TNF-α enhances the currents of voltage gated sodium channels in uninjured dorsal root ganglion neurons following motor nerve injury
Research Highlights
► Lumbar 5 ventral root transection (L5-VRT) increased the currents of TTX-S, Nav1.8 and firing rate in DRG neurons. ► rrTNF enhanced TTX-S and Nav1.8 currents in cultured DRG neurons. ► TNFR-1 knockout attenuated mechanical allodynia and increase in sodium currents.
Introduction
The ectopic discharges in injured (Blumberg and Janig, 1984, Liu et al., 2000, Wall et al., 1974, Yoon et al., 1996) and uninjured DRG neurons (Ali et al., 1999, Wu et al., 2001) induced by peripheral nerve injury may contribute to neuropathic pain, manifested as allodynia, hyperalgesia and ongoing pain. Because the behavioral signs of neuropathic pain cannot be prevented or reversed by acute section of the dorsal root of the injured DRG neurons (Li et al., 2000), it has been suggested that the ectopic discharges arising from the uninjured but not from injured DRG neurons play a crucial role in neuropathic pain. A large body of evidence has demonstrated that the abnormal expression of voltage-gated sodium channels (VGSCs) in DRG neurons is responsible for the ectopic discharges (Chahine et al., 2005, Matzner and Devor, 1994). For example, the Nav1.3, a tetrodotoxin-sensitive (TTX-S) sodium channel, which is hardly detectable in adult DRG neurons, is up-regulated in several neuropathic pain models (Black et al., 1999, Dib-Hajj et al., 1999, Kim et al., 2001, Waxman et al., 1994). Consistent with this observation, the inhibition of Nav1.3 expression by intrathecal injection of Nav1.3 specific antisense oligodeoxynucleotides results in the attenuation of mechanical allodynia and thermal hyperalgesia induced by peripheral nerve injury (Hains et al., 2004). Thus, over-expression of Nav1.3 may contribute to neuropathic pain. Mature DRG neurons contain two subtypes of TTX-resistant (TTX-R) sodium channels, Nav1.8 and Nav1.9, both of which are expressed almost exclusively in nociceptors (Amaya et al., 2000). Nav1.8 is substantially down-regulated in injured DRG neurons (Cummins and Waxman, 1997, Dib-Hajj et al., 1996) and is up-regulated in uninjured DRG neurons following spinal nerve ligation (Zhang et al., 2004) but is not affected in chronic nerve constriction injury (CCI) or spared nerve injury (SNI) models (Decosterd et al., 2002). Importantly, blockade (Ekberg et al., 2006, Jarvis et al., 2007) or specific knockdown of Nav1.8 with antisense oligodeoxynucleotides (Dong et al., 2007, Lai et al., 2002) can effectively suppress or reverse neuropathic pain. Similar to Nav1.8, the expression of Nav1.9 is reduced in injured DRG neurons (Dib-Hajj et al., 1998) and is up-regulated in uninjured neurons after peripheral nerve injury (Berta et al., 2008). In the Nav1.9 knockout mice inflammatory pain behavior is substantially attenuated, while pain behavior is unchanged in neuropathic pain models (Amaya et al., 2006).
Consistent with the previous studies, our recent work (He et al., 2010) showed that selective injury of motor neurons by L5-VRT, which induces behavioral signs of neuropathic pain in bilateral hind paws (Li et al., 2002), up-regulates Nav1.3 and Nav1.8 in uninjured DRG neurons at both mRNA and protein levels, indicating that nerve injury may modulate the expression of the sodium channels indirectly. Furthermore, we have shown that TNF-α and TNFR-1 are up-regulated in L4 and L5 DRGs following L5-VRT (Xu et al., 2006) and that inhibition of TNF-α synthesis or genetic deletion of TNFR1 prevents the up-regulation of Nav1.3 and Nav1.8 in DRG neurons (He et al., 2010). Thus, elevated levels of TNF-α may be responsible for the up-regulation of these sodium channels. In the present work using the L5-VRT animal model, cultured adult DRG neurons and TNFR-1 knockout mice, we tested whether the up-regulated sodium channels are functional and whether the augmented sodium channel currents result in the excitability of DRG neurons with patch clamp technique. The role of elevated TNF-α in the functional change was evaluated.
Section snippets
Animals
Male Sprague–Dawley rats weighing 150–250 g were used. Animals were housed in separate cages at a room temperature kept at 24 ± 1 °C and 50–60% humidity, under a 12:12-h light/dark cycle. They had access to food and water ad libitum. Experimental procedures were approved by the local animal care committee and were carried out in accordance with the guidelines of the National Institutes of Health on animal care and the ethical guidelines for investigation of experimental pain in conscious animals (
The current densities of TTX-sensitive and Nav1.8 but not Nav1.9 channels in DRG neurons are increased following L5-VRT
In our previous work we showed that both Nav1.3 and Nav1.8 channels are up-regulated in uninjured L4 and L5 DRG neurons at mRNA and protein levels following selective damage of motor neurons by L5-VRT (He et al., 2010). To test whether the over-expressed sodium channels are functional, we measured the peak current densities (pA/pF) of TTX-S, Nav1.8, and Nav1.9 in L4/L5 DRG neurons isolated from L5-VRT rats and sham-operated rats. As shown in Fig. 2, the currents of TTX-S and Nav1.8 but not
Discussion
In the present study, we demonstrated that the TTX-S, Nav1.8 but not Nav1.9 channel currents in uninjured DRG neurons increased following selective damage of motor neurons. The changes were associated with the increase in excitability of the sensory neurons. TNF-α in CSF and in DRG tissue increased after L5-VRT, and rrTNF increased TTX-S, Nav1.8 and Nav1.9 currents in cultured DRG neurons. Genetic deletion of TNFR-1 attenuated the mechanical allodynia and prevented the increase in sodium
Conclusions
In summary, following peripheral nerve injury, the increased TNF-α may contribute to neuropathic pain by the up-regulation of VGSCs in uninjured DRG neurons via activation of TNFR-1.
Competing interests
The authors declare that they have no competing interests.
Acknowledgments
This work was supported by National Natural Science foundation of China (Nos.30800336, 30970957, 30900436).
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