Inhibition of ³H-GABA and ³H-Dopamine uptake into rat brain synaptosomes by Tityusserrulatus venom and its toxin TsTX-V

Cecchini, a.l.(1,2,3), vasconcelos, f.(1), giglio, j.r.(2), amara, s.g.(3), arantes, e.c.(1)

(1)Departamento de Física e Química, FCFRP-USP, (2)Departamento de Bioquímica e Imunologia, FMRP, USP, Ribeirão Preto, SP, Brazil, (3)Vollum Institute, HHMI, Oregon Health and Science University, Portland, Oregon, USA.

Objectives: Scorpion venoms contain toxins that have well-established actions on the function of ionic channels of excitable cells.  However, the effects of these toxins on neurotransmitter transport have not yet been examined.  This study evaluates the effects of Tityus serrulatus scorpion venom (TsV) and TsTX-V, an a-toxin, on GABA and dopamine (DA) uptake in isolated rat brain synaptosomes.

METHODS AND RESULTS: TsTX-V was purified from TsV by a combination of ion exchange chromatography and reverse phase HPLC. Homogeneity of TsTX-V was evidenced by a single band by PAGE, SDS-PAGE (Mr=7230) and isoelectric focusing (pI=8.0). Both the venom and its toxin were assayed for their effects on ³H-GABA uptake into cortical synaptosomes and on ³H-DA uptake into striatal synaptosomes.  TsV(0.43ng/mL) inhibited both GABA and DA uptake (~50%).  TsTX-V showed IC₅₀s= 9.37nM and 22.2nM for GABA and DA uptake, respectively. In contrast, maximal concentrations of TsV(1300ng/mL) and TsTX-V(2770nM) had no effect on ³H-L-glutamate uptake.  The inhibitory effects of TsV and TsTX-V were still observed when external Ca²⁺ was replaced by Sr²⁺⁽7.6mM) and EGTA(1mM), but not when synaptosomes were pre-treated with TTX(1mM).  TsTX-V did not inhibit ³H-GABA uptake in COS-7 cells expressing GAT-1 or GAT-3, the major brain GABA transporters, suggesting that the toxin indirectly inhibits transport.

Conclusions: TsVand TsTX-V appear not to affect transporters expressed in heterologous expression systems, including COS-7 cells, but can inhibit GABA and DA transport activity in a Ca²⁺-independent manner in native systems such as synaptosomes.

Suupportedby: Fapesp, Capes 

CORRESPONDENCE TO:

Eliane Candiani Arantes, Avenida do Café s/n, Departamento de Física e Química, FCFRP, USP, Ribeirão Preto, SP, CEP: 14040-903, Brasil, Email: ecabraga@fcfrp.usp.br