Microinjection of myo-Inositol(1,4,5)trisphosphate and Other Calcium-Mobilizing Agents into Intact Adherent Cells

Wilcox, R. A.; Forsythe, Ian D.; McCann, Terence J.

doi:10.1385/1-59259-250-3:193

Cited by 2 publications

(3 citation statements)

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“…The intracellular locations of Ins(1,4,5)P 3 R 1 and Ins(1,4,5)P 3 R 2 were determined using antibodies specific for these proteins and immunofluorescence. The specificity of the antibodies for Ins-(1,4,5)P 3 R 1 and Ins(1,4,5)P 3 R 2 was confirmed by Western blot analysis using extracts of L15 mouse fibroblasts [27], which express Ins(1,4,5)P 3 R 1 , but do not express significant levels of Ins(1,4,5)P 3 R 2 [29], and extracts of SaOS-2 human osteoblasts, which predominantly express Ins(1,4,5)P 3 R 2 and Ins(1,4,5)P 3 R 3 [28]. A single band corresponding to the expected size of the Ins(1,4,5)P 3 R was observed in each case (results not shown).…”

Section: Intracellular Distribution Of Ins(145)p 3 R 1 and Ins(14mentioning

confidence: 81%

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Inositol trisphosphate analogues selective for types I and II inositol trisphosphate receptors exert differential effects on vasopressin-stimulated Ca2+ inflow and Ca2+ release from intracellular stores in rat hepatocytes

Gregory

Hughes

Riley

et al. 2004

Biochemical Journal

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Previous studies have shown that adenophostin A is a potent initiator of the activation of SOCs (store-operated Ca2+ channels) in rat hepatocytes, and have suggested that, of the two subtypes of Ins(1,4,5)P3 receptor predominantly present in rat hepatocytes [Ins(1,4,5)P3R1 (type I receptor) and Ins(1,4,5)P3R2 (type II receptor)], Ins(1,4,5)P3R1s are required for SOC activation. We compared the abilities of Ins(1,4,6)P3 [with higher apparent affinity for Ins(1,4,5)P3R1] and Ins(1,3,6)P3 and Ins(1,2,4,5)P4 [with higher apparent affinities for Ins(1,4,5)P3R2] to activate SOCs. The Ins(1,4,5)P3 analogues were microinjected into single cells together with fura 2, and dose-response curves for the activation of Ca2+ inflow and Ca2+ release from intracellular stores obtained for each analogue. The concentration of Ins(1,4,6)P3 which gave half-maximal stimulation of Ca2+ inflow was substantially lower than that which gave half-maximal stimulation of Ca2+ release. By contrast, for Ins(1,3,6)P3 and Ins(1,2,4,5)P3, the concentration which gave half-maximal stimulation of Ca2+ inflow was substantially higher than that which gave half-maximal stimulation of Ca2+ release. The distribution of Ins(1,4,5)P3R1 and Ins(1,4,5)P3R2 in rat hepatocytes cultured under the same conditions as those employed for the measurement of Ca2+ inflow and release was determined by immunofluorescence. Ins(1,4,5)-P3R1s were found predominantly at the cell periphery, whereas Ins(1,4,5)P3R2s were found at the cell periphery, the cell interior and nucleus. It is concluded that the idea that a small region of the endoplasmic reticulum enriched in Ins(1,4,5)P3R1 is required for the activation of SOCs is consistent with the present results for hepatocytes.

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Section: Intracellular Distribution Of Ins(145)p 3 R 1 and Ins(14mentioning

confidence: 81%

“…SaOS-2 human osteosarcoma cells (A.T.C.C., Rockville, MD, U.S.A.) were kindly provided by Dr T. J. McCann, Babraham Institute, Cambridge, U.K., and L15 mouse fibroblasts [27] by Professor K. Mikoshiba. SaOS-2 cells [28] and L15 mouse fibroblasts [29] were cultured as described previously.…”

Section: Methodsmentioning

confidence: 99%

Inositol trisphosphate analogues selective for types I and II inositol trisphosphate receptors exert differential effects on vasopressin-stimulated Ca2+ inflow and Ca2+ release from intracellular stores in rat hepatocytes

Gregory

Hughes

Riley

et al. 2004

Biochemical Journal

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“…Another important factor determining [Ca 2+ ]i dynamics is inositol 1,4,5-trisphosphate receptors (IP3Rs), which mediates the release of Ca 2+ from intracellular storage sites in the ER [11]. Studies have shown that the cytosolically generated inositol 1,4,5-trisphosphate (IP3) also triggers putative nuclear membrane IP3Rs [12][13][14]. Furthermore, as IP3Rs have high-affinity Ca 2+ -binding domains, any rise in cytosolic Ca 2+ could, in principle, activate channels, yet its role in fluoride-exposed OBs is still unknown.…”

Section: Introductionmentioning

confidence: 99%

Mechanisms of Intracellular Calcium Homeostasis in MC3T3-E1 Cells and Bone Tissues of Sprague-Dawley Rats Exposed to Fluoride

Duan

Zhang

et al. 2015

Biol Trace Elem Res

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Calcium homeostasis of osteoblasts (OBs) has an important role in the physiology and pathology of bone tissue. In order to study the mechanisms of intracellular calcium homeostasis, MC3T3-E1 cells and Sprague-Dawley rats were treated with different concentrations of fluoride. Then, we examined intracellular-free calcium ion ([Ca(2+)]i) in MC3T3-E1 cells as well as mRNA and protein levels of Cav1.2, the main subunit of L-type voltage-dependent calcium channels (VDCCs), Na(+)/Ca(2+) exchange carriers (NCS), and plasma membrane Ca(2+)-ATPase (PMCA), inositol 1,4,5-trisphosphate receptor (IP3R) channels, sarco/endoplasmic reticulum calcium ATPase 2b (SERCA2b)/ATP2A2 in vitro, and rat bone tissues in vivo. Our results showed that [Ca(2+)]i of fluoride-treated OBs increased in a concentration-dependent manner with an increase in the concentration of fluoride. We also found that the low dose of fluoride led to high expression levels of Cav1.2, NCS-1, and PMCA and low expression levels of IP3R and SERCA2b/ATP2A2, while the high dose of fluoride induced an increase in SERCA2b/ATP2A2 levels and decrease in Cav1.2, PMCA, NCS-1, and IP3R levels. These results demonstrate that calcium channels and calcium pumps of plasma and endoplasmic reticulum (ER) membranes keep intracellular calcium homeostasis by regulating Cav1.2, NCS-1, PMCA, IP3R, and SERCA2b/ATP2A2 expression.

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Microinjection of myo-Inositol(1,4,5)trisphosphate and Other Calcium-Mobilizing Agents into Intact Adherent Cells

Cited by 2 publications

References 0 publications

Inositol trisphosphate analogues selective for types I and II inositol trisphosphate receptors exert differential effects on vasopressin-stimulated Ca2+ inflow and Ca2+ release from intracellular stores in rat hepatocytes

Inositol trisphosphate analogues selective for types I and II inositol trisphosphate receptors exert differential effects on vasopressin-stimulated Ca2+ inflow and Ca2+ release from intracellular stores in rat hepatocytes

Mechanisms of Intracellular Calcium Homeostasis in MC3T3-E1 Cells and Bone Tissues of Sprague-Dawley Rats Exposed to Fluoride

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