Distinction among Neuronal Subtypes of Voltage-Activated Sodium Channels by μ-Conotoxin PIIIA

Abstract

The functional properties of most sodium channels are too similar to permit identification of specific sodium channel types underlying macroscopic current. Such discrimination would be particularly advantageous in the nervous system in which different sodium channel family isoforms are coexpressed in the same cell. To test whether members of the mu-conotoxin family can discriminate among known neuronal sodium channel types, we examined six toxins for their ability to block different types of heterologously expressed sodium channels. PIIIA mu-conotoxin blocked rat brain type II/IIA (rBII/IIA) and skeletal muscle sodium current at concentrations that resulted in only slight inhibition of rat peripheral nerve (rPN1) sodium current. Recordings from variant lines of PC12 cells, which selectively express either rBII/IIA or rPN1 channel subtypes, verified that the differential block by PIIIA also applied to native sodium current. The sensitivity to block by PIIIA toxin was then used to discriminate between rBII/IIA and rPN1 sodium currents in NGF-treated PC12 cells in which both mRNAs are induced. During the first 24 hr of NGF-treatment, PN1 sodium channels accounted for over 90% of the sodium current. However, over the ensuing 48 hr period, a sharp rise in the proportion of rBII/IIA sodium current occurred, confirming the idea, based on previous mRNA measurements, that two distinct sodium channel types appear sequentially during neuronal differentiation of PC12 cells.

Characterization and Developmental Changes of Na+Currents of Petrosal Neurons With Projections to the Carotid Body

“…4A). Assuming a single binding/ blocking site for TTX and using a Langmuir inhibition isoform equation, this corresponds to a half-maximal inhibition of 5.5 nM for TTX, which is similar to that reported for recombinant rat Na v 1.1 (9.6 nM) , Na v 1.6 (6.4 nM) , and Na v 1.7 (4 nM) (Safo et al 2000) channels expressed in Xenopus oocytes. Two of the labeled petrosal ganglion cells exposed to TTX exhibited both fastinactivating current, which was blocked by TTX, and slowly inactivating current, which was not (data not shown).…”

Structure-Activity Relationships of Hainantoxin-IV and Structure Determination of Active and Inactive Sodium Channel Blockers

“…Therefore, we hypothesize that HNTX-IV binds to the sodium channels with the surface encompassing mainly Lys 27 Structural Comparison of HNTX-IV with -Conotoxins--Conotoxins are receptor site 1 sodium channel blockers purified from the venom of marine cone snails that act selectively to occlude the pore of VGSCs by competing with TTX and saxitoxin. Among the -conotoxins, GIIIA/B and GS specifically inhibit rat skeletal muscle sodium channels (rNa(v)1.4), whereas PIIIA inhibits neuronal as well as muscle TTX-S sodium channels (33)(34)(35)(36)(37)(38)(39). The three-dimensional structure of HNTX-IV has little resemblance to the three-loop -conotoxins (GIIIA/B and PIIIA) in agreement with the low sequence identity between them and their different cystine frameworks (27,40,41).…”

ConusVenoms: A Rich Source of Novel Ion Channel-Targeted Peptides

“…A recent example is the demonstration that Na v 1.2 and Na v 1.7 channels appear sequentially during neuronal differentiation of PC12 cell lines (158). Interestingly, in rat brain neurons -PIIIA seems to preferentially inhibit persistent over transient voltage-activated Na currents (135).…”