Na_Vβ Subunits Modulate the Inhibition of Na_V1.8 by the Analgesic Gating Modifier μO-Conotoxin MrVIB

Abstract

Voltage-gated sodium channels (VGSCs) consist of a pore-forming ␣-subunit and regulatory ␤-subunits. Several families of neuroactive peptides of Conus snails target VGSCs, including Oconotoxins and -conotoxins. Unlike -conotoxins and the guanidinium alkaloid saxitoxin (STX), which are pore blockers, O-conotoxins MrVIA and MrVIB inhibit VGSCs by modifying channel gating. O-MrVIA/B can block Na V 1.8 (a tetrodotoxinresistant isoform of VGSCs) and have analgesic properties. The effect of Na V ␤-subunit coexpression on susceptibility to block by O-MrVIA/B and STX has, until now, not been reported. Here, we show that ␤1-, ␤2-, ␤3-, and ␤4-subunits, when individually coexpressed with Na V 1.8 in Xenopus laevis oocytes, increased the k on of the block produced by O-MrVIB (by 3-, 32-, 2-, and 7-fold, respectively) and modestly decreased the apparent k off . Strong depolarizing prepulses markedly accelerated MrVIB washout with rates dependent on ␤-subunit coexpression. Thus, coexpression of ␤-subunits with Na V 1.8 can strongly influence the affinity of the conopeptide for the channel. This observation is of particular interest because ␤-subunit expression can be dynamic, e.g., ␤2-expression is up-regulated after nerve injury (J Neurosci, 25: 10970 -10980, 2005); therefore, the effectiveness of a O-conotoxin as a channel blocker could be enhanced by the conditions that may call for its use therapeutically. In contrast to MrVIB's action, the STX-induced block of Na V 1.8 was only marginally, if at all, affected by coexpression of any of the ␤-subunits. Our results raise the possibility that O-conotoxins and perhaps other gating modifiers may provide a means to functionally assess the ␤-subunit composition of VGSC complexes in neurons.

Conus Venom Peptide Pharmacology

“…Indeed, voltage-dependent relief of Na v 1.4 inhibition by MrVIB was attributed to the voltage sensor of domain II (Leipold et al, 2007), another site shared with ␤ scorpion toxins. Similar effects were also reported for Na v 1.8, where the K off of MrVIB was also accelerated by strong depolarizations (Wilson et al, 2011b). However, compared with Na v 1.4, the K off of MrVIB at Na v 1.8 was significantly slower and required repeated depolarizations to relieve block (Wilson et al, 2011b).…”

“…Similar effects were also reported for Na v 1.8, where the K off of MrVIB was also accelerated by strong depolarizations (Wilson et al, 2011b). However, compared with Na v 1.4, the K off of MrVIB at Na v 1.8 was significantly slower and required repeated depolarizations to relieve block (Wilson et al, 2011b).…”

“…Significant differences in their affinity at native Na v 1.8 expressed in DRG neurons and heterologously expressed Na v 1.8 have been reported. These presumably arise because of the presence of auxiliary ␤ subunits, particularly ␤2, which has recently been shown to significantly increase the rate of Na v 1.8 inhibition by MrVIB and to increase its potency accordingly (Wilson et al, 2011b). It is noteworthy that O-conotoxins can also differentiate between the TTXresistant subtypes Na v 1.8 and -1.9, because the persistent Na v 1.9 current in DRG neurons was unaffected by MrVIB (Ekberg et al, 2006).…”

“…As observed for -conotoxins (see section II.C), interactions of O-conotoxins with Na v channel types are also influenced by auxiliary subunit expression. Na v ␤2 subunits are up-regulated in sensory neurons in neuropathic pain models (Pertin et al, 2005) and interact with Na v s to increase the on-rate of MrVIB in heterologous systems (Wilson et al, 2011b), providing the O-conotoxins with a window of functional selectivity. It is noteworthy that Na v ␤3 subunits are also up-regulated in pain states (Shah et al, 2000), and their expression levels influence gating of Na v 1.3 (Cusdin et al, 2010) to potentially influence O-conotoxin pharmacology in a diseasedependent manner.…”

The Sophisticated Peptide Chemistry of Venomous Animals as a Source of Novel Insecticides Acting on Voltage-Gated Sodium Channels

“…These results suggest that the auxiliary subunits are functionally conserved among different insect species. A recent study has shown that that the channel affinity of the conotoxin MrVIB, a mammalian sodium channel blocker isolated from the cone snail Conus marmoreus, is strongly influenced by the co expression of -subunits (Wilson et al, 2011). Their high conservation among different insect species, their wide expression in distinct tissues together with the observation that they are involved in the neurotoxin-channel interaction raises the intriguing question whether the insect auxiliary subunits might represent interesting new phyla-selective targets for neurotoxin insecticides.…”