Abstract

Background: 

Chronic stress (CS) is associated with a decrease in pain threshold caused by the changes in neural pain circuits. It can be associated to glucocorticoid imbalance with alterations in neural circuitry. Inhibition of stress-induced pain-related neural changes by using techniques that safely induce neuroplasticity such as transcranial direct current stimulation (tDCS) may prevent hyperalgesia triggered by CS. Objective: 

This study aimed to verify the effect of tDCS performed prior to CS exposure on nociceptive response. Methods: Thirty-two rats were distributed in the following groups: control; stress; sham-tDCS þ stress; and tDCS þ stress. Bicephalic active tDCS was performed for 8 consecutive days before the CS exposure. The pain threshold was evaluated using a hot plate and tail flick latency (TFL) tests.

Results: 

The tDCS exposure increased the pain threshold on stressed rats. Conclusion: The data obtained indicate that the treatment with bicephalic active tDCS before chronic stress exposure prevents stress-induced hyperalgesia.

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 Chronic stress (CS) exposure is associated with alterations in neural pain circuits that may result in hyperalgesia and/or allodynia. It important to note that, in the context of pain, stress exerts paradoxical effects: while acute stress induces analgesia [1], chronic stress classically triggers hyperalgesia [2,3] and allodynia [3,4].

Post-traumatic stress disorder (PTSD) can induce exacerbation of emotional stress symptoms associated with decreased pain threshold [5,6]. Glucocorticoid imbalances and structural/functional alterations in neural circuits that resemble those seen following chronic stress suggest that inappropriate processing of information during stress may be a part of pathological process [7] (Herman, 2013). In humans, tDCS reduces pain related chronic disorders such as fibromyalgia and spinal cord injury [8,9]. Studies in rats performed by our research group showed that tDCS reverses chronic inflammatory pain [10], hyperalgesia induced by chronic stress [3], and neuropathic pain [11]. Its therapeutic effect is based on the modulation of neural activity by small electrical currents that lead to the polarization of the neural tissue. In consequence, a significant change in the resting membrane threshold occurs [12], leading to synaptic plasticity [13]. This study was conducted to examine the effect of tDCS treatment prior to chronic stress exposure by measuring on pain threshold. Material and methods Animals Thirty-two 60-day-old male Wistar rats were randomized for weight and based on pain threshold by using a hot plate basal test (HPT); they were divided into four groups: Control; Stress; ShamtDCS þ Stress, and tDCS þ stress. All procedures were approved by the Institutional Committee for Animal Care and Use (GPPG-HCPA protocol No. 100.381) in accordance with the Guide for the Care and Use of Laboratory Animals 8th edition (2011). The final pain threshold was evaluated using HPT and tail flick latency (TFL) tests [14,15]. The latency for both tests was recorded in seconds (s), and a cutoff of 20 s and 10 s, respectively, was used to avoid tissue damage. Repeated sessions of bicephalic active tDCS were performed for 20 min/day (Sham tDCS was performed for 20 s/day) for 8 days before the CS exposure between 08:00 h and 09:00 h, 1 h/day/5 days/week/11 weeks) [16]. Twenty-four hours after the last stress session, HP test and the TFL test was performed. Animals were killed by decapitation 24 h after the final HP test. Brainderived neurotrophic factor (BDNF) and interleukin (IL)-10 serum levels were determined using Enzyme-linked Immunosorbent assay. Data were analyzed using one-way analysis of variance (ANOVA) followed by Student Neumann Keuls test, and values of p < 0.05 were considered as statistically significant. Results Chronic stress exposure during the HPT (one-way ANOVA, F(3,28) ¼ 17.520; p < 0.001, n ¼ 8) and in the TFL (one-way ANOVA, F(3,28) ¼ 13.553; p < 0.001, n ¼ 8) triggered thermal hyperalgesia (Fig. 1). The tDCS prior to exposure prevented the thermal hyperalgesia observed in the final HP test, but not in final TFL test. There was no difference between the groups in BDNF and IL10 serum levels (one-way ANOVA, F(3,21) ¼ 0.103, n ¼ 6e7; p ¼ 0.958 and one-way ANOVA, F(3,21) ¼ 0.967; p ¼ 0.424, n ¼ 6e7, respectively) (data not shown) (see Table 1). Discussion The results of this study indicate that the treatment using bicephalic tDCS for eight days prior to chronic stress exposure prevents the hyperalgesia induced by CS. This is the first study showing that tDCS prevents thermal hyperalgesia triggered by CS. This effect was established by measuring the nociceptive response during the HP. HP is used to evaluate the changes in latency behaviors (licking or jumping responses) as indicators of modifications in the supraspinal pain process [17] resulting from supraspinal sensory integration [18]. However, tDCS did not affect the nociceptive response during the TFL, suggesting that TFL presents lower sensitivity for detecting the preventive effect of tDCS. Experimental pain is usually mediated by Ad fibers, and it can disappear completely before the onset of pain mediated by C fibers. A thermal model of pain, such as TFL, usually reflects the moment the first pain wave is produced by Ad fibers, while the second pain wave, triggered by C fibers, is not yet detected [15]. The CS exposure induces thermal hyperalgesia by modifying signaling pathways in the primary afferent nociceptors, and this effect is mediated by neuroendocrine release of both glucocorticoids and catecholamines [19]. The activation of stress-related circuitry in the hypothalamus activates pain-facilitating neurons in the rostral ventromedial medulla to produce hyperalgesia [20], suggesting possible changes in brain activity. In addition, a previous study by our group showed that tDCS stimulation was effective in reversing the hyperalgesia/ allodynia induced in a rat model of chronic restraint stress [3]. This constitutes the rationale for testing the anti-hyperalgesic effect of tDCS treatment. The tDCS modulatory effect can be mediated by several neurotransmitter systems, including opioid, adrenergic, substance P, glutamate, and neurokinin receptors [21,22]. It leads to a cascade of events resulting in the modulation of synaptic neural chains that include several thalamic nuclei, the limbic system, brainstem nuclei, and the spinal cord [23]. Thus, we suggest that bicephalic tDCS treatment applied prior to exposure to CS modifies maladaptive neuroplastic processes involved in the etiology of hyperalgesia induced by chronic stress. In conclusion, our findings provide pioneering evidence that hyperalgesia triggered by chronic stress can be prevented by previous treatment by tDCS. This finding opens a path to new pharmacological studies aiming to apply neuromodulation techniques as a tool to decrease the use of pain-reducing drugs. Future studies should be carried out in order to evaluate other molecular biomarkers to elucidate the neuroplastic mechanisms underlying the effect of prior exposure to neuromodulation on prevention of pain responses triggered by chronic stress exposure. 

Conflicts of interest The authors declare that they do not have conflicts of interest associated with the publication of this manuscript. The authors alone are responsible for the content and writing of the paper. Acknowledgements This study was supported by the following Brazilian funding agencies: the National Council for Scientific and Technological Development e CNPq (Torres ILS, Scarabelot VL); the Committee for the Improvement of Higher Education Personnel e CAPES (Spezia Adachi LN); the Graduate Research Group of the Hospital de Clínicas de Porto Alegre e GPPG (I.L.S. Torres, Grant 100.381); MCT/ FINEP e COENG/2013 - CAPES/FAPERGS (Macedo IC).

 [1] Cecconello AL, Torres IL, Oliveira C, Zanini P, Niches G, Ribeiro MF. DHEA administration modulates stress-induced analgesia in rats. Physiol Behav 2016;157:231e6. [2] Torres ILS, Vasconcellos AP, Silveira Cucco SN, Dalmaz C. Effect of repeated stress on novelty-induced antinociception in rats. Braz J Med Biol Res 2001;34:241e4. [3] Adachi LNS, Wolnei W, Laste G, Medeiros LF, Rozisky JR, Souza A, et al. Reversal of chronic stress induced pain by transcranial direct current stimulation (tDCS) in an animal model. Brain Res 2012;1489:17e26. [4] Bardin L, Malfetes N, Newman-Tancredi A, Depoortere R. Chronic restraint stress induces mechanical and cold allodynia,and enhances inflammatory pain in rat: relevance to human stressassociated painful pathologies. Behav Brain Res 2009;205:360e6. [5] Corrigan FM, Fisher JJ, Nutt DJ. Autonomic dysregulation and the Window of Tolerance model of the effects of complex emotional trauma. J Psychopharmacol 2011;25:17e25. [6] Holley AL, Wilson AC, Noel M, Palermo TM. Post-traumatic stress symptoms in children and adolescents with chronic pain: a topical review of the literature and a proposed framework for future research. Eur J Pain 2016;20:1371e83. [7] Herman JP. Neural control of chronic stress adaptation. Front Behav Neurosci 2013;7:61. [8] Fregni F, Boggio PS, Lima MC, Ferreira MJ, Wagner T, Rigonatti SP, et al. A sham-controlled, phase II trial of transcranial direct current stimulation for the treatment of central pain in traumatic spinal cord injury. Pain 2006;122: 197e209. [9] Fregni F, Gimenes R, Valle AC, Ferreira MJ, Rocha RR, Natalle L, et al. A randomized, sham-controlled, proof of principle study of transcranial direct current stimulation for the treatment of pain in fibromyalgia. Arthritis Rheum 2006;54:3988e98. [10] Laste G, Caumo W, Adachi LN, Rozisky JR, de Macedo IC, Filho PR, et al. Aftereffects of consecutive sessions of transcranial direct current stimulation (tDCS) in a rat model of chronic inflammation. Exp Brain Res 2012;221:75e83. [11] Cioato SG, Medeiros LF, Marques-Filho PR, Vercelino R, de Souza A, Scarabelot VL, et al. Long-lasting effect of transcranial direct current stimulation in the reversal of hyperalgesia and cytokine alterations induced by the neuropathic pain model. Brain Stimul 2016;9:209e17. [12] Zaghi S, Acar M, Hultgren B, Boggio PS, Fregni F. Noninvasive brain stimulation with low-intensity electrical currents: putative mechanisms of action for direct and alternating current stimulation. Neuroscientist 2010;16:285e307. [13] Fritsch B, Reis J, Martinowich K, Schambra HM, Ji Y, Cohen LG, et al. Direct current stimulation promotes BDNF-dependent synaptic plasticity: potential implications for motor learning. Neuron 2010;66:198e204. [14] Woolfe G, MacDonald AD. The evaluation of the analgesic action of pethidine hydrochloride (Demerol). J Pharmacol Exp Ther 1944;80:300e7. [15] Le Bars D, Gozariu M, Cadden SW. Animal models of nociception. Pharmacol Rev 2001;53:597e652. [16] Ely DR, Dapper V, Marasca J, Correa JB, Gamaro GD, Xavier MH, et al. Effect of ^ restraint stress on feeding behavior of rats. Physiol Behav 1997;61:395e8. [17] Ossipov MH, Kovelowski CJ, Nichols ML, Hruby VJ, Porreca F. Characterization of supraspinal antinociceptive actions of opioid delta agonists in the rat. Pain 1995;62:287e93. [18] Rubinstein M, Mogil JS, Japon M, Chan EC, Allen RG, Low MJ. Absence of opioid stress-induced analgesia in mice lacking beta-endorphin by site-directed mutagenesis. Proc Natl Acad Sci 1996;1996(93):3995e4000. [19] Khasar SG, Burkham J, Dina OA, Brown AS, Bogen O. Stress induces a switch of intracellular signaling in sensory neurons in a model of generalized pain. J Neurosci 2008;28:5721e30. [20] Martenson ME, Cetas JS, Heinricher MM. A possible neural basis for stressinduced hyperalgesia. Pain 2009;142:236e44. [21] Morgan PJ, Barrett P, Howell HE, Helliwell R. Melatonin receptors: localization, molecular pharmacology and physiological significance. Neurochem Int 1994;24:101e46. [22] Wu FS, Yang YC, Tsai JJ. Noncompetitive inhibition of the glycine receptormediated current by melatonin in cultured neurons. Brain Res 2000;881: 208e11. [23] Lima MC, Fregni F. Motor cortex stimulation for chronic pain: systematic review and meta-analysis of the literature. Neurology 2008;70:2329e37.