The mechanism of analgesia in Na_V1.7 null mutants

Abstract

Deletion of SCN9A encoding the voltage-gated sodium channel NaV1.7 in humans leads to profound pain insensitivity and anosmia. Conditional deletion of NaV1.7 in sensory neurons of mice also abolishes pain suggesting the locus of analgesia is the nociceptor.Here we demonstrate that NaV1.7 knockout mice have essentially normal nociceptor activity using in vivo calcium imaging and extracellular recording. However, glutamate and substance P release from nociceptor central terminals in the spinal cord is greatly reduced by an opioid-dependent mechanism. Analgesia is also substantially reversed by central but not peripheral application of opioid antagonists. In contrast, the lack of neurotransmitter release from olfactory sensory neurons is opioid-independent. Male and female humans with NaV1.7 null mutations show naloxone reversible analgesia. Thus opioid-dependent inhibition of neurotransmitter release is the principal mechanism of NaV1.7 null analgesia in mice and humans.deficits in thermal, mechanical, inflammatory and some forms of neuropathic pain. 15,16 To determine the mechanism of NaV1.7 null analgesia, we used complementary optical, electrophysiological and pharmacological methods to study nociceptor function in vivo in mice and humans lacking NaV1.7. ResultsDeletion of NaV1.7 in sensory neurons decreases pain sensitivity without silencing peripheral nociceptorsWe deleted Scn9a encoding NaV1.7 in peripheral sensory neurons of mice. Advillin-Cre or Wnt1-Cre was used to excise the floxed Scn9a allele, resulting in knockout of NaV1.7 restricted, respectively, to sensory neurons or to neural crest-derived neurons. 15 We performed whole-cell voltage-clamp recordings of voltage-gated sodium currents in cultured sensory neurons from solution contained 125 mM K-gluconate, 6 mM KCl, 10 mM HEPES, 0.1 mM EGTA, 2 mM Mg-ATP, pH 7.3 with KOH, and osmolarity of 290-310 mOsm. Cells were targeted for patching in the inner and outer Lamina II and visualized through an Eclipse E600FN Nikon microscope (Nikon, Japan) equipped with infrared differential interference contrast (IR-DIC) connected to a digital camera (Nikon, DS-Qi1Mc). Cells were voltage-clamped at -70 mV and spontaneous excitatory post synaptic currents (sEPSCs) recorded. sEPSCs were automatically detected using ClampFit in a 5 s window for each cell. Ex vivo olfactory bulb electrophysiology:Acute MOB slices were prepared from 4 -11 week old mice (male and female) anesthetized with CO2 before decapitation. OBs were rapidly dissected in ice-cold oxygenated (95% O2, 5% CO2) solution containing the following (in mM): 83 NaCl, 26.2 NaHCO3, 1 NaH2PO4, 2.5 KCl, 3.3 MgCl2, 0.5 CaCl2, 70 sucrose, pH 7.3 (osmolarity, 300 mOsm/l). The tissue was mounted on a

Long-lasting analgesia via targeted in situ repression of Na V 1.7 in mice

“…(ii) Although long-term studies were performed (308 days after a single intrathecal injection), studies to evaluate the actual duration of treatment and whether any compensatory mechanisms take place because of Na V 1.7 repression must be performed. In particular, previous work has reported compensatory changes in the endogenous opioid system (proenkephalin up-regulation) in response to Na V 1.7 knockout in mice (68)(69)(70). (iii) Further studies must be performed to explore the properties of repeat dosing at the spinal level.…”

Examination of the contribution of Nav1.7 to axonal propagation in nociceptors

“…However, this is in contrast to numerous studies reporting that propagation in a proportion (75-96%) of C-fibres is maintained when Nav1.7 (and other TTX-S channels) are blocked following axonal TTX application [24; 27; 37; 45; 52]. Studies employing genetic knockdown of Nav1.7 also report that a proportion of C-fibres are capable of propagation without the channel [21; 33].…”

Peripheral mechanisms of peripheral neuropathic pain

“…For example, researchers at Genentech have used tarantula toxins to investigate the structural basis of NaV 1.7 inhibition with the end goal of accelerating the development of next generation modulators (Xu et al, 2019). Some work proposes that NaV 1.7 activity also modulates endogenous opioid peptide release, suggesting that the combined action of a NaV 1.7 inhibitor and an opioid molecule may improve synergistic analgesia with fewer side effects (MacDonald et al, 2021). However, a publication has sought to disprove this mechanism suggesting that upregulation of opioid peptides play no role in the analgesic effects of NaV 1.7 blockers (Bankar et al, 2018).…”