Tetrodotoxin-resistant Sodium Channel Nav1 Research Articles

Growth Differentiation Factor-15 Produces Analgesia by Inhibiting Tetrodotoxin-Resistant Nav1.8 Sodium Channel Activity in Rat Primary Sensory Neurons

Growth differentiation factor 15 (GDF-15) is a member of the transforming growth factor-β superfamily. It is widely distributed in the central and peripheral nervous systems. Whether and how GDF-15 modulates nociceptive signaling remains unclear. Behaviorally, we found that peripheral GDF-15 significantly elevated nociceptive response thresholds to mechanical and thermal stimuli in naïve and arthritic rats. Electrophysiologically, we demonstrated that GDF-15 decreased the excitability of small-diameter dorsal root ganglia (DRG) neurons. Furthermore, GDF-15 concentration-dependently suppressed tetrodotoxin-resistant sodium channel Nav1.8 currents, and shifted the steady-state inactivation curves of Nav1.8 in a hyperpolarizing direction. GDF-15 also reduced window currents and slowed down the recovery rate of Nav1.8 channels, suggesting that GDF-15 accelerated inactivation and slowed recovery of the channel. Immunohistochemistry results showed that activin receptor-like kinase-2 (ALK2) was widely expressed in DRG medium- and small-diameter neurons, and some of them were Nav1.8-positive. Blockade of ALK2 prevented the GDF-15-induced inhibition of Nav1.8 currents and nociceptive behaviors. Inhibition of PKA and ERK, but not PKC, blocked the inhibitory effect of GDF-15 on Nav1.8 currents. These results suggest a functional link between GDF-15 and Nav1.8 in DRG neurons via ALK2 receptors and PKA associated with MEK/ERK, which mediate the peripheral analgesia of GDF-15.

Modulation of Nav1.8 by Lysophosphatidic Acid in the Induction of Bone Cancer Pain.

Given that lysophosphatidic acid (LPA) and the tetrodotoxin-resistant sodium channel Nav1.8 are both involved in bone cancer pain, the present study was designed to investigate whether crosstalk between the LPA receptor LPA1 (also known as EDG2) and Nav1.8 in the dorsal root ganglion (DRG) contributes to the induction of bone cancer pain. We showed that the EDG2 antagonist Ki16198 blocked the mechanical allodynia induced by intrathecal LPA in naïve rats and attenuated mechanical allodynia in a rat model of bone cancer. EDG2 and Nav1.8 expression in L4-6 DRGs was upregulated following intrathecal or hindpaw injection of LPA. EDG2 and Nav1.8 expression in ipsilateral L4-6 DRGs increased with the development of bone cancer. Furthermore, we showed that EDG2 co-localized with Nav1.8 and LPA remarkably enhanced Nav1.8 currents in DRG neurons, and this was blocked by either a protein kinase C (PKC) inhibitor or a PKCε inhibitor. Overall, we demonstrated the modulation of Nav1.8 by LPA in DRG neurons, and that this probably underlies the peripheral mechanism by which bone cancer pain is induced.

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Activation of Tetrodotoxin-Resistant Sodium Channel NaV1.9 in Rat Primary Sensory Neurons Contributes to Melittin-Induced Pain Behavior

Tetrodotoxin-resistant (TTX-R) sodium channels NaV1.8 and NaV1.9 in dorsal root ganglion (DRG) neurons play important roles in pathological pain. We recently reported that melittin, the major toxin of whole bee venom, induced action potential firings in DRG neurons even in the presence of a high concentration (500nM) of TTX, indicating the contribution of TTX-R sodium channels. This hypothesis is fully investigated in the present study. After subcutaneous injection of melittin, NaV1.8 and NaV1.9 significantly upregulate mRNA and protein expressions, and related sodium currents also increase. Double immunohistochemical results show that NaV1.8-positive neurons are mainly medium- and small-sized, whereas NaV1.9-positive ones are only small-sized. Antisense oligodeoxynucleotides (AS ODNs) targeting NaV1.8 and NaV1.9 are used to evaluate functional significance of the increased expressions of TTX-R sodium channels. Behavioral tests demonstrate that AS ODN targeting NaV1.9, but not NaV1.8, reverses melittin-induced heat hypersensitivity. Neither NaV1.8 AS ODN nor NaV1.9 AS ODN affects melittin-induced mechanical hypersensitivity. These results provide previously unknown evidence that upregulation of NaV1.9, but not NaV1.8, in small-sized DRG neurons contributes to melittin-induced heat hypersensitivity. Furthermore, melittin-induced biological effect indicates a potential strategy to study properties of TTX-R sodium channels.

Increased expression of tetrodotoxin-resistant sodium channels Nav1.8 and Nav1.9 within dorsal root ganglia in a rat model of bone cancer pain

In attempt to understand the underlying mechanisms of cancer-induced bone pain, we investigated the presence of two tetrodotoxin-resistant sodium channels, Nav1.8 (SNS/PN3) and Nav1.9 (SNS2/NaN), in dorsal root ganglia (DRG) neurons in an animal model of bone cancer pain. Thirty-six female Sprague-Dawley rats were randomized into three groups: Sham operation group (Sham), cancer-bearing animals killed after 7 days (C7) and cancer-bearing animals killed after 14 days group (C14). After establishment of bone cancer pain model, behavioral tests were carried out to determine the paw withdrawal threshold (PWT) of mechanical and thermal hyperalgesia, respectively. Real-time RT-PCR, Western bolt and Immunofluorescence were used to determine the mRNA and protein expression of Nav1.8 and Nav1.9 in ipsilateral lumbar 4-5 DRG. Compared to Sham group, PWT of mechanical and thermal hyperalgesia in C14 group displayed a significant decrease (P<0.01) from post-operation day (POD) 5 and POD7 to the end point of the observation, respectively. Compared to Sham group, the relative mRNA expression of Nav1.8 and Nav1.9 exhibited a significant up-regulation in C14 group (8.9 times and 9 times, respectively, P<0.01) but not C7 group (1.5 times and 2.4 times, respectively). Western blot and Immunofluorescence revealed an apparent increase of Nav1.8 (P<0.05) and Nav1.9 (P<0.05) protein in C14 group compared with Sham group. The up-regulation of mRNA and protein levels of Nav1.8 and Nav1.9 suggested their potential involvement in the development and maintenance of bone cancer pain.

Ion channels involved in cold detection in mammals: TRP and non-TRP mechanisms.

Substantial progress in understanding thermal transduction in peripheral sensory nerve endings was achieved with the recent cloning of six thermally gated ion channels from the TRP (transient receptor potential) super-family. Two of these channels, TRP melastatin 8 (TRPM8) and TRP ankyrin 1 (TRPA1), are expressed in dorsal root ganglion (DRG) and trigeminal ganglion (TG) neurons, are activated by various degrees of cooling, and are candidates for mediating gentle cooling and noxious cold, respectively. However, accumulating evidence suggests that more than just these two channels are involved in cold sensing in mammals. A recent report described a critical role of the voltage-gated tetrodotoxin-resistant sodium channel Nav1.8 in perceiving intense cold and noxious stimuli at cold temperatures. Other ion channels, such as two-pore domain background potassium channels (K2P), are known to be expressed in peripheral nerves, have pronounced temperature dependence, and may contribute to cold sensing and/or cold hypersensitivity in pain states. This article reviews the evidence supporting a role for each of these channels in cold transduction, focusing on their biophysical properties, expression pattern, and modulation by pro-inflammatory mediators.

Discovery of potent furan piperazine sodium channel blockers for treatment of neuropathic pain

The synthesis and pharmacological characterization of a novel furan-based class of voltage-gated sodium channel blockers is reported. Compounds were evaluated for their ability to block the tetrodotoxin-resistant sodium channel Na v1.8 (PN3) as well as the Na v1.2 and Na v1.5 subtypes. Benchmark compounds from this series possessed enhanced potency, oral bioavailability, and robust efficacy in a rodent model of neuropathic pain, together with improved CNS and cardiovascular safety profiles compared to the clinically used sodium channel blockers mexiletine and lamotrigine.

Differential expression of tetrodotoxin-resistant sodium channels Na v1.8 and Na v1.9 in normal and inflamed rats

In an attempt to understand mechanisms underlying peripheral sensitization of primary afferent fibers, we investigated the presence of the tetrodotoxin-resistant Na + channel subunits Na v1.8 (SNS) and Na v1.9 (SNS2) on axons in digital nerves of normal and inflamed rat hindpaws. In normal animals, 14.3% of the unmyelinated and 10.7% of the myelinated axons labeled for the Na v1.8 subunit. These percentages significantly increased in 48 h inflamed animals to 22.0% (1.5-fold increase) and 57.5% (6-fold increase) for unmyelinated and myelinated axons, respectively. In normal animals, Na v1.9 labeled 9.9% of the unmyelinated and 2.1% of the myelinated axons and following inflammation, the proportion of Na v1.9-labeled unmyelinated axons significantly decreased to 3.0% with no change in the proportion of labeled myelinated axons. These data indicate that Na v1.8 and Na v1.9 subunits are transported to the periphery in normal animals and are differentially regulated during inflammation. The massive increase in Na v1.8 expression in myelinated axons suggests that these may contribute to peripheral sensitization and inflammatory hyperalgesia.

Tetrodotoxin-resistant sodium channels Na v1.8/SNS and Na v1.9/NaN in afferent neurons innervating urinary bladder in control and spinal cord injured rats

Tetrodotoxin-resistant (TTX-R) sodium channels Na v1.8/SNS and Na v1.9/NaN are preferentially expressed in small diameter dorsal root ganglia (DRG) neurons. The urinary bladder is innervated by small afferent neurons from L6/S1 DRG, of which ∼75% exhibit high-threshold action potentials that are mediated by TTX-R sodium channels. Following transection of the spinal cord at T8, the bladder becomes areflexic and then gradually hyper-reflexic, and there is an attenuation of the TTX-R sodium currents in bladder afferent neurons. In the present study, we demonstrate that Na v1.8 is expressed in both bladder and non-bladder afferent neurons, while Na v1.9 is expressed in non-bladder afferent neurons but is rarely observed in bladder afferent neurons. In spinal cord transected rats 28–32 days following transection, there is a decreased expression of Na v1.8 sodium channels in bladder afferents, but no change in the expression of Na v1.8 in non-bladder afferent neurons. Both bladder and non-bladder afferent neurons exhibit limited increases in Na v1.9 expression following spinal cord transection. These results demonstrate that the expression of TTX-R channels in bladder afferent neurons changes after spinal cord transection, and these changes may contribute to the increased excitability of these neurons following spinal cord injury.