Abbreviations used:α-BTx, α-bungarotoxin; MLA, methyllycaconitine; nAChR, nicotinic acetylcholine receptor; RT, reverse transcription; SCLC, small cell lung carcinoma; d-TC, d-tubocurarine.
Abstract: SCC-37 is a small cell lung carcinoma line that aberrantly expresses muscle-type nicotinic acetylcholine receptors (nAChRs). It was established from a patient with a paraneoplastic autoimmune neuromuscular disorder, myasthenia gravis. When grown as a xenograft tumor, SCC-37 cells express plasma membrane receptors that bind 125I-labeled α-bungarotoxin (125I-α-BTx), cosediment with 9S nAChR pentamers, and bind to a monoclonal antibody (MAb 35) specific for muscle-type (α1 subunit) α-BTx receptors. The agonist carbamylcholine (carbachol) stimulates influx of 22Na+ in SCC-37 cells; this is inhibited by α-BTx and by d-tubocurarine. Long-term cultured SCC-37 cells have functional and ligand-binding evidence for surface coexpression of both α1 and neuronal-type (α7 subunit) α-BTx receptors. A subclone of SCC-37, designated SCC-A9, expresses only the neuronal-type (α7 subunit) α-BTx receptors on its surface. Carbachol does not stimulate 22Na+ influx in SCC-A9 cells, but cytisine initiates a sustained influx of Ca2+. Activation of this response is inhibited by α-BTx and by the α7-selective antagonist methyllycaconitine. Addition of Co2+ abrogates the sustained elevation of intracellular free Ca2+ concentration, implying that the cytisine-stimulated influx of Ca2+ is sustained by secondary opening of voltage-sensitive channels in the plasma membrane. Surface receptors for 125I-α-BTx are blocked by methyllycaconitine and d-tubocurarine. Solubilized α-BTx receptors from plasma membranes of SCC-A9 cells cosediment with 10S neuronal nAChR pentamers and bind to an α7-specific monoclonal antibody (MAb P27) but not to the muscle nAChR-reactive MAb 35. However, MAb P27 and MAb 35 both bind to α-BTx receptors solubilized from the cytoplasmic compartments of SCC-A9 and the parental SCC-37 line. Reverse transcription-PCR analysis revealed RNA transcripts for α7 and α1 subunits in both SCC-A9 and SCC-37 cells. The nAChRs that are expressed in these novel human cell lines can regulate cation fluxes directly as well as indirectly by synergizing with the activity of voltage-sensitive Ca2+ channels. These activities may influence the secretion of autocrine growth factors and the transcription of growth regulatory genes and thus be pertinent to the growth and metastasis of malignant neuroendocrine neoplasms.