Biomedical subjects

C Paillart

Publications and source records attributed to C Paillart.

4 recordsLinked to original sources

PubMed1999

Specific distribution of sodium channels in axons of rat embryo spinal motoneurones.

1. The distribution of Na+ channels and development of excitability were investigated in vitro in purified spinal motoneurones obtained from rat embryos at E14, using electrophysiological, immunocytochemical and autoradiographical methods. 2. One hour after plating the motoneurones (DIV0), only somas were present. They expressed a robust delayed rectifier K+ current (IDR) and a fast-inactivating A-type K+ current (IA). The rapid neuritic outgrowth was paralleled by the emergence of a fast-activating TTX-sensitive sodium current (INa), and by an increase in both K+ currents. 3. The change in the three currents was measured daily, up to DIV8. The large increase in INa observed after DIV2 was accompanied by the onset of excitability. Spontaneous activity was observed as from DIV6. 4. The occurrence of axonal differentiation was confirmed by the fact that (i) only one neurite per motoneurone generated antidromic action potentials; and (ii) 125I-alpha-scorpion toxin binding, a specific marker of Na+ channels, labelled only one neurite and the greatest density was observed in the initial segment. Na+ channels therefore selectively targeted the axon and were absent from the dendrites and somas. 5. The specific distribution of Na+ channels was detectable as soon as the neurites began to grow. When the neuritic outgrowth was blocked by nocodazole, no INa developed. 6. It was concluded that, in spinal embryonic motoneurone in cell culture, Na+ channels, the expression of which starts with neuritic differentiation, are selectively addressed to the axonal process, whereas K+ channels are present in the soma prior to the neuritic outgrowth.

Animals↗

PubMed1997

Direct block of voltage-sensitive sodium channels by genistein, a tyrosine kinase inhibitor.

Genistein, an isoflavone inhibitor of tyrosine-specific protein kinases, was shown to specifically block the 22Na+ influx through voltage-sensitive Na+ channels in cultured rat brain neurons, whereas other tyrosine kinase antagonists such as lavendustin A, compound 5, tyrphostin A47 and an erbstatin analog were inactive at concentrations known to block kinase activity in other neuronal systems. Dose-response curves for genistein indicated a half-maximum effect at 60 microM. Daidzein, an inactive analog of genistein, had a similar inhibitory effect on the 22Na+ influx with a half-maximum effect at 195 microM. The time course of genistein action was rapid, because maximum effect on 22Na+ influx was obtained in less than 20 s at 100 microM. Analysis of Na+ currents by the whole-cell recording technique showed that 20 microM genistein reduced the sodium current and shifted the voltage dependence of both activation and inactivation curves. No competition with [3H]saxitoxin binding was observed, whereas the binding of [3H]batrachotoxinin A 20-alpha-benzoate to rat brain synaptosomal membranes was partially inhibited, which suggested a direct or allosteric interaction with neurotoxin binding site 2. These data taken together clearly indicate that the inhibition of voltage-sensitive sodium channels by genistein is not mediated by tyrosine kinase inhibition.

Animals↗

PubMed1996

Activity-induced internalization and rapid degradation of sodium channels in cultured fetal neurons.

A regulatory mechanism for neuronal excitability consists in controlling sodium channel density at the plasma membrane. In cultured fetal neurons, activation of sodium channels by neurotoxins, e.g., veratridine and alpha-scorpion toxin (alpha-ScTx) that enhance the channel open state probability induced a rapid down-regulation of surface channels. Evidence that the initial step of activity-induced sodium channel down-regulation is mediated by internalization was provided by using 125I-alpha-ScTx as both a channel probe and activator. After its binding to surface channels, the distribution of 125I-alpha-ScTx into five subcellular compartments was quantitatively analyzed by EM autoradiography. 125I-alpha-ScTx was found to accumulate in tubulovesicular endosomes and disappear from the cell surface in a time-dependent manner. This specific distribution was prevented by addition of tetrodotoxin (TTX), a channel blocker. By using a photoreactive derivative to covalently label sodium channels at the surface of cultured neurons, we further demonstrated that they are degraded after veratridine-induced internalization. A time-dependent decrease in the amount of labeled sodium channel alpha subunit was observed after veratridine treatment. After 120 min of incubation, half of the alpha subunits were cleaved. This degradation was prevented totally by TTX addition and was accompanied by the appearance of an increasing amount of a 90-kD major proteolytic fragment that was already detected after 45-60 min of veratridine treatment. Exposure of the photoaffinity-labeled cells to amphotericin B, a sodium ionophore, gave similar results. In this case, degradation was prevented when Na+ ions were substituted by choline ions and not blocked by TTX. After veratridine- or amphotericin B-induced internalization of sodium channels, breakdown of the labeled alpha subunit was inhibited by leupeptin, while internalization was almost unaffected. Thus, cultured fetal neurons are capable of adjusting sodium channel density by an activity-dependent endocytotic process that is triggered by Na+ influx.

Amphotericin B↗

PubMed1994

Sodium channel internalization in developing neurons.

Neurotoxin-induced activation of voltage-dependent Na+ channels provoked rapid (t1/2 = 15-20 min) channel down-regulation in cultured rat brain neurons, resulting in a 50%-70% decrease in [3H]saxitoxin and 125I-alpha-scorpion toxin binding capacities as well as a decrease in Na+ peak current. Experiments using 125I-alpha-scorpion toxin as both a Na+ channel activator and a surface channel probe showed that a fraction of the bound toxin was internalized, since it was not releasable by acidic washing. Internalization was inhibited by tetrodotoxin, abolished in Na(+)-free medium, and induced by amphotericin B, a Na+ ionophore. Moreover, down-regulation occurred only in immature neuronal tissue, either cultured fetal neurons or postnatal hippocampal slices, but was absent in adult brain. These observations indicate that Na+ channel internalization is triggered by Na+ influx into neurons and may be involved in the control of electrical activity during development.

Aging↗