Characterization of nicotinic receptors in immortalized hippocampal neurons

Nicotinic acetylcholine receptors, particularly nicotinic a-bungarotoxin (a-BGT) receptors, are present in relatively high concentrations in rat hippocampus. Because of the difficulties encountered in studying receptors using primary cells in culture, especially for biochemical work, we investigated the possibility of using an immortalized cell line from embryonic rat hippocampus (H19-7). RNase protection assays show that α4, α7 and β2 neuronal nicotinic receptor subunit mRNAs are present in differentiated but not undifferentiated H19-7 cells, while α2, α3, α5 and β3 subunit mRNAs were not detectable under either condition. In line with these results, the present data demonstrate that the H19-7 cells express cell surface nicotinic a-BGT binding sites, which were maximal after seven days of differentiation in culture. The receptors were saturable, of high affinity (K_d = 1.30 nM and B_max = 11.70 fmol/10⁵ cells) and had a pharmacological profile similar to that observed for CNS a-BGT receptors. On the other hand, although α4 and β2 neuronal nicotinic subunit mRNAs were present in differentiated H19-7 cells, no 3H]cytisine binding was observed. Because immortalized cell lines have the advantage that they provide a limitless supply of cells as compared to primary cell cultures, but yet are not malignant in origin, the present results may suggest that the H19-7 immortalized hippocampul cell line represent a useful CNS model system for examining α-BGT nicotinic receptors.