Deoxycholate induces the preferential hydrolysis of polyphosphoinositides by human platelet and rat corneal phospholipase C

Deoxycholate promotes phospholipase C degradation of endogenous phosphatidyl[3H]inositol (Pl), phosphatidyl[3H]inositol monophosphate (PIP) and phosphatidyl[3H]inositol bisphosphate (PIP2) in rat cornea and human platelets. Hydrolysis of phosphatidyl[3H]inositol significantly lags polyphospho[3H]inositide degradation. Concomitantly, formation of [3H]inositol monophosphate (IP1) lags behind [3H]inositol bisphosphate (IP2) and [3H]inositol trisphosphate (IP3) production. These results demonstrate that rat cornea and human platelet phospholipase C cause a preferential hydrolysis of the endogenous polyphosphoinositides rather than phosphatidylinositol.     

Inositol 1,3,4,5-tetrakisphosphate and not phosphatidylinositol 3,4-bisphosphate is the probable precursor of inositol 1,3,4-trisphosphate in agonist-stimulated parotid gland.

When [3H]inositol-prelabelled rat parotid-gland slices were stimulated with carbachol, noradrenaline or Substance P, the major inositol trisphosphate produced with prolonged exposure to agonists was, in each case, inositol 1,3,4-trisphosphate. Much lower amounts of radioactivity were present in the inositol 1,4,5-trisphosphate fraction separated by anion-exchange h.p.l.c. Analysis of the inositol trisphosphate head group of phosphatidylinositol bisphosphate in [32P]Pi-labelled parotid glands showed the presence of phosphatidylinositol 4,5-bisphosphate, but no detectable phosphatidylinositol 3,4-bisphosphate. Carbachol-stimulated [3H]inositol-labelled parotid glands contained an inositol polyphosphate with the chromatographic properties and electrophoretic mobility of an inositol tetrakisphosphate, the probable structure of which was determined to be inositol 1,3,4,5-tetrakisphosphate. Since an enzyme in erythrocyte membranes is capable of degrading this tetrakisphosphate to inositol 1,3,4-trisphosphate, it is suggested to be the precursor of inositol 1,3,4-trisphosphate in parotid glands.     

Evidence for the incorporation of [32P]orthophosphate into leaf inositol phospholipids

Leaf discs of brinjal, tomato, sugar cane and maize rapidly incorporated [32P]orthophosphate into total phospholipids. Analyses of the labelled lipid extracts by thin-layer chromatography, autoradiography and comparison with inositol phospholipid standards demonstrated the labelling of phosphatidylinositol monophosphate and phosphatidylinositol bisphosphate in addition to other phospholipids. The presence of polyphosphoinositides was further confirmed by deacylation of phosphatidylinositol monophosphate and phosphatidylinositol bisphosphate and separation of the water-soluble products, glycerophosphoinositol phosphate and glycerophosphoinositol bisphosphate by formate exchange chromatography. Incorporation of [32P]orthophosphate into inositol phospholipids was time-dependent, with monoester phosphate groups attaining isotopic equilibrium within 90 min of incubation. After 2 h, incorporation of label into phosphatidylinositol, phosphatidylinositol monophosphate and phosphatidylinositol bisphosphate was about 15, 10 and 3%, respectively, of the total phospholipids. The ratio of radioactivity in phosphatidylinositol/phosphatidylinositol monophosphate/phosphatidylinositol bisphosphate was about 5:5:1 in brinjal leaves. However, this ratio may be an overestimate of the amounts of inositol phospholipids present, as other lysophospholipids may comigrate with standards.     

The Role of Phosphatidylinositol 4,5 Bisphosphate Breakdown in Cell-Surface Receptor Activation

Abstract

The activation of Ca²⁺-mobilising receptors on hepatocytes and many other cells leads to a prompt reduction in the cellular content of inositol phospholipids. The primary event which underlies these changes is, most probably, a phospholipase C-catalysed attack upon phosphatidylinositol 4,5 bisphosphate. The receptor-mediated breakdown of this lipid in stimulated cells is: (i) not mediated by an increase in cytosol [Ca²⁺] and (ii) closely coupled to receptor occupation. Phosphatidylinositol 4,5 bisphosphate degradation may be studied by measuring the appearance of the water-soluble product, inositol trisphosphate (and its metabolites: inositol bisphosphate and inositol monophosphate), in stimulated cells. Recent evidence indicates that inositol trisphosphate and the lipid soluble product of phosphatidylinositol 4,5 bisphosphate breakdown, 1,2 diacylglycerol, may act as ‘second messengers’ which mediate the effects of many extracellular signals in stimulated cells.     

Phosphatidylinositol Metabolism During In Vitro Hypoxia

The effects of in vitro histotoxic hypoxia (0.5 mM KCN) on potassium-stimulated phosphatidylinositol turnover were determined. In rat cortical slices that were prelabeled with [2-3H]inositol, depolarization with 60 mM KCl increased [2-3H]inositol monophosphate and [2-3H]inositol bisphosphate accumulation in a Ca2+-dependent manner. At early times (10 s and 1 min), histotoxic hypoxia enhanced potassium-stimulated [2-3H]inositol monophosphate and inositol bisphosphate accumulation. Under basal conditions, hypoxia did not alter the accumulation of [2-3H]inositol phosphates. These results are consistent with the following hypothesis. The hypoxic-induced increase in cytosolic free calcium that we reported previously may lead to the early stimulation of inositol phosphates formation during hypoxia through activation of phospholipase C. The impairment of inositol phosphates formation during more prolonged hypoxia may be due to negative feedback regulation of the phosphatidylinositol cascade by protein kinase C or to a reduction in ATP levels.     

Requirement for calcium ions in acetylcholine-stimulated phosphodiesteratic cleavage of phosphatidyl-myo-inositol 4,5-bisphosphate in rabbit iris smooth muscle

1. The mechanism of acetylcholine-stimulated breakdown of phosphatidyl-myo-inositol 4,5-bisphosphate and its dependence on extracellular Ca(2+) was investigated in the rabbit iris smooth muscle. 2. Acetylcholine (50mum) increased the breakdown of phosphatidylinositol bisphosphate in [(3)H]inositol-labelled muscle by 28% and the labelling of phosphatidylinositol by 24% of that of the control. Under the same experimental conditions there was a 33 and 48% increase in the production of (3)H-labelled inositol trisphosphate and inositol monophosphate respectively. Similarly carbamoylcholine and ionophore A23187 increased the production of these water-soluble inositol phosphates. Little change was observed in the (3)H radioactivity of inositol bisphosphate. 3. Both inositol trisphosphatase and inositol monophosphatase were demonstrated in subcellular fractions of this tissue and the specific activity of the former was severalfold higher than that of the latter. 4. The acetylcholine-stimulated production of inositol trisphosphate and inositol monophosphate was inhibited by atropine (20mum), but not tubocurarine (100mum); and it was abolished by depletion of extracellular Ca(2+) with EGTA, but restored on addition of low concentrations of Ca(2+) (20mum). 5. Calcium-antagonistic agents, such as verapamil (20mum), dibenamine (20mum) or La(3+) (2mm), also abolished the production of the water-soluble inositol phosphates in response to acetylcholine. 6. Release of inositol trisphosphate from exogenous phosphatidylinositol bisphosphate by iris muscle microsomal fraction (;microsomes') was stimulated by 43% in the presence of 50mum-Ca(2+). 7. The results indicate that increased Ca(2+) influx into the iris smooth muscle by acetylcholine and ionophore A23187 markedly activates phosphatidylinositol bisphosphate phosphodiesterase and subsequently increases the production of inositol trisphosphate and its hydrolytic product inositol monophosphate. The marked increase observed in the production of inositol monophosphate could also result from Ca(2+) activation of phosphatidylinositol phosphodiesterase. However, there was no concomitant decrease in the (3)H radioactivity of this phospholipid.     

Polyphosphoinositides are present in plant tissue culture cells

Polyphosphoinositides have been isolated from wild carrot cells grown in suspension culture. This is the first report of polyphosphoinositides in plant cells. The phospholipids were identified by comigration with known standards on thin-layer plates. After overnight labeling of the cells with myo-[2-3H] inositol, the phosphoinositides as percent recovered inositol were 93% phosphatidylinositol., 3.7% lysophosphatidylinositol, 1.7% phosphatidylinositol monophosphate, 0.8% phosphatidylinositol bisphosphate.     

Metabolism and functions of inositol phosphates

Activation of Ca2+-mobilizing receptors rapidly increases the cytoplasmic Ca2+ concentration both by releasing Ca2+ stored in endoplasmic reticulum and by stimulating Ca2+ entry into the cells. The mechanism by which Ca2+ release occurs has recently been elucidated. Receptor activation of phospholipase C results in the hydrolysis of the plasma membrane lipid, phosphatidylinositol 4,5-bisphosphate (PIP2), to yield two intracellular messengers, diacylglycerol (DAG) and (1,4,5)inositol trisphosphate [(1,4,5)IP3]. DAG remains in the plasma membrane where it stimulates protein phosphorylation via the phospholipid-dependent protein kinase C. (1,4,5)IP3 diffuses to and interacts with specific sites on the endoplasmic reticulum to release stored Ca2+. Receptor stimulation of phospholipase C appears to be mediated by one or more guanine nucleotide-dependent regulatory proteins by a mechanism analogous to hormonal activation of adenylyl cyclase. The actions of (1,4,5)IP3 on Ca2+ mobilization are terminated by two metabolic pathways, sequential dephosphorylation to inositol bisphosphate (IP2), inositol monophosphate (IP) and inositol or by phosphorylation to inositol tetrakisphosphate (IP4) and sequential dephosphorylation to different inositol phosphates. A sustained cellular response also requires Ca2+ entry into the cell from the extracellular space. The mechanism by which hormones increase Ca2+ entry is not known; a recent proposal involving movement of Ca2+ through the endoplasmic reticulum, possibly regulated by IP4, will be considered here.     

Bombesin stimulates inositol polyphosphate production in GH4C1 pituitary tumor cells: comparison with TRH

The hormones bombesin and thyrotropin-releasing hormone (TRH) stimulated formation of inositol- monophosphate, bisphosphate, trisphosphate and tetrakisphosphate with parallel time courses in GH4C1 cells, while a more polar inositol polyphosphate peak, consisting of inositol-pentakisphosphate and perhaps also inositol-hexakisphosphate, was unaffected by either hormone. Although bombesin and TRH had similar potencies in stimulating inositol trisphosphate production (Km = 30 nM and 40 nM, respectively), TRH was significantly more efficacious than bombesin. Maximal stimulation of inositol-1,4,5-trisphosphate formation by TRH was not further increased by addition of a maximally effective dose of bombesin, suggesting that the two hormones act through stimulation of a common pool of phospholipase C, and this enzyme pool can be fully stimulated by TRH, alone.     

Antigen-stimulated metabolism of inositol phospholipids in the cloned murine mast-cell line MC9.

Cells of the murine mast-cell clone MC9 grown in suspension culture were sensitized with an anti-DNP (dinitrophenol) IgE and subsequently prelabelled by incubating with [32P]Pi. Stimulation of these cells with DNP-BSA (bovine serum albumin) caused marked decreases in [32P]polyphosphoinositides (but not [32P]phosphatidylinositol) with concomitant appearance of [32P]phosphatidic acid. Whereas phosphatidylinositol monophosphate levels returned to baseline values after prolonged stimulation, phosphatidylinositol bisphosphate levels remained depressed. Stimulation of sensitized MC9 cells with DNP-BSA increased rates of incorporation of [32P]Pi into other phospholipids in the order: phosphatidylcholine greater than phosphatidylinositol greater than phosphatidylethanolamine. In sensitized cells prelabelled with [3H]inositol, release of inositol monophosphate, inositol bisphosphate and inositol trisphosphate, was observed after stimulation with DNP-BSA. When Li+ was added to inhibit the phosphatase activity that hydrolysed the phosphomonoester bonds in the sugar phosphates, greater increases were observed in all three inositol phosphates, particularly in inositol trisphosphate. The IgE-stimulated release of inositol trisphosphate was independent of the presence of extracellular Ca2+. In addition, the Ca2+ ionophore A23187 caused neither the decrease in [32P]polyphosphoinositides nor the stimulation of the release of inositol phosphates. These results demonstrate that stimulation of the MC9 cell via its receptor for IgE causes increased phospholipid turnover, with effects on polyphosphoinositides predominating. These data support the hypothesis that hapten cross-bridging of IgE receptors stimulates phospholipase C activity, which may be an early event in stimulus-secretion coupling of mast cells. The results with the Ca2+ ionophore A23187 indicate that an increase in intracellular Ca2+ alone is not sufficient for activation of this enzyme.     

Stimulation of Inositol Incorporation into Lipids of PC 12 Cells by Nerve Growth Factor and Bradykinin

The effects of bradykinin (BK) and lithium on the phosphatidylinositol cycle were examined in PC12 cells cultured for 20 h in the presence [PC12(+)] or in the absence [PC12(-)] of nerve growth factor (NGF). BK (1 microM) induced a small stimulation of the incorporation of myo-[2-3H]inositol into the lipids of PC12(-) cells and a three- to fourfold stimulation of such incorporation into the lipids of PC12 (+) cells. About 15 h of incubation with NGF and greater than 10 min of incubation with BK were needed for maximal stimulation of inositol incorporation by BK. In the presence of 25 mM LiCl, BK stimulated the inositol monophosphate levels nine-fold in PC12 (-) and 30-fold in PC12 (+) cells. After incubation for 20 h with NGF, an increased binding of [3H]BK to the PC12 (+) cells was observed at 4 degrees C. Exposure of the cells for 30 min to 25 mM LiCl enhanced the effect of BK on the inositol incorporation into total inositol lipids, especially in PC12(+) cells. In these cells, LiCl in the presence of BK also increased several-fold the intracellular levels of inositol bisphosphate and inositol trisphosphate.     

Prolactin induces the formation of inositol bisphosphate and inositol trisphosphate in cultured mouse mammary gland explants

The effect of prolactin on [3H]inositol metabolism in cultured mouse mammary gland explants derived from 12-14-day pregnant mice was determined. In mammary gland explants that were prelabeled by culturing the tissues with 3 microCi/ml myo-[3H]inositol for 48 h, the levels of 3H in inositol derivatives were determined. The temporal effect of prolactin on the quantity of 3H present in phosphatidylinositol (PI), phosphatidylinositol monophosphate (PIP), phosphatidylinositol bisphosphate (PIP2) and various inositol phosphate containing fractions were examined. Prolactin significantly stimulated the accumulation of 3H label in inositol monophosphate (IP1), inositol bisphosphate (IP2) and inositol trisphosphate (IP3) 1-3 h after addition of prolactin. An effect of prolactin on the accumulation of inositol derivatives was not apparent at prolactin-exposure periods of less than 60 min; nor was an effect of prolactin apparent when exposure periods of 4 h or longer were employed. Prolactin did not significantly decrease the 3H label in PI, PIP or PIP2 except at 1 and 2 h. These data when considered with other apropos studies are compatible with the conclusion that the turnover of inositol lipid derivatives may be involved in the mechanism by which prolactin regulates metabolic processes in the mammary gland. The primary action of prolactin on mammary cells, however, would not appear to involve its action on the metabolism of the inositol derivatives in view of the extended time required (1 h) before effects of prolactin on perturbations of inositide metabolism are manifested.     

Stretch increases inositol trisphosphate and inositol tetrakisphosphate in cultured pulmonary vascular smooth muscle cells.

There are no reports of the effect of stretch on inositol phosphates in smooth muscle. Phosphoinositide and inositol phosphate metabolism was studied in cultured rat vascular smooth muscle cells subjected to stretching. The masses of inositol trisphosphate and tetrakisphosphate increased (+34 +/- 7% and +58 +/- 12%, respectively; p less than 0.001) after 25 s of a single 20% stretch and had returned to control levels by 45 s; phosphatidylinositol, phosphatidylinositol phosphate and bisphosphate did not change. Repetitive stretch did not alter the masses of any of the compounds. A single stretch also increased 45Ca2+ efflux (+52 +/- 5%, p less than 0.01). These data suggest that stretch of cultured vascular smooth muscle can elicit a rapid, short-lived increase in inositol phosphates, which may subsequently affect Ca2+.     

Metabolism of inositol phosphates in parotid cells: implications for the pathway of the phosphoinositide effect and for the possible messenger role of inositol trisphosphate

Rat parotid acinar cells were used to investigate the time course of formation and breakdown of inositol phosphates in response to receptor-active agents. In cells preincubated with [3H]inositol and in the presence of 10 mM LiCl (which blocks hydrolysis of inositol phosphate), methacholine (10(-4)M) caused a substantial increase in cellular content of [3H]inositol phosphate, [3H]inositol bisphosphate and [3H]inositol trisphosphate. Subsequent addition of atropine (10(-4) M) caused breakdown of [3H]inositol trisphosphate and [3H]inositol bisphosphate and little change in accumulated [3H]inositol phosphate. The data could be fit to a model whereby inositol trisphosphate and inositol bisphosphate are formed from phosphodiesteratic breakdown of phosphatidylinositol bisphosphate and phosphatidylinositol phosphate respectively, and inositol phosphate is formed from hydrolysis of inositol bisphosphate rather than from phosphatidyl-inositol. Consistent with this model was the finding that [3H]inositol trisphosphate and [3H]inositol bisphosphate levels were substantially increased in 5 sec while an increase in [3H]inositol phosphate was barely detectable at 60 sec. These results indicate that in the parotid gland the phosphoinositide cycle is activated primarily by phosphodiesteratic breakdown of the polyphosphoinositides rather than phosphatidyl-inositol. Also, the results show that formation of inositol trisphosphate is probably sufficiently rapid for it to act as a second messenger signalling internal Ca2+ release in this tissue.     

Chemoattractant and guanosine 5''-[gamma-thio]triphosphate induce the accumulation of inositol 1,4,5-trisphosphate in Dictyostelium cells that are labelled with [3H]inositol by electroporation.

The analysis of the inositol cycle in Dictyostelium discoideum cells is complicated by the limited uptake of [3H]inositol (0.2% of the applied radioactivity in 6 h), and by the conversion of [3H]inositol into water-soluble inositol metabolites that are eluted near the position of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] on anion-exchange h.p.l.c. columns. The uptake was improved to 2.5% by electroporation of cells in the presence of [3H]inositol; electroporation was optimal at two 210 microseconds pulses of 7 kV. Cells remained viable and responsive to chemotactic signals after electroporation. The intracellular [3H]inositol was rapidly metabolized to phosphatidylinositol and more slowly to phosphatidylinositol phosphate and phosphatidylinositol bisphosphate. More than 85% of the radioactivity in the water-soluble extract that was eluted on Dowex columns as Ins(1,4,5)P3 did not co-elute with authentic [32P]Ins(1,4,5)P3 on h.p.l.c. columns. Chromatography of the extract by ion-pair reversed-phase h.p.l.c. provided a good separation of the polar inositol polyphosphates. Cellular [3H]Ins(1,4,5)P3 was identified by (a) co-elution with authentic [32P]Ins(1,4,5)P3 and (b) degradation by a partially purified Ins(1,4,5)P3 5-phosphatase from rat brain. The chemoattractant cyclic AMP and the non-hydrolysable analogue guanosine 5'-[gamma-thio]triphosphate induced a transient accumulation of radioactivity in Ins(1,4,5)P3; we did not detect radioactivity in inositol 1,3,4-trisphosphate or inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4]. In vitro, Ins(1,4,5)P3 was metabolized to inositol 1,4- and 4,5-bisphosphate, but not to Ins(1,3,4,5)P4 or another tetrakisphosphate isomer. We conclude that Dictyostelium has a receptor- and G-protein-stimulated inositol cycle which is basically identical with that in mammalian cells, but the metabolism of Ins(1,4,5)P3 is probably different.     

Activation of phosphoinositide hydrolysis by nerve growth factor and lysophosphatidylserine in rat peritoneal mast cells

Histamine secretion in rat peritoneal mast cells stimulated by nerve growth factor requires a synergistic signal delivered by lysophosphatidylserine. To study the signal-transducing system activated by these compounds, phospholipid metabolism has been investigated in these cells. Phospholipid labeling with 32PO4 reveals a 5-9-fold stimulation of phosphatidic acid, phosphatidylinositol and phosphatidylcholine synthesis. Increased synthesis of phosphatidylinositol is also monitored using [3H]inositol incorporation. When [3H]inositol-labeled mast cells are incubated in the presence of Li+, nerve growth factor and lysophosphatidylserine enhance the accumulation of inositol monophosphate, inositol bisphosphate and inositol trisphosphate. Similar to the induced histamine release, accumulation of inositol phosphates (a) does not occur when the two agonists are added separately; (b) is inhibited when lysophosphatidyl-L-serine is replaced by lysophosphatidyl-D-serine; and (c) is enhanced in the presence of extracellular Ca2+. The data suggest that the interactive stimulus of nerve growth factor and lysophosphatidylserine is transmitted through the polyphosphoinositide-phospholipase C system.     

Thrombin's effects on osteoblastic cells. I. Cytosolic calcium and phosphoinositides

Thrombin, a blood coagulation factor, has been shown to be a very effective in vitro bone resorbing agent whose mechanism of action on osteoblastic cells remains to be elucidated. In the present study, the effects of highly purified human thrombin on Saos-2 and G292 cells, two human osteoblast-like osteosarcoma cell lines, were investigated. Thrombin (0.6-16 U/ml) caused a significant, dose-dependent increase in osteoblastic cell proliferation. Thrombin also elicited a dose-dependent increase in cytosolic calcium concentration in both Saos-2 and G292 cells (maximal increases were 38% and 200% over baseline, respectively). Addition of thrombin to the osteoblast-like cells resulted in significant time- and dose-dependent changes in phosphoinositide levels: the percentage of inositol monophosphate levels were decreased, whereas the percentage of inositol bisphosphate, inositol trisphosphate and inositol tetrakisphosphate levels were increased. The relative magnitude of the changes in phosphoinositide levels was similar to the changes in cytosolic calcium concentration. These results suggest that thrombin's mechanism of action on bone cells may involve increases in cytosolic calcium levels and in phosphoinositide metabolism.     

Lithium Selectively Inhibits Muscarinic Receptor-Stimulated Inositol Tetrakisphosphate Accumulation in Mouse Cerebral Cortex Slices

The in vitro effects of Li on agonist- and depolarization-stimulated accumulation of inositol phosphates were determined in mouse cerebral cortex slices. Of the agents examined, only the cholinergic agonist carbachol produced a significant accumulation of inositol tetrakisphosphate (InsP4) in the absence of Li. Lithium at 5 mM enhanced the accumulation of inositol monophosphate (InsP1) and inositol bisphosphate (InsP2) due to all the stimuli used and potentiated inositol trisphosphate (InsP3) accumulation due to histamine and noradrenaline, although at lower Li concentrations, carbachol-stimulated InsP3 accumulation was reduced. Li also enhanced InsP4 accumulation in the presence of noradrenaline, histamine, and elevated KCl level but, in marked contrast, reduced carbachol-stimulated InsP4 accumulation with an IC50 of 100 microM. There was a significant time delay between the initiation of carbachol stimulation and the beginning of the InsP4 inhibition due to Li. The phorbol ester 4 beta-phorbol 12 beta-myristate 13 alpha-acetate did not mimic the effects of Li. The results suggest that muscarinic receptor-mediated InsP4 production might be one of the targets for the therapeutic action of Li.     

A sensitive method for the quantification of the mass of inositol phosphates using gas chromatography-mass spectrometry

A sensitive method to directly measure the mass of inositol phosphates from biologic samples is described. The procedure uses ammonium sulfate gradient elution anion exchange column chromatography to isolate inositol monophosphate, bisphosphate, trisphosphate, and tetrakisphosphate. The isolated fractions are dephosphorylated and subsequently desalted by a novel approach using solid barium hydroxide in a 1:1 stoichiometric ratio to the amount of ammonium sulfate present in the dephosphorylated sample. The myo-inositol derived from each inositol phosphate species was quantified by stable isotope dilution gas chromatography-mass spectrometry of the hexakis(trimethylsilyl) derivative using hexadeutero-myo-inositol as the internal standard. The applicability and sensitivity of this method are illustrated by measuring the mass of individual inositol phosphates in isolated adult canine cardiac myocytes.     

The effects of different hexachlorocyclohexanes and cyclodienes on glucose uptake and inositol phospholipid synthesis in rat brain cortex.

The inositol lipids from rat brain miniprisms were deacylated and separated by anion-exchange chromatography in order to determine whether or not gamma-hexachlorocyclohexane (gamma-HCH, lindane) and related compounds affect the different phosphatidylinositols. The incorporation of myo-[2-3H]inositol into phosphatidylinositol, phosphatidylinositol monophosphate and phosphatidylinositol bisphosphate were inhibited by lindane and its delta-HCH isomer. The inhibitory effects on phosphatidylinositol synthesis are not prominent in alpha-HCH and they are not significant with the beta-HCH and cyclodienes. The results presented here indicate that the inhibitory effect of lindane and delta-HCH on the phosphatidylinositol metabolism was no exclusively due to an interference with glucose transport. Lindane-treated miniprisms showed decreased myo-[2-3H]inositol uptake and, proportionately, an even greater inhibition of inositol phospholipid synthesis. Cellular uptake can, therefore, not account for all of the lindane inhibition.