Lan-1: A human neuroblastoma cell line with M₁ and M₃ muscarinic receptor subtypes coupled to intracellular Ca²⁺ elevation and lacking Ca²⁺ channels activated by membrane depolarization

The LAN-1 clone, a cell line derived from a human neuroblastoma, possesses muscarinic receptors. The stimulation of these receptors with increasing concentrations of carbachol (CCh; 1-1,000 μM) caused a dose-dependent increase of the intracellular free Ca²⁺ concentration ([Ca²⁺]_i). This increase was characterized by an early peak phase (10 s) and a late plateau phase. The removal of extracellular Ca²⁺ reduced the magnitude of the peak phase to ∼ 70% but completely abolished the plateau phase. The muscarinic-activated Ca²⁺ channel was gadolinium (Gd³⁺) blockable and nimodipine and ω-conotoxin insensitive. In addition, membrane depolarization did not cause any increase in [Ca²⁺]_i. The CCh-induced [Ca²⁺]_i elevation was concentration-dependently inhibited by pirenzepine and 4-diphenylacetoxy-N-methylpiperidine methiodide, two rather selective antagonists of M, and M₃ muscarinic receptor subtypes, respectively, whereas methoctramine, an M₂ antagonist, was ineffective. The coupling of M₁ and M₃ receptor activation with [Ca²⁺]_i elevation does not seem to be mediated by a pertussis toxin-sensitive guanine nucleotide-binding protein or by the diacylglycerol-protein kinase C system. The mobilization of [Ca²⁺]_i elicited by M₁ and M₃ muscarinic receptor stimulation seems to be dependent on an inositol trisphosphate-sensitive intracellular store. In addition, ryanodine did not prevent CCh-induced [Ca²⁺]_i mobilization, and, finally, LAN-1 cells appear to lack caffeine-sensitive Ca²⁺ stores, because the methylxanthine was unable to elicit intracellular Ca²⁺ mobilization, under basal conditions, after a subthreshold concentration of CCh (0.3 μM), or after thapsigargin.