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R F Irvine

Publications and source records attributed to R F Irvine.

At least 91 records · Page 5Linked to original sources
PubMed1990

Nuclear inositol lipids. Relationship between growth factor induced metabolic changes and protein kinase C activity.

We have sought to establish the effect of mitogen treatment on nuclear inositol lipids and the relationship between inositol cycle products and hyperphosphorylation of nuclear proteins via PKC during the lag phase leading to the onset of DNA synthesis. Swiss 3T3 cells were labelled for 36 hr with high levels of [3H]-myo-inositol and the radioactivity in nuclear inositol phospholipids was measured. Treatment of cells for 2 min, but not for 4 hr, with mitogenic concentrations of insulin-like growth factor I and bombesin caused a marked decrease in PtdInsP and PtdInsP2. Moreover, in vivo phosphorylation of some nuclear proteins occurs later on. Among these proteins, histone H1 and 0.75 M PCA soluble polypeptide, with an apparent Mr of 21,000 as revealed by electrophoretic analysis, are phosphorylated in vitro by protein kinase C in isolated nuclei purified from 3T3 cells treated for 90 min with IGF-I and bombesin. Since these phosphorylative events follow the earlier changes in nuclear polyphoinositide metabolism induced by the same mitogen combination, it seems possible that these two phenomena are related to each other and trigger the synthetic machinery responsible for replicating DNA.

Adenosine Triphosphate↗
PubMed1990

Ram spermatozoa produce inositol 1,4,5-trisphosphate but not inositol 1,3,4,5-tetrakisphosphate during the Ca2+/ionophore-induced acrosome reaction.

An investigation was made of the production of inositol tris- and tetrakisphosphates concomitant with ionophore-stimulated breakdown of PtdIns(4,5)P2 in ram spermatozoa. As spermatozoa displayed very low rates of incorporation of [3H]inositol into their phosphoinositides, the studies were carried out using 32P-labelled cells. Using a specially developed procedure, inositol tris- and tetrakisphosphates were isolated, free of labelled ATP and P(i); they were then separated from each other (and from other minor labelled compounds) and analysed, using ionophoresis and HPLC. Levels of 32P-labelled material with the chromatographic characteristics of Ins(1,4,5)P3 were very low in untreated cells, but rose sharply with ionophore treatment, in parallel with rapid PtdInsP2 breakdown. No 32P-labelled material with the characteristics of Ins(1,3,4,5)P4 or Ins(1,3,4)P3 was found, and there was no evidence for phosphorylation of Ins(1,4,5)P3 in sperm homogenates. The implications of our findings are discussed with respect to the physiological modulation of Ca2+ influx that is required to initiate the acrosome reaction at fertilization.

Acrosome↗
PubMed1990

Spatial dynamics of intracellular calcium in agonist-stimulated vascular smooth muscle cells.

Vasoconstrictor agonists stimulate smooth muscle contraction by inducing a rise in intracellular free Ca2+. Digital-imaging microscopy of fura-2 fluorescence from single vascular smooth muscle cells cultured from the human internal mammary artery has allowed us to record the subcellular alterations in Ca2+ that occur immediately after stimulation by receptor agonists. The thrombin-induced rise in cytoplasmic free Ca2+ begins in a discrete region typically located close to the end of the cell. Subsequently, this region of elevated Ca2+ expands until Ca2+ is elevated throughout the cell cytoplasm. The rate of spreading in the region of elevated Ca2+ in a linear direction averaged 10.1 microns/s, enabling it to traverse the length of most cells within approximately 5 s, and involved rises in Ca2+ of between 200 and 500 nM. In some cells, the Ca2+ rise began at both ends and collided midway. Similar dynamic changes in the spatial distribution of Ca2+ were recorded in cells stimulated by acetylcholine. The novel observation that vasoconstrictor agonists induce an elevation of Ca2+ in a localized region which subsequently expands throughout the cytoplasm of single smooth muscle cells may provide new insight into the nature of Ca2+ signaling in vascular tissue.

Acetylcholine↗
PubMed1990

Care and cleaning of the non-contemporary photographic image.

Continuing the series on various aspects of archival permanence, this paper suggest approaches which can be safely made in the care and cleaning of non-contemporary photographic images. In particular the 19th century produced images such as daguerreotype, collodion positive and the ferrotype.

Image Enhancement↗
PubMed1989

Inositol phosphates and cell signalling.

Inositol 1,4,5-trisphosphate is a second messenger which regulates intracellular calcium both by mobilizing calcium from internal stores and, perhaps indirectly, by stimulating calcium entry. In these actions it may function with its phosphorylated metabolite, inositol 1,3,4,5-tetrakisphosphate. The subtlety of calcium regulation by inositol phosphates is emphasized by recent studies that have revealed oscillations in calcium concentration which are perhaps part of a frequency-encoded second-messenger system.

Animals↗
PubMed1989

Automated isocratic high-performance liquid chromatography of inositol phosphate isomers.

Isomers of inositol phosphates from biological samples can be analysed by anion-exchange h.p.l.c., by using isocratic elution with phosphate buffers. The method involves the preliminary processing of the extracted samples with conventional soft-gel anion-exchange resins, including the commonly used Dowex resins, followed by direct analysis with h.p.l.c. of a portion of relevant fractions. Run times (up to 20 min) and collected fraction numbers (up to 24) are minimal, so that if the method is used in conjunction with automated h.p.l.c. injection a high throughput of samples is achieved.

Chromatography, High Pressure Liquid↗
PubMed1989

Inositol 1,3,4,5-tetrakisphosphate and inositol 1,4,5-trisphosphate act by different mechanisms when controlling Ca2+ in mouse lacrimal acinar cells.

In internally perfused single lacrimal acinar cells the competitive inositol 1,4,5-trisphosphate (Ins 1,4,5-P3)-antagonist heparin inhibits the ACh-evoked K+ current response mediated by internal Ca2+ and also blocks both the Ins 1,4,5-P3-evoked transient as well as the sustained K+ current increase evoked by combined stimulation with internal Ins 1,4,5-P3 and inositol 1,3,4,5-tetrakisphosphate (Ins 1,3,4,5-P4). When, during sustained stimulation with both Ins 1,4,5-P3 and Ins 1,3,4,5-P4, one of the inositol polyphosphates is removed, the K+ current declines; whereas removal of Ins 1,4,5-P3 results in an immediate termination of the response, removal of Ins 1,3,4,5-P4 only causes a very gradual and slow reduction in the current. Ins 1,3,4,5-P4 is therefore not an acute controller of Ca2+ release from stores into the cytosol, but modulates the release of Ca2+ induced by Ins 1,4,5,P3 by an unknown mechanism, perhaps by linking Ins 1,4,5 P3-sensitive and insensitive Ca2+ stores.

Acetylcholine↗
PubMed1989

Inositol 1,3,4,5-tetrakisphosphate causes release of Ca2+ from permeabilized mouse lymphoma L1210 cells by its conversion into inositol 1,4,5-trisphosphate.

Addition of Ins(1,3,4,5)P4 at micromolar concentrations causes release of Ca2+ from electroporated L1210 cells, but not from digitonin-permeabilized cells. This was shown to be due to its conversion into Ins(1,4,5)P3, because only the electroporated cells convert Ins(1,3,4,5)P4 into Ins(1,4,5)P3. Thus electroporation appears to activate or expose an Ins(1,3,4,5)P4 3-phosphatase.

Animals↗
PubMed1989

Changes in nuclear inositol phospholipids induced in intact cells by insulin-like growth factor I.

Swiss 3T3 cells were labelled for 36 hours with high levels of 3H-myo-inositol and the radioactivity in nuclear inositol phospholipids was measured. Treatment of cells for 2 minutes, but not for 4 hours, with mitogenic concentrations of insulin-like growth factor I and bombesin caused a slight decrease in PtdIns and more marked decreases in PtdInsP and PtdInsP2. These effects were not seen if isolated nuclei were incubated with IGF-I and bombesin. We interpret these results to mean that rapid mass changes occur in nuclear inositol phospholipids in the early stages of the mitotic response.

Animals↗
PubMed1989

Inositol 1:2-cyclic,4,5-trisphosphate is only a weak agonist at inositol 1,4,5-trisphosphate receptors.

The activity of inositol 2,4,5-trisphosphate and inositol 1:2-cyclic,4,5-trisphosphate relative to inositol 1,4,5-trisphosphate was examined by two assays; firstly, in a binding assay using rat cerebellar membranes, and secondly, in a Ca2+-mobilization assay using permeabilized Swiss 3T3 cells. In both assays the first two phosphates have a potency at least an order of magnitude less than inositol 1,4,5-trisphosphate. The possible reasons for differences between these results and previous data are discussed, as are the implications for any putative physiological role for the cyclic trisphosphate.

Animals↗
PubMed1989

Inositol 1,3,4,5-tetrakisphosphate is essential for sustained activation of the Ca2+-dependent K+ current in single internally perfused mouse lacrimal acinar cells.

We have examined the effects of various inositol polyphosphates, alone and in combination, on the Ca2+-activated K+ current in internally perfused, single mouse lacrimal acinar cells. We used the patch-clamp technique for whole-cell current recording with a set-up allowing exchange of the pipette solution during individual experiments so that control and test periods could be directly compared in individual cells. Inositol 1,4,5-trisphosphate (Ins 1,4,5 P3) (10-100 microM) evoked a transient increase in the Ca2+-sensitive K+ current that was independent of the presence of Ca2+ in the external solution. The transient nature of the Ins 1,4,5 P3 effect was not due to rapid metabolic breakdown, as similar responses were obtained in the presence of 5 mM 2,3-diphosphoglyceric acid, that blocks the hydrolysis of Ins 1,4,5 P3, as well as with the stable analogue DL-inositol 1,4,5-trisphosphorothioate (Ins 1,4,5 P(S)3) (100 microM). Ins 1,3,4 P3 (50 microM) had no effect, whereas 50 microM Ins 2,4,5 P3 evoked responses similar to those obtained by 10 microM Ins 1,4,5 P3. A sustained increase in Ca2+-dependent K+ current was only observed when inositol 1,3,4,5-tetrakisphosphate (Ins 1,3,4,5 P4) (10 microM) was added to the Ins 1,4,5 P3 (10 microM)-containing solution and this effect could be terminated by removal of external Ca2+. The effect of Ins 1,3,4,5 P4 was specifically dependent on the presence of Ins 1,4,5 P3 as it was not found when 10 microM concentrations of Ins 1,3,4 P3 or Ins 2,4,5 P3 were used.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Phosphatidylinositol(4,5)bisphosphate and Phosphatidylinositol(4)phosphate in Plant Tissues.

Pea (Pisum sativum) leaf discs or swimming suspensions of Chlamydomonas eugametos were radiolabeled with [(3)H]myo-inositol or [(32)P]Pi and the lipids were extracted, deacylated, and their glycerol moieties removed. The resulting inositol trisphosphate and bisphosphate fractions were examined by periodate degradation, reduction and dephosphorylation, or by incubation with human red cell membranes. Their likely structures were identified as d-myo-inositol(1,4,5)trisphosphate and d-myo-inositol(1,4,)-bisphosphate. It is concluded that plants contain phosphatidylinositol(4)phosphate and phosphatidylinositol(4,5)bisphosphate; no other polyphosphoinositides were detected.

Journal Article↗
PubMed1988

Inositol phosphate production and Ca2+ mobilization in human umbilical-vein endothelial cells stimulated by thrombin and histamine.

Human umbilical-vein endothelial cells (HUVECs) were cultured, and their inositol phosphate formation and Ca2+ mobilization in response to thrombin and histamine were studied. Evidence from measurement of intracellular Ca2+ in the absence of extracellular Ca2+ established that the two agonists were both acting on a single cell population, and suggested that a Ca2+-influx component was stimulated which was dependent on receptor-occupancy. After 30 s of stimulation in the presence of 10 mM-LiCl, the effects of 20 microM-histamine and 1 unit of thrombin/ml on formation of inositol phosphates were additive, but at 5 min they were not. HUVECs labelled with myo-[3H]inositol for 72 h synthesized radiolabelled inositol pentakis- and hexakis-phosphate. The predominant isomers of inositol mono-, bis- and tris-phosphates whose formation was stimulated were the 4-phosphate, the 1,4-bisphosphate and the 1,3,4-trisphosphate.

Calcium↗
PubMed1988

Rapid changes in phospholipid metabolism in the nuclei of Swiss 3T3 cells induced by treatment of the cells with insulin-like growth factor I.

When highly-purified nuclei of Swiss-mouse 3T3 cells are incubated with [32P]-gamma ATP, radioactivity is incorporated into phosphatidic acid and the two polyphosphoinositol lipids, phosphatidylinositol(4)P and phosphatidylinositol(4,5)P2. If the cells are pre-treated with insulin-like growth factor I, this incorporation into the phospholipids is decreased. The effect is maximal by 2 minutes, is transient in that it has disappeared by one hour, and is increased markedly by the co-addition of bombesin, even though bombesin alone has no effect. We suggest that some aspect of polyphosphoinositide metabolism is altered in the nucleus (leading to a decreased radiolabelling) when the cells are treated with mitogenic growth factors, and that this change in inositide metabolism is a very early event in the sequence leading to cell division.

Adenosine Triphosphate↗
PubMed1988

Formation of methylphosphoryl inositol phosphates by extractions that employ methanol.

Fixatives that contain methanol extract an unknown compound from several tissues including the retinas of squid (Loligo). We have determined that the compound probably contains (1) a myo-inositol ring that is phosphorylated in more than one position (including at the 5-hydroxyl), (2) a charged moiety that is not susceptible to alkaline phosphatase, and (3) a methyl group. We have found that the compound can be made by treating either phosphatidylinositol bisphosphate or human red cell ghosts with acidic methanol. We have confirmed the observation of Lips, Bross & Majerus [Proc. Natl. Acad. Sci. U.S.A. 85, 88-92] that the compound also can be made by methanolysis of inositol (cyclic 1:2,4,5)trisphosphate; however, we have not found inositol (cyclic 1:2,4,5)trisphosphate in either stimulated or unstimulated squid retinas. We tentatively identify the compound as (1-methylphosphoryl)inositol 4,5-bisphosphate formed by methanolysis of phosphatidylinositol 4,5-bisphosphate. By using this methanolysis to incorporate label from [14C]methanol, we have estimated the mass of inositol 1,4,5-trisphosphate in squid retinas to be approx. 30 mumol/l of retinal volume.

Alkaline Phosphatase↗