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  • Title: B103 neuroblastoma cells predominantly express endothelin ET(B) receptor; effects of extracellular Ca(2+) influx on endothelin-1-induced mitogenesis.
    Author: Kawanabe Y, Hashimoto N, Masaki T.
    Journal: Eur J Pharmacol; 2001 Aug 17; 425(3):173-9. PubMed ID: 11513835.
    Abstract:
    We sought to examine the effects of endothelin-1 on the intracellular free Ca(2+) concentration ([Ca(2+)](i)) and mitogenic response in the neuroblastoma cell line, B103 (B103 cells). The results obtained from an [125I] endothelin-1 binding assay demonstrated that B103 cells express the endothelin receptor. The B(max) and K(d) values for [125I]endothelin-1 binding were 70+/-36 fmol/mg protein and 52+/-13 pM, respectively. Endothelin-1 failed to stimulate cAMP formation, but it did inhibit forskolin-induced cAMP formation. Endothelin-1 also stimulated the accumulation of [3H]inositol phosphates. These results indicate that the endothelin receptor in B103 cells couples with G(i) and G(q) but not with G(s). Monitoring of [Ca(2+)](i) showed that endothelin-1 evoked a transient increase in [Ca(2+)](i); this remained even in the absence of extracellular Ca(2+). However, no sustained, endothelin-1-induced increase in [Ca(2+)](i) due to extracellular Ca(2+) influx was detected. The endothelin B receptor-selective antagonist, 2,6-Dimethylpiperidinecarbonyl-gamma-Methyl-Leu-N(in)-[Methoxycarbonyl]-D-Trp-D-Nle (BQ 788), abolished the endothelin-1-induced increase in [Ca(2+)](i), while the endothelin ET(A) receptor-selective antagonist, cyclo-D-Asp-Pro-D-Val-Leu-D-Trp (BQ 123), failed to inhibit it. These results indicate that B103 cells express endothelin ET(B) receptor or an endothelin ET(B)-like receptor predominantly and have no Ca(2+) channels activated by endothelin-1. Endothelin-1 activated mitogen-activated protein kinase in B103 cells. However, based on the data for 3-(4,5-dimethy-2-thiazolyl)-2,5-diphenyl tetrazolium bromide, [3H]thymidine incorporation, and apoptosis screening assays, endothelin-1 induces neither mitogenesis nor apoptosis. These results suggest that endothelin-1 has no role in the mitogenic response in B103 cells, and this is consistent with the notion that an endothelin-1-induced sustained increase in [Ca(2+)](i) plays a role in endothelin-1-induced cell proliferation.
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