A Scorpion α-Like Toxin That Is Active on Insects and Mammals Reveals an Unexpected Specificity and Distribution of Sodium Channel Subtypes in Rat Brain Neurons

Abstract

Several scorpion toxins have been shown to exert their neurotoxic effects by a direct interaction with voltage-dependent sodium channels. Both classical scorpion alpha-toxins such as Lqh II from Leiurus quiquestratus hebraeus and alpha-like toxins as toxin III from the same scorpion (Lqh III) competitively interact for binding on receptor site 3 of insect sodium channels. Conversely, Lqh III, which is highly toxic in mammalian brain, reveals no specific binding to sodium channels of rat brain synaptosomes and displaces the binding of Lqh II only at high concentration. The contrast between the low-affinity interaction and the high toxicity of Lqh III indicates that Lqh III binding sites distinct from those present in synaptosomes must exist in the brain. In agreement, electrophysiological experiments performed on acute rat hippocampal slices revealed that Lqh III strongly affects the inactivation of voltage-gated sodium channels recorded either in current or voltage clamp, whereas Lqh II had weak, or no, effects. In contrast, Lqh III had no effect on cultured embryonic chick central neurons and on sodium channels from rat brain IIA and beta1 subunits reconstituted in Xenopus oocytes, whereas sea anemone toxin ATXII and Lqh II were very active. These data indicate that the alpha-like toxin Lqh III displays a surprising subtype specificity, reveals the presence of a new, distinct sodium channel insensitive to Lqh II, and highlights the differences in distribution of channel expression in the CNS. This toxin may constitute a valuable tool for the investigation of mammalian brain function.

Molecular Basis of the High Insecticidal Potency of Scorpion α-Toxins

“…One interaction is between the conserved Core-domain and a channel region that may be conserved among insects and mammals. This interaction may enable recognition of receptor site-3 by all toxins of the ␣-group and could explain the ability of anti-mammalian ␣-toxins to displace at high concentrations ␣-insect toxins from their binding site on the insect channel (9,47,48). The second interaction is between the variable NCdomain and another channel region.…”

“…The core of Aah2 Lqh␣IT(face) is made of an ␣-helix (residues 19 -28) packed against a threestranded anti-parallel ␤-sheet (residues 2-4, 32-37, and [45][46][47][48][49][50][51] and is very similar to that of Aah2 (Fig. 4A) (38).…”

Combinatorial Interaction of Scorpion Toxins Lqh-2, Lqh-3, and LqhαIT with Sodium Channel Receptor Sites-3

“…␣-Toxins inhibit Na V channel inactivation in various excitable preparations, but they show vast differences in preference for insect and mammalian Na V channels. Accordingly, they are divided into classic ␣-toxins, which are highly active in mammalian brain [e.g., Lqh-2 (Leiurus quinquestriatus hebraeus)]; ␣-toxins, which are very active in insects (e.g., Lqh␣IT); and ␣-like toxins, which are active in both the mammalian and insect central nervous system (CNS) (e.g., Lqh-3) (Gordon et al, 1996Sautiere et al, 1998;Gilles et al, 1999Gilles et al, , 2000aKrimm et al, 1999).…”

“…For example, Lqh␣IT is ϳ2000-fold less active in mouse brain than Lqh-2 and competes only at micromolar concentrations for binding to rat brain synaptosomes. The ␣-like toxin, Lqh-3, is highly toxic in the mouse brain but is a very weak competitor for Lqh-2 binding to rat brain synaptosomes (Gordon et al, 1996Gilles et al, 1999Gilles et al, , 2000a. Moreover, Lqh-3 and Lqh␣IT barely affect the Na ϩ current mediated by the heterologously expressed rNa v 1.2 in contrast to the strong inhibition of inactivation induced by Lqh-2 (Gilles et al, , 2000a(Gilles et al, , 2001.…”

“…␣-Toxins inhibit Na V channel inactivation in various excitable preparations, but they show vast differences in preference for insect and mammalian Na V channels. Accordingly, they are divided into classic ␣-toxins, which are highly active in mammalian brain [e.g., Lqh-2 (Leiurus quinquestriatus hebraeus)]; ␣-toxins, which are very active in insects (e.g., Lqh␣IT); and ␣-like toxins, which are active in both the mammalian and insect central nervous system (CNS) (e.g., Lqh-3) (Gordon et al, 1996Sautiere et al, 1998;Gilles et al, 1999Gilles et al, , 2000aKrimm et al, 1999).Despite differences in toxicity and binding properties, all scorpion ␣-toxins bind to receptor site 3, the structural features of which are still elusive, but known to involve the extracellular loops S5-S6 of D1 and D4 (Tejedor and Catterall, 1988;Thomson and Catterall, 1989) and S3-S4 of D4 of rat Na V 1.2 (Catterall, 1992(Catterall, , 2000Rogers et al, 1996). Mutagenesis within the external linker S3-S4 in domain 4 of rat brain Na V 1.2 identified a negatively charged residue, Glu1613, as a major determinant that affects the binding of a classic scorpion ␣-toxin (Rogers et al, 1996).…”