Calcium‐dependence of histamine‐ and carbachol‐induced inositol phosphate formation in human U373 MG astrocytoma cells: comparison with HeLa cells and brain slices

Abstract

Histamine (1 mm) induced an accumulation of inositol monophosphate ([3H]‐IP1) in the U373 MG human astrocytoma cell line which increased with time in the presence of 30 mm Li+. After a 30 min incubation period with 1 mm histamine [3H]‐IP1 was the major product detected (84 ± 1 % of total [3H]‐IPx) and was present at a level 11 (±1) fold of basal accumulation. Concentration‐response curves for histamine‐induced [3H]‐IP1 accumulation in U373 MG cells (EC50 5.4 ± 0.5 μm) were shifted to the right in a parallel fashion by mepyramine (slope of a Schild plot 0.99 ± 0.08), yielding a Kd for mepyramine of 3.5 ± 0.3 nm, consistent with the involvement of histamine H1‐receptors. The temelastine‐sensitive binding of [3H]‐mepyramine to a membrane fraction from U373 MG cells was hyperbolic and had a mean Kd of 2.5 ± 1.0 nm. The maximum amount of temelastine‐sensitive binding was 86 ± 19 pmol g−1 membrane protein. Carbachol also induced [3H]‐IP1 accumulation in U373 MG cells, 2.8 (± 0.1) fold of basal with 1 mm carbachol, with an EC50 of 48 ± 8 μm. Pirenzepine shifted carbachol concentration‐response curves to the right (slope of Schild plot 0.89 ± 0.07) giving a Kd for pirenzepine of 0.10 ± 0.01 μm, suggesting that phosphoinositide hydrolysis in U373 MG cells is mediated by the M3‐, rather than the M1‐, muscarinic receptor subtype. [3H]‐IP1 accumulation induced by both 1 mm histamine and by 1 mm carbachol increased when the Ca2+ concentration of the medium was increased from ‘zero’ (no added Ca2+) to 0.3 mm. Histamine‐stimulated [3H]‐IP1 accumulation was further increased, although not so markedly, as the Ca2+ was raised to 4 mm. The same pattern was apparent with histamine‐induced accumulations of [3H]‐IP2 and [3H]‐IP3. In contrast, [3H]‐IPx accumulation in response to carbachol increased between 0.3 and 1.3 mm, but thereafter remained unchanged ([3H]‐IP1) or declined ([3H]‐IP2 and [3H]‐IP3). In HeLa cells, [3H]‐IP1 accumulations induced by 1 mm histamine and 1 mm carbachol showed the same pattern of Ca2+ dependence and were independent of extracellular Ca2+ above 0.3 mm (histamine) or 1.3 mm (carbachol). The response to carbachol appeared to be mediated by an M3‐muscarinic receptor (apparent Kd for pirenzepine 0.09 μm). In cross‐chopped slices of guinea‐pig cerebral cortex and guinea‐pig cerebellum, [3H]‐IP1 accumulation induced by 1 mm histamine in the presence of 10 mm Li+ increased as the extracellular Ca2+ was increased from 0.3 to 2.5 mm, but a further increase to 4 mm had no further effect. In contrast the response to histamine in rat cerebral cortex increased markedly between 1.3 and 4 mm Ca2+. Accumulations of [3H]‐IP1 induced by carbachol in guinea‐pig or rat cerebral cortical slices were not increased as extracellular Ca2+ was raised from 0.3 to 4 mm. Nimodipine (100 nm) and ω‐conotoxin (3 μm) had no significant effect on histamine‐induced [3H]‐IP1 accumulation in rat cerebral cortical slices or in U373 MG cells. We conclude that histamine‐induced [3H]‐IP1 accumulation in U373 MG cells does appear to have a compo...

Modulation of Histamine-induced Ca2+ Release by Protein Kinase C

“…Fig. 6 (27). Consistently, the kinetics of both the cytosolic and mitochondrial [Ca 2ϩ ] peaks induced by carbachol were slower than those induced by histamine, although the height of the peaks were similar.…”

Characterisation of α1B-adrenoceptors linked to inositol phosphate formation and calcium mobilisation in human astrocytoma U373 MG cells

“…In U373 MG cells the pattern of the Ca 2+ dependence of NA-stimulated [ 3 H]-IP 1 accumulation (Fig. 4) is similar to that for carbachol (Arias-Montaño et al 1994) and gives no indication of any α 1A -receptor contribution.…”

Histamine‐induced Ca2+ entry in human astrocytoma U373 MG cells: Evidence for involvement of store‐operated channels

“…Previous work showed the presence of H 1 receptors in this cell line (Arias-Montaño et al, 1994), and exposure to histamine (30 mM) in the presence of Ca 21 (1.8 mM) in the extracellular medium induced an increase in [Ca 21 ] i that occurred in two temporally distinct phases: a fast transitory response with a peak of 413 6 73 nM followed by a much slower increase in [Ca 21 ] i , which remained while histamine was present ( Fig. 2A).…”

“…This action is mediated by the activation of H 1 receptors, expressed in relatively high density in the plasma membrane of U373 MG cells (Arias-Montaño et al, 1994) because MEP, a selective antagonist at H 1 receptors, abolished the increase in intracellular Ca 21 induced by histamine application (Fig. 2).…”

“…At the cellular level, H 1 and H 2 receptors have been identified not only in neurons, but also in astrocytes and blood vessel cells (Inagaki and Wada, 1994). Histamine-induced Ca 21 entry in glial cells and glioma-derived cell lines has been described, and the presence of store-operated and receptor-operated channels has been previously suggested (Arias-Montaño et al, 1994;Weiger et al, 1997;Jung et al, 2000;Mauban et al, 2006). However, very little is known regarding the molecular nature of the channels that participate in this physiological process and their regulation.…”