Effects of endothelin (ET) family peptides and their derivatives on cellular contraction and calcium-ion level were examined by using cultured human vascular smooth muscle cells (VSM). Contraction of cultured human VSM, isolated from human fetal aortic segments, was induced within 1 min after the treatment with ET-1 (100 nM) as seen in the changes of cytosolic calcium-ion localization. In parallel with the cell contraction, cytosolic calcium-ion level in the human VSM increased very rapidly and then dropped with some oscillation as determined by Anchorage Cell Analyzing System. It was noted that transient calcium-ion mobilization rather than sustained calcium-ion influx was significant in the contraction of cultured human VSM. Vasoactive intestinal contractor (VIC), three amino acids different from ET-1, had less activity in increase of intracellular calcium-ion level and in percent of response cells than ET-1, ET-2, and VIC-S4L6 (one amino acid different from ET-1). EC50 of ET-1, VIC-S4L6, ET-2, and VIC were 0.5 nM, 0.6 nM, 2.0 nM, and 20 nM, respectively. VIC-like peptide (VIC-LP), 16 amino acids fragment of VIC precursor protein, had no effect with a single administration of up to 10 mu M. However, the increase in calcium-ion level by VIC was suppressed with a prior treatment of cells with high concentration (10 mu M) of VIC-LP. The establishment of cultured human VSM for the simultaneous examination of the contraction and calcium-ion level will provide a new system to study signal transduction of vasocontractor peptides.
