Effects of Tityus serrulatus scorpion toxin gamma on voltage-gated Na+ channels

The effects of Brazilian scorpion Tityus serrulatus toxin gamma (TiTx gamma) were studied on voltage-gated Na+ channels from human heart (hHl) and rat skeletal muscle (rSkM1). The Na+ channels were expressed in Xenopus laevis oocytes, and Na+ currents were recorded using two-microelectrode voltage-clamp techniques. In control experiments, the threshold of activation of hH1 is more negative than that of rSkM1 by approximately 20 mV. The toxin induces a shift of the voltage dependence of activation toward more negative potential values and reduces the amplitude of the current when administered to rSkM1. In contrast, TiTx gamma has little discernible effect on the current-voltage curve for hH1 at 100 nmol/L. Chimeric channels formed from these two isoforms were constructed to localize the binding site of TiTx gamma on rSkM1. TiTx gamma shifts the activation of a chimera (SSHH) in which domains 1 (D1) and 2 (D2) derive from rSkM1 and domain 3(D3) and 4 (D4) derive from hH1. This finding suggests that the toxin acts on the activation of rSkM1 by binding either to D1 and/or D2. TiTx gamma shifted the activation of another chimera with D2-D3-D4 from rSkM1 (HSSS) toward more hyperpolarizing potentials and had no effect on the activation of other chimeras with only D1-D3-D4 from rSkM1 (SHSS) or only D3 from rSkM1 (HHSH). Finally, a chimera in which D2 is from rSkM1 and all others domains are from hH1 (HSHH) provides further compelling support for our hypothesis. TiTx gamma shifts the activation of this chimera toward more negative potential values. Thus, TiTx gamma action on chimeras segregates with the source of D2: when D2 is from rSkM1, the toxin affects activation. We infer that D2 plays an important role in the activation process of voltage-gated Na+ channels.