Effect of ANEPIII, a novel recombinant neurotoxic polypeptide, on sodium channels in primary cultured rat hippocampal and cortical neurons.
ABSTRACT Previous studies have shown that the recombinant neurotoxic polypeptide BmK ANEP (ANEPIII) displayed good anti-neuroexcitation activity as demonstrated by pharmacological tests of the blockade of chemical-induced convulsive seizures. In order to search for further anticonvulsant mechanism of action of ANEPIII, the effects of ANEPIII on sodium channels were assessed using the whole-cell patch clamp recordings in primary cultures of rat hippocampal and cortical neurons. ANEPIII decreased the sodium currents in a voltage-dependent manner, which appeared as a shift of the current-voltage relation to positive potentials. The effect was reversible after washing. The concentration-responsiveness measured in hippocampal and cortical neurons revealed an IC(50) value of 124.6 nM and 192.7 nM, respectively. Furthermore, ANEPIII 1000 nM significantly shifted the activation curves of sodium current in hippocampal and cortical neurons to more positive potentials and the recovery from inactivation of sodium current was significantly slower. Voltage-dependent inactivation curves of sodium channels in hippocampal and cortical neurons did not change in the presence of 1000 nM ANEPIII. Thus, our results demonstrated that ANEPIII in submicromolar concentrations was a voltage-dependent, reversible blocker of sodium current in hippocampal and cortical neurons. It is concluded that these phenomena may explain, at least in part, the anti-neuroexciting properties of this peptide.
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ABSTRACT: The effect of Radix paeoniae rubra (RPR) on voltage-gated sodium channel (VGSC) currents (I(Na)) was examined in freshly isolated rat hippocampal CA1 neurons using whole-cell patch-clamp technique under voltage-clamp conditions. RPR suppressed I(Na) without affecting the current activation, inactivation and deactivation. The amplitude of I(Na) decreased by approximately 18.4% within a few seconds of 0.8 mg/ml RPR exposure. RPR (0.8 mg/ml) shifted the steady-state inactivation curves of I(Na) to negative potentials, with hyperpolarizing direction shift of V(1/2) of 10.0 mV. The time course of I(Na) recovery from inactivation was prolonged significantly by 0.8 mg/ml RPR. RPR (0.8 mg/ml) also enhanced the activity-dependent attenuation of I(Na) and decreased the fraction of activated channels. These results suggested that RPR suppressed hippocampal CA1 I(Na) by shifting the inactivation curve in hyperpolarizing direction, slowing the recovery time course from inactivation, enhancing the activity-dependent attenuation and decreasing the number of activatable channels. RPR suppression on I(Na) might predict the protective effect during brain ischemia in hippocampal CA1 neurons.Brain Research 07/2002; 940(1-2):1-9. · 2.88 Impact Factor
- Toxin Reviews - TOXIN REV. 01/1998; 17(2):131-159.
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ABSTRACT: Most scorpion toxins are ligand peptides that recognize and bind to integral membrane proteins known as ion-channels. To date there are at least 202 distinct sequences described, obtained from 30 different species of scorpions, 27 from the family Buthidae and three from the family Scorpionidae. Toxins that recognize potassium and chloride channels are usually from 29 to 41 amino acids long, stabilized by three or four disulfide bridges, whereas those that recognize sodium channels are longer, 60 to 76 amino acid residues, compacted by four disulfide bridges. Toxins specific for calcium channels are scarcely known and have variable amino acid lengths. The entire repertoire of toxins, independently of their specificity, was analyzed together by computational programs and a phylogenetic tree was built showing two separate branches. The K(+) and Cl(-) channel specific toxins are clustered into 14 subfamilies, whereas those of Na(+) and Ca(2+) specific toxins comprise at least 12 subfamilies. There are clear similarities among them, both in terms of primary sequence and the main three-dimensional folding pattern. A dense core formed by a short alpha helix segment and several antiparallel beta-sheet stretches, maintained by disulfide pairing, seems to be a common structural feature present in all toxins. The physiological function of these peptides is manifested by a blockage of ion passage through the channels or by a modification of the gating mechanism that controls opening and closing of the ion pore.Biochimie 01/2000; 82(9-10):861-8. · 3.14 Impact Factor