| Budget Amount *help |
¥3,710,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥210,000)
Fiscal Year 2007: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2006: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 2005: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
In the first study, we assessed local anesthetics toxicity cell biologically. The local anesthetics have direct neurotoxicity and induce growth cone collapse when applied to neurons at large concentrations. However, the effects of prolonged exposure to local anesthetics at a small concentration have never been studied. We examined whether neurite growth was slowed by tetracaine at small concentrations in chick embryo dorsal root ganglions. The effects of tetracaine were examined microscopically and by a neurite growth rate assay, quantitative morphologic assay, growth cone collapse assay, and Western blot assay. Neurite growth 24 and 48 h after application was delayed significantly when tetracaine was applied at a concentration larger than 5 microM. Filopodia of growth cones retracted, and their number was significantly decreased 24 and 48 h after the application of 10 and 20 microM of tetracaine. The quantity of actin in cell bodies increased, contrary to the effect on neurites and gr
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owth cones, where actin decreased 48 h after the application of 5, 10, and 20 microM of tetracaine. In conclusion, continuous exposure to tetracaine at small concentrations delayed neurite growth, reduced the number of filopodia, and decreased actin content.
The second study was undertaken to determine whether propofol has neurotoxic effects on peripheral, retinal, and autonomic neurons, and which neurons are particularly liable to injury by propofol. Dorsal root ganglia, retinal ganglion cell layers, and sympathetic ganglion chains were isolated from day eight chick embryos and cultured for 20 hr. Thereafter, propofol was added at various concentrations [5-300 microM (0.9-53 microg x mL (-1) )] to investigate its effects on these three types of neuronal tissue. Morphological changes were examined quantitatively by growth cone collapse assay. Propofol concentrations were measured using high performance liquid chromatography. Propofol induced growth cone collapse and neurite destruction. The three types of neurons tested exhibited significantly different dose-response relationships two hours after the application of propofol (P < 0.001) but not at 24 hr after application. The growth cone-collapsing effect was at least partially reversible in all three types of neurons after exposure to 100 microM propofol up to six hours, though reversibility was not observed after 24-hr exposure. While the clinical safety profile of propofol has been well documented, at high concentrations propofol has potential neurotoxicity on growing neurons in vitro.
From these observations, we concluded nerve growth cones and neurites are sensitive outer environment. Some externally applied trophic factors and/or cells that release trophic factors, may be effective to promote nerve regeneration. Less
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