Experiments with Radiolabeled Toxins and Radioactive Isotopes

Experiments with Radiolabeled Toxins and Radioactive Isotopes 

The experiments underlying the site definition involved either the binding of radiolabeled toxins or the displacement of such toxins. As for site-2 toxins the binding of [ H]veratridine is unsatisfactory due to its comparatively low affinity and the high nonspecific binding to membrane lipids 

In many papers neurotoxin effects on cells, synaptosomes or lipid vesicles containing purified channels were studied by means of Na+ uptake. Recently the intravesicular increase in Na concentration has also been monitored with fluorescent indicators (Daniell 1992 Deri and Adma-Vizi 

in synaptosomes of rainbow trout brain aconitine is clearly more potent than veratridine K0.5 of 7.7 vs 66 XM) and maximum uptake, likewise, is 76% vs 29% of that in batrachotoxin (Stuart et al. 1987). Another deviating result has been reported for purified channels from rat brain, reconstituted into phosphatidylcholine vesicles where batrachotoxin activates a smaller fraction of channels than veratridine, and Kd for the latter is larger (30 pM) than in native channels (Tamkun et al. 1984) a similarly high value (Ko.5=35 pM) is observed with purified channels from rat sar-colemma (Weigele and Barchi 1982). In N18 cells neurotoxin-induced activation is inhibited by divalent cations, with Mn being most effective and Sr least effective. The inhibition constants for Ca are 0.84 mM and 

2 mM (veratridine- and batrachotoxin-dependent uptake Catterall 1975a). In a rat brain preparation stimulated by veratridine 55 pM Ca has been shown to be sufficient for reducing uptake by one-half (Matthews et al. 1981). Na+ uptake through several types of tetrodotoxin-sensitive channels activated by veratridine is also inhibited by Zn + and Cd + with a half-inhibitory concentration (IC50) of ca. 2 and 5 mM, respectively, whereas the IC50 of tetrodotoxin-resistant channels, for example, from heart cells, is much lower 50 pM and 0.2 mM (Frelin et al. 1986). 

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