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Basic fibroblast growth factor induction of neuronal ion channel expression in ascidian ectodermal blastomeres

Takeshi Inazawa *, Yasushi Okamura ¹ and Kunitaro Takahashi ²

1. Cleavage-arrested anterior animal (a4-2) blastomeres isolated from eight-cell embryos of Halocynthia aurantium differentiated into neuronal type cells expressing neuron-specific ion channels when they were treated with basic fibroblast growth factor (bFGF). This induction process was very similar to that when a4-2 blastomeres were cultured in contact with anterior vegetal (A4-1) blastomeres from the same embryos or when treated with subtilisin, a serine protease.

2. Other growth factors, transforming growth factor (TGF) 1, activin A, epidermal growth factor (EGF) and nerve growth factor (NGF), had no effect on the default epidermal differentiation of cleavage-arrested a4-2 blastomeres.

3. Messenger RNA of the ascidian neuronal Na⁺ channel, TuNa I, was detected using RT-PCR in a4-2-derived partial embryos of Halocynthia aurantium as well as in the cleavage-arrested a4-2 blastomeres treated with bFGF, confirming the neural inducer activity of bFGF during ascidian embryogenesis.

4. bFGF was effective at concentrations as low as 1 ng ml^-1 in inducing neuronal ion channels in cleavage-arrested a4-2 blastomeres. EC₅₀ for neuronal differentiation was estimated to be around 8 ng ml^-1, and the maximum effect of 90 % neuronalization was obtained with above 100 ng ml^-1.

5. For induction of neuronal differentiation, bFGF was required to be continuously present 8 to 14 h after fertilization. A similar time window was required for cell-contact induction, but it was considerably shorter for subtilisin induction.

6. We discuss whether activation of receptor tyrosine kinase is a common pathway for neural induction by bFGF, subtilisin, and cell-contact with A4-1 blastomeres.

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