induced neurotoxicity.