Sodium Channel Nav1.7 in Sympathetic Neurons Contributes to Pain Phenotypes in Mice
In people, loss-of-function mutations in the voltage-gated sodium channel Nav1.7 result in total insensitivity to pain. Replicating that absolute loss of pain perception in animals has proven a challenge—globally knocking out Nav1.7 in mice is lethal, and targeted deletion of the channel in a subset of sensory neurons results in only partial deficits in pain sensation. Now, a team led by John Wood, University College London, UK, reports that rendering mice impervious to most types of pain requires deleting Nav1.7 not only in sensory neurons, but also in sympathetic neurons. In particular, removing the channel from both cell types protected the animals from neuropathic pain.
The mice give the field a much-needed animal model for dissecting the biology of Nav1.7 in pain, and may shed light on how to better target the channel therapeutically. In an immediate illustration of that idea, Wood and collaborators used the mice to uncover a novel role for Nav1.7 in neurotransmitter release in the spinal cord. The study appeared online April 24 in Nature Communications.
“This is an elegant study that provides a boost to the field,” said Sulayman Dib-Hajj, Yale University School of Medicine, New Haven, Connecticut, US, and the Veterans Affairs Medical Center, West Haven, Connecticut, US. Dib-Hajj, whose recent work has implicated Nav1.7 in the painful disorders inherited erythromelalgia and small fiber neuropathy (see PRF related news story; Dib-Hajj et al., 2010; Han et al., 2012), was not involved in the new study.
The results help explain the long-standing discrepancy between the phenotypes of Nav1.7 loss in people versus mice, he said. “Wood and coauthors produce reasonable data that you have to knock [Nav1.7] out in both sensory and sympathetic neurons to see phenotypes relevant to neuropathic pain or response to noxious heat that was not abrogated in the other knockouts.” However, he said, “Exactly what [Nav1.7] is doing in sympathetic neurons, time will tell.”
Targeted knockouts reveal cell-specific roles for Nav1.7
Nav1.7, encoded by the gene SCN9A, is required for the initiation of action potentials in sensory neurons. In humans, loss-of-function mutations produce congenital insensitivity to pain (CIP; see Cox et al., 2006). To create a mouse model of Nav1.7 deficiency, the Wood lab previously produced a knockout targeted to sensory neurons that express the related channel Nav1.8. Those mice showed diminished pain response to mechanical insults and deficits in inflammatory pain, but had normal responses to acute thermal stimuli and nerve injury (Nassar et al., 2004; Nassar et al., 2005).
In the new study, first author Michael Minett and colleagues generated additional knockout strains to probe the role of Nav1.7 in a larger population of sensory neurons, and in sympathetic neurons. In one strain, the investigators targeted Cre recombinase-catalyzed Nav1.7 deletion to sensory neurons using the promoter of the actin-binding protein Advillin. They confirmed expression of Advillin-driven Cre in all dorsal root ganglion (DRG) neurons in the transgenic mice, supporting the idea that the knockouts affect all sensory neurons. In the other strain, channel deletion was targeted using the promoter of the signaling protein Wnt1, which drives expression in the precursors of sensory and sympathetic neurons. After confirming loss of SCN9A mRNA in either DRG neurons (Advillin mice) or DRG and sympathetic cervical ganglia neurons (Wnt1 mice), the researchers extensively characterized both strains for acute and inflammatory pain responses, and for neuropathic pain after spinal nerve transection.
Unexpectedly, deleting Nav1.7 in all sensory neurons did not render the animals insensitive to pain. The Advillin mice remained responsive to some types of painful heat or cold, and developed neuropathic pain similar to their wild-type littermates. However, deleting Nav1.7 in both sympathetic and sensory neurons abolished the remaining responses to painful heat and nerve injury. The mice were still not completely pain free: Both strains retained their ability to respond to extreme cold, which appeared to require Nav1.8 rather than Nav1.7.
Wood says he and his colleagues were “amazed” to see that the mice lacking Nav1.7 in all sensory neurons showed a response to noxious heat. “So we wondered if we took the thing out in sympathetic neurons as well, if that would have an effect, and it did. We were very surprised, actually.”
The idea that the autonomic nervous system—better known for mounting “fight or flight” responses to stress—contributes to neuropathic pain is not new. “There is a lot of literature from the ’70s, ’80s, and ’90s saying that sympathetic neurons are very important in neuropathic pain,” Wood told PRF. Sympathetic sprouting into the DRG, and interaction between sympathetic and sensory neurons, have been documented in rodent models of chronic pain (for a recent example, see Xie et al., 2011), including the spinal nerve transection model used for the current study.
Interest in the sympathetic nervous system died down, however, when sympathetic nerve blocks proved to be generally ineffective to control pain in human studies. The new findings raise the possibility that it may be necessary to block both sympathetic and sensory nerves to relieve pain, and could serve to refocus attention on sensory-sympathetic interactions in neuropathic pain, Wood said.
A novel role for Nav1.7 in the central nervous system?
The study held other surprises, too. The team found that Nav1.7-deficient sensory neurons failed to release the neurotransmitter substance P, or to display synaptic potentiation in the dorsal horn of the spinal cord in response to electrical stimulation of the sciatic nerve—indicating that Nav1.7 is required not only to generate action potentials in the periphery, but also to send the signals on to postsynaptic neurons in the central nervous system. (A similar phenomenon was previously reported for Nav1.7 in olfactory neurons; see PRF related news story.)
Nav1.7’s role in neurotransmitter release is intriguing, said Dib-Hajj. The mechanism by which Nav1.7 affects synaptic signaling, however, remains to be determined. “Nav1.7 in central termini may be important to keep the action potential strong enough to induce release of neurotransmitters,” he told PRF.
Wood, on the other hand, said Nav1.7 may contribute to synaptic signaling directly, because electrical stimulation of nerve trunks evokes action potentials in all neurons. He said his group is looking to see whether Nav1.7 associates with calcium channels that control neurotransmitter release.
Translational efforts
Since the discovery of Nav1.7’s role in CIP and additional diseases involving gain-of-function mutations such as erythromelalgia and paroxysmal extreme pain disorder, the channel has been the target of much drug discovery activity, and some compounds have reached early-phase clinical testing for neuropathic pain conditions (see PRF related news story). The new knockouts offer researchers a model for exploring the contribution of Nav1.7 to neuropathic and other forms of pain, and should help efforts to develop Nav1.7 blockers for pain treatment.
One example is the finding that Nav1.7 plays a role in the central nervous system, and not just in the periphery, where its function is already appreciated. Does that mean that Nav1.7-targeting drugs will need to cross the blood-brain barrier? That, Wood said, “is an interesting question.”
Top image: Staining of Advillin-Cre–positive DRG neurons.
Credit for both images: Minett et al., 2012, Nature Communications, distributed under a Creative Commons license.
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