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PubMed2000

A model of inositol compartmentation in astrocytes based upon efflux kinetics and slow inositol depletion after uptake inhibition.

Intracellular compartmentation of inositol was demonstrated in primary cultures of mouse astrocytes, incubated in isotonic medium, by determination of efflux kinetics after "loading" with [3H]inositol. Three kinetically different compartments were delineated. The largest and most slowly exchanging compartment had a halflife of approximately 9 hr. This slow release leads to retention of a sizeable amount of pre-accumulated inositol in the tissue 24 hr after the onset of uptake inhibition, as confirmed by the observation that the inositol uptake inhibitor fucose caused a larger inhibition of unidirectional inositol uptake than of inositol pool size, measured as accumulated [3H]inositol after 24 hr of combined exposure to the inhibitor and the labeled isotope. Based upon the present observations and literature data, it is suggested that the large, slowly exchanging compartment is largely membrane-associated and participating in signaling via the phosphatidylinositide second messenger system, whereas inositol functioning as an osmolyte is distributed in the cytosol and located in one or both of the compartments showing a faster release.

Animals↗
PubMed1997

Effect of osmolality and myo-inositol deprivation on the transport properties of myo-inositol in primary astrocyte cultures.

myo-Inositol uptake measured in primary astrocyte cultures was saturable in the presence of Na+ with a Km of 13-18 microM and a Vmax of 9.4 nmoles/mg protein/hour in myo-inositol-fed cells, indicating a high affinity transport system. In myo-inositol-deprived cells, Km was about 53 microM with a Vmax of 13.2 nmoles/mg protein/hour. Decreasing osmolality decreased the Vmax to about 1.9 nmoles/mg protein/hour whereas increasing osmolality increased Vmax about 5-fold, while Kms were essentially unchanged in myo-inositol fed cells. In cells deprived of myo-inositol, Vmax decreased in hypotonic medium and increased in hypertonic medium almost 10-fold, but with more than a doubling of the Km regardless of the osmolality. Glucose (25 mM) inhibited myo-inositol uptake 51% whereas the other hexoses used inhibited uptake much less. Our findings indicate that myo-inositol uptake in astrocytes occurs through an efficient carrier-mediated Na(+)-dependent co-transport system that is different from that of glucose and its kinetic properties are affected by myo-inositol availability and osmotic stress.

Animals↗
PubMed1986

Stimulation, by vasopressin and other agonists, of inositol-lipid breakdown and inositol phosphate accumulation in WRK 1 cells.

WRK 1 cells were labelled to equilibrium with 2-myo-[3H]inositol and stimulated with vasopressin. Within 3 s of hormone stimulation there was a marked accumulation of 3H-labelled InsP2 and InsP3 (inositol bis- and tris-phosphate), but not of InsP (inositol monophosphate). There was an associated, and rapid, depletion of 3H-labelled PtdInsP and PtdInsP2 (phosphatidylinositol mono- and bis-phosphates), but not of PtdIns (phosphatidylinositol), in these cells. Some 4% of the radioactivity in the total inositol lipid pool of WRK 1 cells was recovered in InsP2 and InsP3 after 10 s stimulation with the hormone. The selectivity of the vasopressin receptors of WRK 1 cells for a variety of vasopressin agonists and antagonists revealed these to be of the V1a subtype. There was no receptor reserve for vasopressin-stimulated inositol phosphate accumulation in WRK 1 cells. The accumulation of inositol phosphates was enhanced in the presence of Li+ions. Half-maximal accumulation of InsP, InsP2 and InsP3 in vasopressin-stimulated cells was observed with 0.9, 3.0 and 3.6 mM-Li+ respectively. Bradykinin and 5-hydroxytryptamine also provoked inositol phosphate accumulation in WRK 1 cells. The effects of sub-optimal concentrations of bradykinin and vasopressin upon inositol phosphate accumulation were additive, but those of optimal concentrations of the hormones were not.

Animals↗
PubMed1991

Studies of inositol analogues as inhibitors of the phosphoinositide pathway, and incorporation of 2-deoxy-2-fluoro-myo-inositol to give analogues of phosphatidylinositol intermediates.

The incorporation of [3H]Ins into PtdIns by exchange of free and lipid-bound inositol moieties occurs via the action of at least two types of Mg2+/Mn(2+)-dependent enzymes in turkey erythrocytes. One is a nucleotide-independent PtdIns/Ins exchange enzyme and its function is, as yet, unknown, whereas the other is CMP-dependent and appears to be an exchange reaction catalysed by PtdIns synthase. The effects of analogues with modifications of the substituent at the 1-, 2-, 3-, 4- and 5-positions on the incorporation of [3H]Ins into PtdIns under both synthase and exchange reaction conditions were investigated in turkey erythrocytes. Analogues causing substantial inhibition of [3H]Ins incorporation were then used in kinetic experiments to determine the type of inhibition involved. The analogues 1-deoxy-1-fluoro-scyllo-inositol and 5-O-methyl-myo-inositol exhibited the greatest effects on the incorporation of [3H]Ins via both the synthase and exchange reactions, and the kinetic analysis indicated that they were competitive inhibitors of Ins. Ki values of 0.37 mM and 2.87 mM were observed for 1-deoxy-1-fluoro-scyllo-inositol under exchange and synthase reaction conditions respectively; similar Ki values of 0.26 mM and 2.80 mM were observed for 5-O-methyl-myo-inositol in the exchange and synthase reactions respectively. The ability of 1-deoxy-1-fluoro-scyllo-inositol and its diastereoisomer, 2-deoxy-2-fluoro-myo-inositol, to act as substrates for the synthase and exchange reactions in turkey erythrocytes was investigated. The radiolabelled derivative of the former analogue was not incorporated into phospholipids, whereas the radiolabelled derivative of the latter analogue was a poor substrate for the synthase and exchange enzymes. In the presence of ATP, the labelled analogue of PtdIns, derived from 2-deoxy-2-fluoro-myo-[2-3H]inositol, appeared to be converted into phosphorylated PtdIns analogues, presumably by the enzymes of the phosphoinositide pathway.

Animals↗
PubMed1997

Properties of the inositol 3,4,5,6-tetrakisphosphate 1-kinase purified from rat liver. Regulation of enzyme activity by inositol 1,3,4-trisphosphate.

Inositol 3,4,5,6-tetrakisphosphate is a novel intracellular signal that regulates calcium-dependent chloride conductance (Xie, W., Kaetzel, M. A., Bruzik, K. S., Dedman, J. R., Shears, S. B., and Nelson, D. J. (1996) J. Biol. Chem. 271, 14092-14097). The molecular mechanisms that regulate the cellular levels of this signal are not characterized. To pursue this problem we have now studied the 1-kinase that deactivates inositol 3,4,5,6-tetrakisphosphate. The enzyme was purified from rat liver 1600-fold with a 1% yield. The native molecular mass was determined to be 46 kDa by gel filtration. The Km values for inositol 3,4,5,6-tetrakisphosphate and ATP were 0. 3 and 10.6 microM, respectively. The kinase was unaffected by either protein kinase A or protein kinase C. Increases in Ca2+ concentration from 0.1 to 1-2 microM inhibited activity by 10-20%. Most importantly, inositol 1,3,4-trisphosphate was shown to be a potent (Ki = 0.2 microM), specific, and competitive inhibitor of the 1-kinase. Our new kinetic data show that typical receptor-dependent adjustments in cellular levels of inositol 1,3,4-trisphosphate provide a mechanism by which the concentration of inositol 3,4,5,6-tetrakisphosphate is dependent on changes in phospholipase C activity. These conclusions also provide a new perspective to our understanding of the physiological importance of the pathway of inositol phosphate turnover initiated by the inositol 1,4, 5-trisphosphate 3-kinase.

Animals↗
PubMed1976

myo-Inositol metabolism in the neonatal and developing rat fed a myo-inositol-free diet.

Neonatal rats of the Holtzman strain, 6 days of age, were fed a myo-inositol restricted liquid formula by gastric intubation for 10 days, after which they were fed a purified myo-inositol-free diet until they were 72 days old. No differences in weight gain were observed between myo-inositol/100 ml of formula or 150 mg myo-inositol/100 g diet. Most tissues examined from rats fed the myo-inositol deprived formula and diet had lower free myo-inositol levels than the controls with the exception of the liver. Despite reduced free and lipid-bound myo-inositol in the liver, there was no evidence of fatty liver in the young rats at any age. The cerebrum and cerebellum of myo-inositol deprived rats had normal myelination and mitochondriogenesis as judged by the levels of 2',3'-cyclic nucleotide-3'-phosphohydrolase (EC 3.1.4.1) and fumarase (EC 4.2.1.2) activity, respectively.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1991

Effect of desipramine on inositol phosphate formation and inositol phospholipids in rat brain and human platelets.

To examine the mechanism of action of antidepressant drugs, we studied the effect of desipramine (DMI) in vitro on agonist-stimulated inositol phosphate formation and inositol phospholipids in rat brain and human platelets. We observed that DMI inhibited thrombin-stimulated 3H-inositol bisphosphate (IP2) and 3H-inositol trisphosphate (IP3) but not 3H-inositol monophosphate (IP1) formation in human platelets. DMI also inhibited norepinephrine (NE) and serotonin (5-HT) stimulated 3H-IP1 formation in rat cerebral cortex. DMI increased levels of all three 3H-inositol phospholipids, 3H-phosphatidyl inositol (PI), 3H-PI-4-phosphate (PIP), and 3H-PI 4,5-bisphosphate (PIP2), in both platelets and rat cortex. The decreased formation of inositol phosphates and increased levels of [3H]-PI, [3H]-PIP, and [3H]-PIP2 by DMI appears to be due to the inhibition of the enzyme phospholipase C rather than its effects on receptors. It is thus possible that interaction of tricyclic antidepressant drugs with the PI-signaling system may be related to their mechanism of action.

Animals↗
PubMed1991

Inositol-1,2-cyclic-phosphate 2-inositolphosphohydrolase. Substrate specificity and regulation of activity by phospholipids, metal ion chelators, and inositol 2-phosphate.

Glycerophosphoinositol (GroPIns) is a major inositol phosphate in many cell types. In this study we have determined the optimal conditions (pH 8.0 and 0.5 mM MnCl2) for the metabolism of this molecule in an extract from human placenta, and we show that the major product is inositol (1)-phosphate (Ins(1)P). The enzyme activity that catalyzes this reaction is contained in the same protein designated previously as inositol-(1,2)-cyclic-phosphate 2-inositolphosphohydrolase (cyclic hydrolase), a phosphodiesterase that catalyzes the conversion of inositol-(1,2)-cyclic phosphate (cIns(1,2)P) to Ins(1)P. In addition, the enzyme also catalyzes the production of Ins(1)P from inositol (1)-methylphosphate. All of these substrates, (cIns(1,2)P, GroPIns, and inositol (1)-methylphosphate), contain a phosphodiester bond at the 1-position of the inositol ring. Additional phosphate groups on the 4- or 5-positions of the inositol ring prevent hydrolysis by cyclic hydrolase. The Km of the enzyme for GroPIns is 0.67 mM, and the Vm is 5 mumol/min/mg of protein. GroPIns competitively inhibits cIns(1,2)P hydrolysis with a Ki equal to its Km as a substrate. Hydrolysis of GroPIns and cIns(1,2)P is stimulated by MnCl2, phosphatidylserine, and [ethylenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA). However, whereas cIns(1,2)P hydrolysis is increased 5-8-fold by phosphatidylserine and EGTA only a 2-fold increase of GroPIns hydrolysis occurs under the same conditions. Hydrolysis of both GroPIns and cIns(1,2)P is inhibited by Ins(2)P; the ID50 values are 12 and 1 microM, respectively. There are significant quantities of GroPIns and Ins(2)P in 3T3 cells, indicating that these compounds that alter cIns(1,2)P hydrolase activity may modulate intracellular levels of cIns(1,2)P. Finally, we present evidence suggesting that the substrate specificity of this enzyme is altered during cell transformation.

Animals↗
PubMed1991

Sorbitol, myo-inositol, and rod outer segment phagocytosis in cultured hRPE cells exposed to glucose. In vitro model of myo-inositol depletion hypothesis of diabetic complications.

The "myo-inositol depletion hypothesis" remains a leading but still controversial contender among proposed pathogenetic mechanisms for the chronic complications of diabetes. The multifaceted interrelationships among altered tissue myo-inositol content and metabolism and tissue function have been difficult to elucidate in diabetic animal models due in part to the complex, heterogeneous nature of tissues prone to diabetic complications. The retinal pigment epithelium consists of a homogenous cell monolayer that exhibits related alterations in myo-inositol metabolism and function in diabetic animals. Nontransformed human retinal pigment epithelial (hRPE) cells, which retain their general phenotypic and morphological characteristics during monolayer culture in vitro, were examined for parallel alterations in myoinositol metabolism and cell function when grown under carefully controlled conditions in medium containing hyperglycemic concentrations of glucose. Exposure of hRPE cells to 20-40 mM glucose produced time- and dose-dependent increases in sorbitol content and decreases in myo-inositol content that were partially blocked by the aldose reductase inhibitor sorbinil. myo-Inositol was taken up by two Na-dependent transport systems, at least one of which was competitively inhibited by glucose. Exposure to 20 mM glucose impaired the ability of hRPE cells to take up human retinal rod outer segments, an important physiological function of these cells. The impairment of rod outer segment uptake by high glucose levels was prevented by an aldose reductase inhibitor or elevated medium myo-inositol that corrected the fall in myo-inositol content. Thus, hRPE cells provide a new in vitro model in which to examine the biochemical-functional interrelationships of the myo-inositol depletion hypothesis.

Adult↗
PubMed1988

Regulation of phospholipid biosynthesis in Saccharomyces cerevisiae by inositol. Inositol is an inhibitor of phosphatidylserine synthase activity.

The addition of inositol to the growth medium of Saccharomyces cerevisiae resulted in rapid changes in the rates of phospholipid biosynthesis. The partitioning of the phospholipid intermediate CDP-diacylglycerol was shifted to phosphatidylinositol at the expense of phosphatidylserine and its derivatives phosphatidylethanolamine and phosphatidylcholine. Serine at 133-fold greater concentrations than that of inositol shifted the partitioning of CDP-diacylglycerol to phosphatidylserine at the expense of phosphatidylinositol but to a much lesser degree. Kinetic experiments with pure phosphatidylserine synthase and phosphatidylinositol synthase indicated that the partitioning of CDP-diacylglycerol between phosphatidylserine and phosphatidylinositol was not governed by the affinities both enzymes have for their common substrate CDP-diacylglycerol. Instead, the main regulation of phosphatidylinositol and phosphatidylserine synthesis was through the exogenous supply of inositol. The Km of inositol (0.21 mM) for phosphatidylinositol synthase was 9-fold higher than cytosolic concentration of inositol (24 microM). The Km of serine (0.83 mM) for phosphatidylserine synthase was 3-fold below the cytosolic concentration of serine (2.6 mM). Therefore, inositol supplementation resulted in a dramatic increase in the rate of phosphatidylinositol synthesis, whereas serine supplementation resulted in little affect on the rate of phosphatidylserine synthesis. Inositol also contributed to the regulation of phosphatidylinositol and phosphatidylserine synthesis by having a direct affect on phosphatidylserine synthase activity. Kinetic experiments with pure phosphatidylserine synthase showed that inositol was a noncompetitive inhibitor of the enzyme with a Ki of 65 microM.

CDPdiacylglycerol-Serine O-Phosphatidyltransferase↗
PubMed1994

Modification at C2 of myo-inositol 1,4,5-trisphosphate produces inositol trisphosphates and tetrakisphosphates with potent biological activities.

Novel 2-position-modified D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] analogues, DL-2-deoxy-2-fluoro-myo-inositol 1,4,5-trisphosphate [DL-2F-Ins(1,4,5)P3], DL-myo-inositol 1,2,4,5-tetrakisphosphate [DL-Ins(1,2,4,5)P4], DL-scyllo-inositol 1,2,4-trisphosphate [DL-sc-Ins(1,2,4)P3], scyllo-inositol 1,2,4,5-tetrakisphosphate [sc-Ins(1,2,4,5)P4] and scyllo-inositol 1,2,4,5-tetrakisphosphorothioate [sc-Ins(1,2,4,5)PS4] were investigated for their ability to bind to the Ins(1,4,5)P3 receptor, mobilise intracellular Ca2+ stores and interact with metabolic enzymes. With the exception of sc-Ins(1,2,4,5)PS4, all the Ins(1,4,5)P3 analogues potently displaced [3H]Ins(1,4,5)P3 from its receptor in bovine adrenal cortex and were apparently potent full agonists at the Ca2+ mobilising Ins(1,4,5)P3 receptor of SH-SY5Y cells, giving respective IC50 and EC50 values of: sc-Ins(1,2,4,5)P4 (IC50 14 nM, EC50 77 nM), DL-2F-Ins(1,4,5)P3 (IC50 25 nM, EC50 105 nM), DL-Ins(1,2,4,5)P4 (IC50 26 nM, EC50 163 nM), DL-sc-Ins(1,2,4)P3 (IC50 52 nM, EC50 171 nM), compared to Ins(1,4,5)P3 (IC50 4 nM, EC50 52 nM). sc-Ins(1,2,4,5)P4 was equipotent to Ins(1,4,5)P3 for Ca2+ release making it the most potent inositol tetrakisphosphate and indeed Ins(1,4,5)P3 analogue yet characterised. In contrast, although sc-Ins(1,2,4,5)P4 (IC50 425 nM, EC50 1603 nM) was a significantly weaker ligand and agonist than Ins(1,4,5)P3, it was a partial agonist of high intrinsic activity with maximally effective concentrations releasing only about 80% of Ins(1,4,5)P3-sensitive Ca2+ stores of SH-SY5Y cells. Ins(1,4,5)P3 and sc-Ins(1,2,4,5)P4 were readily metabolised by Ins(1,4,5)P3 3-kinase and 5-phosphatase activities, DL-2F-Ins(1,4,5)P3 and DL-sc-Ins(1,2,4)P3 were resistant to 5-phosphatase, while sc-Ins(1,2,4,5)PS4 and DL-Ins(1,2,4,5)P4 were resistant to both 3-kinase and 5-phosphatase activity and were potent inhibitors of the 5-phosphatase enzyme (Ki = 300 nM and 2.9 microM, respectively). These results demonstrate that modification of the 2-position of Ins(1,4,5)P3, even with an anionic group, does not critically affect Ins(1,4,5)P3 binding interaction or Ca2+ release, suggesting that the 2-OH of Ins(1,4,5)P3 fails to interact significantly with the binding site of its receptor. However, modification remote from the crucial vicinal 4,5-bisphosphate can affect analogue efficacy in Ca2+ release.

Animals↗
PubMed1980

Metabolism of myo-[2-H]Inositol and scyllo-[R-H]Inositol in Ripening Wheat Kernels.

Injection of myo-[2-(3)H]inositol or scyllo-[R-(3)H]inositol into the peduncular cavity of wheat stalks about 2 to 4 weeks postanthesis led to rapid translocation into the spike and accumulation of label in developing kernels, especially the bran fraction. With myo-[2-(3)H]inositol, about 50 to 60% of the label was incorporated into high molecular weight cell wall substance in the region of the injection. That portion translocated to the kernels was utilized primarily for cell wall polysaccharide formation and phytate biosynthesis. A small amount was recovered as free myo-inositol and galactinol. When scyllo-[R-(3)H]inositol was supplied, most of the label was translocated into the developing kernels where it accumulated as free scyllo-inositol and O-alpha-d-galactopyranosyl-scyllo-inositol in approximately equal amount. None of the label from scyllo-[R-(3)H]inositol was utilized for either phytate biosynthesis or cell wall polysaccharide formation.

Journal Article↗
PubMed1993

Sustained effects of pilocarpine-induced convulsions on brain inositol and inositol monophosphate levels and brain morphology in young and old male rats.

Cerebral inositol and inositol monophosphates, products of phosphoinositide (PI) turnover, and neuronal injury were studied in young (10 weeks) and old (24 months) male Wistar rats after pilocarpine-induced convulsions. The goal was to explore the association between short-term cholinergic convulsions, brain PI signaling, and changes in the brain morphology in the young and the old rats. Pilocarpine caused convulsions in young rats at a dose of 300 mg/kg, whereas a dose of 175 mg/kg was required to obtain the same effect in old rats. A dose of 5 mg/kg of diazepam was used to terminate the convulsions 2 hr after their initiation; the rats were then examined on Day 5 postpilocarpine. Inositol and inositol monophosphate levels were similar in both the young and the old control rats. Pilocarpine-induced convulsions decreased cerebral inositol and increased inositol-1-phosphate in both the young and the old rats. Inositol-4-phosphate was stable in the young rats but increased in the frontal cortex and the hippocampus in the old rats. Delayed neuronal death also occurred in the convulsing rats, i.e., a variable proportion of neurons appeared shrunken with eosinophilic cytoplasm and pyknotic nuclei. The hippocampus was the most severely affected brain area. These results show that old rats are more sensitive than young ones to short-term pilocarpine convulsions, associated with sustained PI turnover, and brain injury. Mechanisms in addition to cholinergic ones are likely to be involved in the prolonged cerebral effects of cholinergic convulsions in rats.

Aging↗
PubMed1991

Lithium-induced decrease of brain inositol and increase of brain inositol-1-phosphate is transient.

The effects of a single dose of LiCl (2.5 or 10 mEq/kg) on brain inositol and inositol-1-phosphate (Ins1P), intermediates of brain phosphoinositide (PI) turnover, were determined in male Han: Wistar rats. There was a remarkable, 36-58 fold elevation of brain Li+ as the single dose of LiCl was increased 4-fold. Moreover, the accumulation of brain lithium was slow during repeated administration of LiCl. Brain lithium did not correlate with changes in brain PI turnover either after a single or repeated doses. Thus, after a single dose of LiCl the increases in brain Ins1P were much less than the decreases in brain inositol. Also, brain inositol was significantly decreased only with the high dose of LiCl whereas brain Ins1P accumulation was more prominent with the lower dose. Moreover, repeated daily doses of LiCl only transiently increased brain Ins1P at 1 and 7 d whereas inositol remained at control levels throughout the 14 d observation period. Lithium probably caused the transient decrease in brain inositol by inhibiting several enzymes, in addition to the inhibition of myo-inositol mono-phosphates, in the PI cycle. Moreover, a slow dampening down of PI turnover by lithium, possible via an inhibitory action on G-protein-coupling, may also explain the present findings.

Analysis of Variance↗
PubMed1985

Thrombin induces the rapid formation of inositol bisphosphate and inositol trisphosphate in human platelets.

Human platelets prelabeled with [3H]inositol were exposed to thrombin. The aqueous soluble inositol phosphates were separated by anion exchange column chromatography, paper chromatography or high-performance liquid chromatography, and identified by cochromatography with authentic standard substances. Thrombin immediately induces the rapid formation of inositol 1,4-bisphosphate and inositol 1,4,5-trisphosphate. Accumulation of inositol-1-monophosphate and inositol-2-monophosphate occurs later after a time lag of 10 sec. The results indicate that the phospholipase C induced polyphosphoinositide hydrolysis rather than the phosphatidylinositol hydrolysis is the triggering event for platelet activation, and support the concept of inositol 1,4,5-trisphosphate as putative second messenger.

Blood Platelets↗
PubMed2006

D-myo-inositol 1-phosphate as a surrogate of D-myo-inositol 1,4,5-tris phosphate to monitor G protein-coupled receptor activation.

Phospholipase C beta (PLC-beta)-coupled G protein-coupled receptor (GPCR) activities traditionally are assessed by measuring Ca2+ triggered by D-myo-inositol 1,4,5-trisphosphate (IP3), a PLC-beta hydrolysis product, or by measuring the production of inositol phosphate using cumbersome radioactive assays. A specific detection of IP3 production was also established using IP3 binding proteins. The short lifetime of IP3 makes this detection very challenging in measuring GPCR responses. Indeed, this IP3 rapidly enters the metabolic inositol phosphate cascade. It has been known for decades that lithium chloride (LiCl) leads to D-myo-inositol 1-phosphate accumulation on GPCR activation by inhibiting inositol monophosphatase, the final enzyme of the IP3 metabolic cascade. We show here that IP1 can be used as a surrogate of IP3 to monitor GPCR activation. We developed a novel homogeneous time-resolved fluorescence (HTRF) assay that correlates perfectly with existing methods and is easily amenable to high-throughput screening. The IP-One assay was validated on various GPCR models. It has the advantage over the traditional Ca2+ assay of allowing the measurement of inverse agonist activity as well as the analysis of PLC-beta activity in any nontransfected primary cultures. Finally, the high assay specificity for D-myo-inositol 1 monophosphate (IP1(1)) opens new possibilities in developing selective assays to study the functional roles of the various isoforms of inositol phosphates.

Animals↗
PubMed1984

Secretagogue-induced formation of inositol phosphates in rat exocrine pancreas. Implications for a messenger role for inositol trisphosphate.

The formation of inositol phosphates in response to secretagogues was studied in rat pancreatic acini preincubated with [3H]inositol. Carbachol caused rapid increases in radioactive inositol phosphate, inositol bisphosphate and inositol trisphosphate . This effect was blocked by atropine, and also elicited by caerulein, but not by ionomycin or phorbol dibutyrate. Thus phospholipase C-mediated breakdown of polyphosphoinositides, with the resulting formation of inositol phosphates, may be an early step in the stimulus-secretion coupling pathway in exocrine pancreas. Inositol trisphosphate may function as a second messenger in the exocrine pancreas, coupling receptor activation to internal Ca2+ release.

Animals↗
PubMed1990

Reduced inositol polyphosphate accumulation and inositol supply induced by lithium in stimulated cerebral cortex slices.

The ability of lithium to interfere with phosphoinositide metabolism in rat cerebral cortex slices has been examined by monitoring the accumulation of CMP-phosphatidate (CMP-PtdOH) and the reduction in Ins(1,4,5)P3 and Ins(1,3,4,5)P4 levels. A small accumulation of [14C]CMP-PtdOH was seen in slices prelabelled with [14C]cytidine and stimulated with carbachol (1 mM) or Li+ (1 mM). However, simultaneous addition of both agents for 30 min produced a 22-fold accumulation, with Li+ producing a half-maximal effect at a concentration of 0.61 +/- 0.19 mM. Kinetic studies revealed that the effects of carbachol and Li+ on CMP-PtdOH accumulation occurred with no initial lag apparent under these conditions and that preincubation with myo-inositol (10 or 30 mM) dramatically attenuated CMP-PtdOH accumulation. myo-Inositol could also attenuate the rate of accumulation of CMP-PtdOH when added 20 min after carbachol and Li+; these effects were not observed when equimolar concentrations of scyllo-inositol were added. Use of specific radioreceptor assays allowed the mass accumulations of Ins(1,4,5)P3 and Ins(1,3,4,5)P4 to be monitored. Following a lag of 5-10 min, Li+ resulted in a marked reduction in the accumulation of both inositol polyphosphates resulting from muscarinic-cholinergic stimulation. Preincubation of cerebral cortex slices with myo- (but not scyllo-) inositol delayed, but did not prevent, the reduction in the accumulation of Ins(1,4,5)P3 or Ins(1,3,4,5)P4. The results suggest that cerebral cortex, at least in vitro, is very sensitive to myo-inositol depletion under conditions of muscarinic receptor stimulation. The relationship of such depletion to the generation of inositol polyphosphate second messengers is discussed.

Animals↗