Involvement of TRPM2 and TRPV1 channels on hyperalgesia, apoptosis and oxidative stress in rat fibromyalgia model: Protective role of selenium

Abstract

Fibromyalgia (FM) results in pain characterized by low selenium (Se) levels, excessive Ca²⁺ influx, reactive oxygen species (ROS) production, and acidic pH. TRPM2 and TRPV1 are activated by ROS and acid; nevertheless, their roles have not been elucidated in FM. Therefore, we investigated the contribution of TRPM2 and TRPV1 to pain, oxidative stress, and apoptosis in a rat model of FM and the therapeutic potential of Se. Thirty-six rats were divided into four groups: control, Se, FM, and FM + Se. The Se treatment reduced the FM-induced increase in TRPM2 and TRPV1 currents, pain intensity, intracellular free Ca²⁺, ROS, and mitochondrial membrane depolarization in the sciatic (SciN) and dorsal root ganglion (DRGN) neurons. Furthermore, Se treatment attenuated the FM-induced decrease in cell viability in the DRGN and SciN, glutathione peroxidase, and reduced glutathione and α-tocopherol values in the DRGN, SciN, brain, muscle, and plasma; however, lipid peroxidation levels were decreased. Se also attenuated PARP1, caspase 3, and 9 expressions in the SciN, DRGN, and muscle. In conclusion, Se treatment decreased the FM-induced increase in hyperalgesia, ROS, apoptosis, and Ca²⁺ entry through TRPM2 and TRPV1 in the SciN and DRGN. Our findings may be relevant to the elucidation and treatment of FM.

Figure 1

Effect of selenium (Se) treatment on paw withdraw threshold of Hot Plate Test (a) and paw withdrawal force of Von Frey test (b) in rats with FM. (n = 8–10 and mean ± SD). (^a p ≤ 0.001 versus basal groups. ^b p ≤ 0.001 5^th day groups. ^c p ≤ 0.001 versus control group of 19^th day group. ^d p ≤ 0.001 versus Se group of 19^th day group. ^e p ≤ 0.001 versus FM group of 19^th day group).

Figure 2

Effect of selenium (Se) treatment on [Ca²⁺]_i concentration through block of TRPV1 gate in dorsal root ganglion neuron (DRGN) (a) and sciatic nerve (SciN) (b) of control and FM-induced rats. (n = 8–10 and mean ± SD). The animals received intraperitoneal Se for 2 weeks after FM induction. Then, these dissected neurons of control and FM groups were further in vitro treated with CAPS (10 μM) and CPZ (0.1 mM) before loading Fura-2 for 120 seconds. (^a p ≤ 0.05 and ^b p ≤ 0.001 versus control. ^c p ≤ 0.001 and ^d p ≤ 0.05 versus control + CPZ and Se groups. ^e p ≤ 0.001 versus FM group (2c).

Figure 3

Effect of selenium (Se) treatment on [Ca²⁺]_i concentration through block of TRPM2 gate in dorsal root ganglion neuron (DRGN) (a) and sciatic nerve (SciN) (b) of control and FM-induced rats. (n = 8 and mean ± SD). The animals received intraperitoneal Se for 2 weeks after FM induction. Then, these dissected neurons of control and FM groups were further in vitro treated with CumHPx (1 mM) and ACA (0.025 mM) before loading Fura-2 for 120 seconds. (^a p ≤ 0.05 and ^b p ≤ 0.001 versus control. (^c p ≤ 0.001 and ^d p ≤ 0.05 versus control + ACA and Se groups. ^e p ≤ 0.001 versus FM group (3c).

Figure 4

Effects of selenium (Se) on TRPV1 channel activation in dorsal root ganglion neuron (DRGN) of control and FM-induced rat. The TRPV1 currents in DRGN were stimulated by extracellular CAPS (10 μM in patch chamber) but they were blocked by extracellular CPZ (0.1 mM) in the patch-chamber. W.C. is whole cell. (a) Control: Original recordings from control neuron. (b) Control + CAPS group (without FM induction). (c) FM group (with FM induction). (d) FM + Se group: Se was administrated to the rats after FM induction. (e) Se group: Se was administrated to the rats without FM induction. (f) TRPV1 channel current densities in the DRGN. The numbers in parentheses indicated n numbers of groups. (^a p ≤ 0.001 versus control. ^b p ≤ 0.001 versus control + CAPS group. ^c p ≤ 0.001 versus control + CAPS + CPZ group. ^d p ≤ 0.001 versus FM + CAPS group. ^e p ≤ 0.001 versus FM + CAPS + CPZ group).

Figure 5

Effects of selenium (Se) on TRPM2 channel activation in dorsal root ganglion neuron (DRGN) of fibromyalgia (FM)-induced rat. The TRPM2 currents in DRGN were stimulated by intracellular ADPR (1 mM in patch pipette) but they were blocked by extracellular TRPM2 antagonist (ACA and 0.025 mM) in the patch-chamber. W.C. is whole cell. Control (without FM induction and stimulation): Original recordings from control neuron. (b) Control + ADPR group (without FM induction). (c) FM group (with FM induction). (d) FM + Se group: The rats received Se after FM induction. (e) Se group: The rats received Se without FM induction. (f) TRPM2 channel current densities in the DRGN. The numbers in parentheses indicated n numbers of groups were indicated by numbers in parentheses. (^a p ≤ 0.001 versus control. ^b p ≤ 0.001 versus control + ADPR group. ^c p ≤ 0.001 versus control + ADPR + ACA group. ^d p ≤ 0.001 versus FM + ADPR group. ^e p ≤ 0.001 versus FM + ADPR + ACA group).

Figure 6

Effects of selenium on the apoptosis, cell viability (MTT), mitochondrial membrane depolarization (JC-1), intracellular ROS production and caspase 3 and 9 values through TRPV1 (a,c and e) and TRPM2 (b,d and f) channel activations in the DRGN of FM-induced rats (mean ± SD and n = 3). Values expressed as fold increase (experimental/control). These neurons were dissected from control, FM and treated animals. The TRPV1 and TRPM2 gates in the neurons were opened with capsaicin (10 μM) and cumene hydroperoxide (1 mM) although they were blocked by CPZ (0.1 mM) and ACA (0.025 mM), respectively. (^a p ≤ 0.05 and ^b p ≤ 0.001 versus control and control + CPZ groups. ^c p ≤ 0.05 versus Se group. ^d p ≤ 0.001 versus Se and Se + CPZ groups. ^e p ≤ 0.001 versus FM and FM + CPZ groups. ^f p ≤ 0.05 versus FM + Se groups).

Figure 7

Effects of selenium on the caspase 3, caspase 9 and PARP1 expression levels in DRGN (a and d), SciN (b and d) and gastrocnemius muscle (c and d) of rats with FM (mean ± SD and n = 3). (^a p ≤ 0.05 and ^bp ≤ 0.001 versus control. ^c p ≤ 0.05 and ^d p ≤ 0.001 versus FM group. ^e p ≤ 0.001 versus FM + Se group).