Complex relationship between Ins(1,4,5)P₃ accumulation and Ca²⁺‐signalling in a human neuroblastoma revealed by cellular differentiation

Abstract

1 Dierentiation of SH-SY5Y neuroblastoma cells induces morphological and biochemical changes consistent with a more neuronal phenotype. These cells may therefore provide a model for studying phenomena such as signal transduction in a neuronal context whilst retaining the advantages of a homogenous cell population expressing a well characterized array of G-protein coupled receptors. 2 This study examined the eects of dierentiating SH-SY5Y cells on muscarinic-and bradykininreceptor-mediated phosphoinositide and Ca 2+ signalling. Retinoic acid (10 mM, 6 days) along with a lowered serum concentration produced phenotypic changes consistent with dierentiation including reduced proliferation and increased neurite outgrowth. 3 Dierentiation increased the magnitude and potency of rapid Ins(1,4,5)P 3 responses to a full muscarinic receptor agonist. Bradykinin receptor-mediated Ins(1,4,5)P 3 signalling was also potentiated following dierentiation. Determination of agonist-evoked accumulation of [ 3 H]-inositol phosphates under lithium-block demonstrated these changes re¯ected enhanced phospholipase C activity which is consistent with observed increases in the expression of muscarinic and bradykinin receptors. 4 Despite the marked alterations in Ins(1,4,5)P 3 signalling following dierentiation, elevations of intracellular [Ca 2+ ] were totally unaltered. Thus, in SH-SY5Y cells, the relationship between the elevations of Ins(1,4,5)P 3 and intracellular [Ca 2+ ] is agonist dependent and aected by the state of dierentiation. This demonstrates that mechanisms other than the measured increase in Ins(1,4,5)P 3 regulate the elevation of intracellular [Ca 2+ ].

Specificity of G Protein-Coupled Receptor Kinase 6-Mediated Phosphorylation and Regulation of Single-Cell M3Muscarinic Acetylcholine Receptor Signaling

“…7A). In SH-SY5Y cell population studies, the EC 50 for MChstimulated InsP 3 accumulation is ϳ5 M (Martin et al, 1999). However, we found that EC 50 values varied between individual SH-SY5Y cells, which caused us to use either 3 or 10 M MCh as an approximate EC 50 concentration depending on the cell studied.…”

Neuronal Differentiation of P19 Embryonal Carcinoma Cells Modulates Kinin B2 Receptor Gene Expression and Function

“…In PC12 cells, BK signaling resulted in translocation of protein kinase C⑀, an enzyme implicated in regulation of neurite outgrowth, and therefore BK was suggested to participate in neuronal differentiation of this cell line (59). The BK-induced signal transduction closely resembles inositol 1,4,5-trisphosphate accumulation followed by calcium release from intracellular stores, which was characterized during differentiation of the human neuroblastoma cell line SH-SY5Y (60,61). Calcium transients have also been observed in the growth cones of developing neurons (62).…”

Characterization of the metabotropic glutamate receptors mediating phospholipase C activation and calcium release in cerebellar granule cells: calcium‐dependence of the phospholipase C response

“…In some cell systems, the InsP 3 ‐elicited increase of cytosolic Ca 2+ concentration ([Ca 2+ ] i ) can be further enhanced by Ca 2+ ‐induced Ca 2+ release through ryanodine receptors (Taylor & Broad, 1998). In neurons, a good quantitative correlation between InsP 3 formation and changes in [Ca 2+ ] i has been reported generally, although recent data indicate this correlation can be dependent on the agonist used (Martin et al ., 1999; del Río et al ., 1999); and it has been suggested that intracellular Ca 2+ mobilization is independent of PLC activation (Chavis et al ., 1996). On the other hand, agonist stimulation of PLC has been shown to be Ca 2+ ‐dependent in several receptor systems.…”