Ultrastructural localization of voltage-sensitive sodium channels using [125I]α scorpion toxin

Cau, Pierre; Massacrier, Annick; Boudier, Jean-Louis; Couraud, F.; Bottini, Janine; Deprez, Paule

doi:10.1016/0006-8993(85)90561-x

Cited by 13 publications

(4 citation statements)

References 33 publications

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“…The deploym ent, spatial d istrib u tio n and localization of sodium channels w ithin neural tissue have been inferred largely from biochemical, physiological and p h a r macological studies, which are n o t able to furnish detailed u ltrastru ctu ra l local ization of the channels. Radio-and fluorescently labelled scorpion toxins have been used as m arkers of sodium channel localization in cultured neurons (C atterall 1981;B oudier et al 1985 ;Cau et al 1985 ;Angelides et al 1988) and in vitro tissue (D ellm ann et al 1983). The high-affinity binding of scorpion toxins is voltage dependent and is appreciably decreased by m em brane depolarization (C atterall 1980), obviating th e ir use w ith aldehyde-fixed tissue.…”

Section: Introductionmentioning

confidence: 99%

Immuno-ultrastructural localization of sodium channels at nodes of Ranvier and perinodal astrocytes in rat optic nerve

Black

Friedman

Waxman

et al. 1989

Proc. R. Soc. Lond. B

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Immuno-electron microscopic localization of sodium channels at nodes of Ranvier within adult optic nerve was demonstrated with polyclonal antibody 7493. The 7493 antisera, which is directed against purified sodium channels from rat brain, recognizes a 260 kDa protein in immunoblots of the crude glycoprotein fraction from adult rat optic nerve. Intense immunoreactivity with 7493 antisera was observed at nodes of Ranvier. Axon membrane at the node was densely stained, whereas paranodal and internodal axon membrane did not exhibit immunoreactivity. The axoplasm beneath the nodal membrane displayed variable immunostaining. Neither terminal paranodal oligodendroglial loops nor oligodendrocyte plasmalemma were immunoreactive with 7493 antisera. However, perinodal astrocyte processes exhibited intense immunoreactivity with the anti-sodium channel antisera. Optic nerves incubated with pre-immune sera, or with 7493 antisera that had been pre-adsorbed with purified sodium channel protein, displayed no immunoreactivity. These results demonstrate localization of sodium channels at high density at mammalian nodes of Ranvier and in some perinodal astrocyte processes. The latter observation offers support for an active role for perinodal astrocyte processes in the aggregation of sodium channels within the axon membrane at the node of Ranvier.

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Section: Introductionmentioning

confidence: 99%

Immuno-ultrastructural localization of sodium channels at nodes of Ranvier and perinodal astrocytes in rat optic nerve

Black

Friedman

Waxman

et al. 1989

Proc. R. Soc. Lond. B

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“…Scorpion toxins can also be used as markers for localization of the voltage-sensitive sodium channels, owing to the following properties: (1) the nonspecific binding is relatively low; (2) they can be labeled whith lz51 to a high specific activity, allowing their use for autoradiographic experiments; (3) because of their polypeptide nature, they can be cross-linked to proteins on their binding site by the usual microscopic fixatives. The autoradiographic localization of voltage-sensitive sodium channels using scorpion toxin as a marker has already been carried out on cultured nerve cells (Catterall, 198 1;Boudier et al, 1985;Cau et al, 1985) and on tissue incubated in vitro (Dellmann et al, 1983).…”

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confidence: 99%

Autoradiographic localization of voltage-dependent sodium channels on the mouse neuromuscular junction using 125I-alpha scorpion toxin. I. Preferential labeling of glial cells on the presynaptic side

1988

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Alpha-scorpion toxins bind specifically to the voltage-sensitive sodium channel in excitable membranes, and binding is potential-dependent (Catterall, 1984). The radioiodinated toxin II from the scorpion Androctonus australis Hector (alpha ScTx) was used to localize voltage-sensitive sodium channels on the presynaptic side of mouse neuromuscular junctions (NMJ) by autoradiography using both light and electron microscopy. Silver grain localization was analyzed by the cross-fire method. At the light-microscopic level, grain density over NMJ appeared 6-8x higher than over nonjunctional muscle membrane. The specificity of labeling was verified by competition/displacement with an excess of native alpha ScTx. Labeling was also inhibited by incubation in depolarizing conditions, showing its potential-dependence. At the electron-microscopic level, analysis showed that voltage-sensitive sodium channels labeled with alpha ScTx were almost exclusively localized on membranes, as expected. Due to washout after incubation, appreciable numbers of binding sites were not found on the postsynaptic membranes. However, on the presynaptic side, alpha ScTx-labeled voltage-sensitive sodium channels were localized on the membrane of non-myelin-forming Schwann cells covering NMJ. The axonal presynaptic membrane was not labeled. These results show that voltage-sensitive sodium channels are present on glial cells in vivo, as already demonstrated in vitro (Chiu et al., 1984; Schrager et al., 1985). It is proposed that these glial channels could be indirectly involved in the ionic homeostasis of the axonal environment.

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“…Among them, radioiodinated scorpion toxins have been used for autoradiographic localization of Na+ channels on nerve cells Cau et al, 1985) and muscle tissue (Boudier et al, 1988;Le Treut et al, 1990). At least five different neurotoxin-binding sites have been described on the Na+ channel (Catterall, 1980;Couraud et al, 1982;Poli et al, 1986).…”

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Autoradiographic localization of voltage-dependent sodium channels on the mouse neuromuscular junction using 125I-alpha scorpion toxin. II. Sodium distribution on postsynaptic membranes

1992

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A radioiodinated alpha-scorpion toxin (toxin II from Androctonus australis Hector) (alpha ScTx) was used as a probe for EM autoradiography to study the distribution of voltage-dependent sodium channels (Na+ channel) on the postsynaptic side of the mouse neuromuscular junction. Silver grain distribution was analyzed by the cross-fire method to assess the relative Na+ channel density in each membrane domain measured by stereology. This analysis showed that the maximum Na+ channel density was located on the edge of the synaptic gutter, where it reached about twice the mean density in the postsynaptic fold membrane. Na+ channel densities have been calculated using ACh receptor (AChR) density in fold crests as reference. Sodium channel density on the edge of the synaptic gutter was estimated at about 5000/microns 2. Sodium channel distribution in the postsynaptic folds was compared to AChR distribution using density distribution analysis (Fertuck and Salpeter, 1976). The results confirmed that, as already observed by immunogold labeling (Flucher and Daniels, 1989), there are no Na+ channels on fold crests. Na+ channels are located in the rest of the fold membrane (bottom) and may be distributed according to two possible models. In the first, density would be uniformly high, although lower than on the gutter edge. In the second, density would decrease from the crest border, where the value was that of the gutter edge, to the fold end, where the value would be 50% lower. Based on the latter model, which was the “best-fit model,” we propose that the postsynaptic membrane includes two domains. The first is the fold crest, which contains almost exclusively AChRs. This domain is devoted to reception-transduction of the chemical signal. The second includes both the fold bottom membrane and the perisynaptic membrane. Sodium channel density is highest along the crest border and decreases moving away. Its functions are the integration of postsynaptic potentials and generation-conduction of the muscle action potential.

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Ultrastructural localization of voltage-sensitive sodium channels using [125I]α scorpion toxin

Cited by 13 publications

References 33 publications

Immuno-ultrastructural localization of sodium channels at nodes of Ranvier and perinodal astrocytes in rat optic nerve

Immuno-ultrastructural localization of sodium channels at nodes of Ranvier and perinodal astrocytes in rat optic nerve

Autoradiographic localization of voltage-dependent sodium channels on the mouse neuromuscular junction using 125I-alpha scorpion toxin. I. Preferential labeling of glial cells on the presynaptic side

Autoradiographic localization of voltage-dependent sodium channels on the mouse neuromuscular junction using 125I-alpha scorpion toxin. II. Sodium distribution on postsynaptic membranes

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