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- BARKER, CJ, et al.
(författare)
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Inositol 1,2,3-trisphosphate and inositol 1,2- and/or 2,3-bisphosphate are normal constituents of mammalian cells
- 1995
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Ingår i: The Biochemical journal. - : Portland Press Ltd.. - 0264-6021 .- 1470-8728. ; 306306 ( Pt 2), s. 557-564
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Tidskriftsartikel (refereegranskat)abstract
- 1. An inositol trisphosphate (InsP3) distinct from Ins(1,4,5)P3 and Ins(1,3,4)P3, which we previously observed in myeloid and lymphoid cells [French, Bunce, Stephens, Lord, McConnell, Brown, Creba and Michell (1991) Proc R. Soc. London B 245, 193-201; Bunce, French, Allen, Mountford, Moore, Greaves, Michell and Brown (1993) Biochem. J. 289, 667-673], is present in WRK1 rat mammary tumour cells and pancreatic endocrine beta-cells. 2. It has been identified as Ins(1,2,3)P3 by a combination of oxidation to ribitol, a structurally diagnostic polyol, and ammoniacal hydrolysis to identified inositol monophosphates. 3. Ins(1,2,3)P3 concentration in HL60 cells changed little during stimulation by ATP or fMetLeuPhe or during neutrophilic or monocytic differentiation, and Ins(1,2,3)P3 was unresponsive to vasopressin in WRK1 cells. 4. Ins(1,2,3)P3 was usually more abundant than Ins(1,4,5)P3, often being present at concentrations between approximately 1 microM and approximately 10 microM. 5. HL60, WRK-1 and lymphoid cells also contain Ins(1,2)P2 or Ins(2,3)P2, or a mixture of these two enantiomers, as a major InsP2 species. 6. Ins(1,2,3)P3 and Ins(1,2)P2/Ins(2,3)P2 are readily detected in cells labelled for long periods, but not in acutely labelled cells. This behaviour resembles that of InsP6, the most abundant cellular inositol polyphosphate that includes the 1,2,3-trisphosphate motif, which also achieves isotopic equilibrium with inositol only slowly. 7. Ins(1,2,3)P3 is the major InsP3 that accumulates during metabolism of InsP6 by WRK-1 cell homogenates. 8. Possible metabolic relationships between Ins(1,2,3)P3, Ins(1,2)P2/Ins(2,3)P2 and other inositol polyphosphates in cells, and a possible role for Ins(1,2,3)P3 in cellular iron handling, are considered.
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| 2. |
- Islam, M. Shahidul, et al.
(författare)
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Effects of caffeine on cytoplasmic free Ca2+ concentration in pancreatic beta-cells are mediated by interaction with ATP-sensitive K+ channels and L-type voltage-gated Ca2+ channels but not the ryanodine receptor
- 1995
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Ingår i: Biochemical Journal. - : Portland Press Ltd.. - 0264-6021 .- 1470-8728. ; 306:Pt 3, s. 679-686
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Tidskriftsartikel (refereegranskat)abstract
- In the pancreatic beta-cell, an increase in the cytoplasmic free Ca2+ concentration ([Ca2+]i) by caffeine is believed to indicate mobilization of Ca2+ from intracellular stores, through activation of a ryanodine receptor-like channel. It is not known whether other mechanisms, as well, underlie caffeine-induced changes in [Ca2+]i. We studied the effects of caffeine on [Ca2+]i by using dual-wavelength excitation microfluorimetry in fura-2-loaded beta-cells. In the presence of a non-stimulatory concentration of glucose, caffeine (10-50 mM) consistently increased [Ca2+]i. The effect was completely blocked by omission of extracellular Ca2+ and by blockers of the L-type voltage-gated Ca2+ channel, such as D-600 or nifedipine. Depletion of agonist-sensitive intracellular Ca2+ pools by thapsigargin did not inhibit the stimulatory effect of caffeine on [Ca2+]i. Moreover, this effect of caffeine was not due to an increase in cyclic AMP, since forskolin and 3-isobutyl-1-methylxanthine (IBMX) failed to raise [Ca2+]i in unstimulated beta-cells. In beta-cells, glucose and sulphonylureas increase [Ca2+]i by causing closure of ATP-sensitive K+ channels (KATP channels). Caffeine also caused inhibition of KATP channel activity, as measured in excised inside-out patches. Accordingly, caffeine (> 10 mM) induced insulin release from beta-cells in the presence of a non-stimulatory concentration of glucose (3 mM). Hence, membrane depolarization and opening of voltage-gated L-type Ca2+ channels were the underlying mechanisms whereby the xanthine drug increased [Ca2+]i and induced insulin release. Paradoxically, in glucose-stimulated beta-cells, caffeine (> 10 mM) lowered [Ca2+]i. This effect was due to the fact that caffeine reduced depolarization-induced whole-cell Ca2+ current through the L-type voltage-gated Ca2+ channel in a dose-dependent manner. Lower concentrations of caffeine (2.5-5.0 mM), when added after glucose-stimulated increase in [Ca2+]i, induced fast oscillations in [Ca2+]i. The latter effect was likely to be attributable to the cyclic AMP-elevating action of caffeine, leading to phosphorylation of voltage-gated Ca2+ channels. Hence, in beta-cells, caffeine-induced changes in [Ca2+]i are not due to any interaction with intracellular Ca2+ pools. In these cells, a direct interference with KATP channel- and L-type voltage-gated Ca(2+)-channel activity is the underlying mechanism by which caffeine increases or decreases [Ca2+]i.
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