Comparison of muscarinic and alpha-adrenergic receptors in rat atria based on phosphoinositide turnover

The effects of muscarinic and alpha-adrenergic receptor stimulation on phosphoinositide turnover in rat atria have been compared. Despite the similar densities of muscarinic receptors in rat left and right atria, 0.1 mM carbachol increased [32P]phosphate incorporation into phosphatidylinositol (PI) by 35% (p less than 0.05) in left atria but had no effect in right atria. By contrast to the small muscarinic receptor effect, stimulation of alpha 1-adrenergic receptors by 0.1 mM methoxamine produced a more than two fold increase in [32P]phosphate incorporation into PI in both left and right atria, despite the reported smaller density of alpha-adrenergic receptors in rat atria compared to muscarinic receptors. Enhanced phosphate labelling by methoxamine did not occur in phospholipids other than PI, and was blocked by the alpha-adrenergic antagonist, phentolamine (20 microM). The results indicate that the majority of the muscarinic receptors in rat atria are not coupled to phosphoinositide turnover. If indeed the observed enhancement in [32P]-phosphate labelling by carbachol reflects phosphoinositide turnover, and assuming equal coupling efficiencies of muscarinic and adrenergic receptors, it is calculated that not more than 2% of the muscarinic receptors in rat left atria are coupled to this response.     

Stimulation by prostaglandin F2 alpha of phosphatidic acid-phosphatidylinositol turnover in rat luteal cells

Prostaglandin F2 alpha (PGF 2 alpha) causes a rapid and marked increase of [32P]-orthophosphate incorporation into phosphatidylinositol (PI) and phosphatidic acid (PA) in rat luteal cells in culture. The incorporation of radioactivity is increased as early as 2 and 5 min after PGF 2 alpha addition into PA and PI, respectively, and by 10 min has reached a 2-fold stimulation over control in both lipid moieties. The labeling of other phospholipids is not affected. PGF 2 alpha exerts its stimulatory effect at an ED50 value of approximately 200 and 60 nM on PI and PA labeling, respectively. By contrast, human chorionic gonadotropin has no effect alone and does not interfere with the PGF 2 alpha-induced stimulation of PA-PI labeling. The striking similarity between the effects of PGF 2 alpha and LHRH on PA-PI labeling suggests that the two agents may exert their direct action on the corpus luteum via a common intracellular mechanism involving acidic phospholipid metabolism.     

The effects of ACTH, serotonin, K+ and angiotensin analogues on 32P incorporation into phospholipids of the rat adrenal cortex: basis for an assay method using zona glomerulosa cells

The effects of various concentrations of serotonin, ACTH, K+, angiotensin II (AII), angiotensin III (AIII) and [Sar1]angiotensin II (SAII) on steroidogenesis and the incorporation of 32P (after preincubation to near equilibrium with the ATP pool) into phosphatidylinositol (PI), phosphatidic acid (PA) and phosphatidylcholine (PC) in a preparation of capsular cells from rat adrenals, consisting of 95% zona glomerulosa (z.g.) and 5% zona fasciculata plus reticularis (z.f.r.) cells, were investigated. Serotonin and ACTH stimulated steroidogenesis in the usual manner but had little or no effect on 32P incorporation into any of the three phospholipids. However, AII, AIII and SAII stimulated steroidogenesis and also 32P incorporation into PA and PI (maximally to about 280% of control values) but not into PC. These results taken together with other data on effects on the cAMP output and Ca2+ fluxes of z.g. cells suggest that stimulation by ACTH and serotonin is mediated by cAMP as second messenger. However, the angiotensins probably act through Ca2+, with associated changes in phospholipid metabolism. The 32P incorporation into PA as a function of lg concentration of AII was linear and showed a reasonable index of precision (0.36 +/- 0.03, eight experiments, 0.23 +/- 0.02 for a further eight experiments) and correlation with steroidogenesis. The corresponding incorporation into PI showed a maximum effect and a much poorer index of precision (1.02 +/- 0.30 (4.69 +/- 3.7] over the same full range of AII concentration used. The effects of AIII and SAII showed similar characteristics for 32P incorporation into both PA and PI, but, as for stimulation of steroidogenesis, at higher concentrations for AIII than for AII. The effects of different doses of AII, AIII and ACTH on the corticosterone output and 32P incorporation into PA, PI and PC of a preparation of cells, consisting of more than 98% z.f.r. cells, from rat decapsulated adrenals were also studied. ACTH, at low doses, which nevertheless markedly stimulated corticosterone output, had a small (maximally to about 125% of control values) but significant effect on 32P incorporation into PA, PI and PC. The maximum effect was usually at about 10(-10) M ACTH and was not significant at 10(-8) M.     

Differential effect of progesterone on the labeling of phosphatidylinositol with [3H]inositol and [32P]phosphate in the uterus of the estrogen-treated ovariectomized rat

In rat uterine mince incubated in vitro [3H]inositol was found to be incorporated into phosphatidylinositol (PI) predominantly via a pathway which could be markedly and dose dependently activated with Mn2+ (0.1-10 mM) and inhibited by Ca2+ (1-10 mM). These ions had no effect on the incorporation of [32P]phosphate (32P) into PI indicating a distinct inositol-exchange mechanism for the labeling of PI with [3H]inositol. Treatment of ovariectomized rats for 5 days with 2 micrograms estradiol dipropionate (EDP) increased about 3-fold (when measured in the presence of 1 mM Mn2+) and 4-5-fold (when measured in the presence of 1 mM Ca2+) the inositol-exchange activity in the rat uterus, and these effects were suppressed by 40 and 30% respectively by the concomitant administration of 2 mg progesterone (P). EDP alone or in combination with P increased to the same extent (by a factor of 2-3) the rate of labeling with 32P of phosphatidylcholine (PC), phosphatidylethanolamine (PE) and plasmenylethanolamine (PmE). The labeling rate of PI was increased 1.5-1.7-fold by treatment with EDP and this increase was selectively augmented further to about 2.5-fold by the simultaneous administration of P. Treatment with P alone had no significant effect on the incorporation of either labeled precursor. Steroid hormone treatments had no effect on the amount of these phospholipids in 100 mg uterine tissue, but they increased about 1.7-fold the rate of labeling of ATP with 32P. We conclude that P, when administered together with estradiol, regulates differentially the turnover of the inositol and phosphate moieties of PI with possible physiological consequences.     

Mechanisms of gonadotropin-releasing hormone and prostaglandin action on luteal cells

Both gonadotropin-releasing hormone (GnRH) and prostaglandin F2 alpha (PGF2 alpha) can inhibit cAMP and progesterone production in the corpus luteum; however, their mechanism of action is not known. GnRH or PGF2 alpha causes a rapid and marked increase of labelling of phosphatidylinositol (PI) and phosphatidic acid (PA) in rat luteal cells in culture. The incorporation of radioactivity is increased as early as 2 and 5 min into PA and PI, respectively. The labelling of the other phospholipids is not affected. GnRH and PGF2 alpha exert their stimulatory effects on PA-PI turnover at a mean effective dose value of ca. 15 and 100 nM, respectively. Their effects appeared to be additive when both agents were present in the same incubations. Interestingly, addition of the calcium ionophore A23187 also causes a dramatic increase of PA-PI turnover in luteal cells. By contrast, human chorionic gonadotropin and isoproterenol, agents that stimulate cAMP and progesterone production in luteal cells, as well as PGE2 (1 microM), all fail to alter phospholipid labelling; dibutyryl or 8-bromo-cAMP (2-5 mM) actually attentuates the GnRH or PGF2 alpha effect on PI and PA. A very similar PA-PI response to GnRH and PGF2 alpha has also been observed using rat granulosa cells in culture. It seems that following their binding to membrane receptors, GnRH and PGF2 alpha may share a common mechanism in the ovarian cell, possibly involving the stimulation of PA-PI metabolism.     

Vasopressin rapidly stimulates phosphatidic acid-phosphatidylinositol turnover in rat anterior pituitary cells

In cultured rat anterior pituitary cells, the agonist [Asu1,6, Arg8]vasopressin (AVP-A) increased by 1.5-fold 32Pi incorporation into phosphatidic acid (PA), as early as 15 s after its addition. Increased phosphatidylinositol (PI) labeling became significant 4 min after AVP-A addition. Dose-response measurements with AVP-A showed ED50 values of 76 and 62 nM for PA and PI labeling, respectively. Peptide corticotropin-releasing factor (CRF) (0.1 microM) did not affect the stimulatory effect of AVP-A on PA and PI labeling. These data suggest that stimulation of PI metabolism in corticotrophs may be one of the early events involved in the stimulation of ACTH release induced by vasopressin.     

Effects of luteinizing hormone on phosphoinositide metabolism in rat granulosa cells

Rat granulosa cells isolated from mature Graafian follicles were incubated with luteinizing hormone under various conditions in order to follow the synthesis and degradation of phospholipids. During acute incubations, luteinizing hormone provoked rapid and concentration-dependent increases in the incorporation of 32PO4 into phosphatidic acid, phosphatidylinositol, and the polyphosphoinositides. Similarly, luteinizing hormone provoked increases in labeling of phosphatidylinositol and the polyphosphoinositides when granulosa cells were incubated with myo-[2-3H]inositol. When granulosa cells were prelabeled with 32PO4 in order to label phosphatidylinositol to constant specific radioactivity (4 h), luteinizing hormone treatment significantly increased 32P-phosphatidylinositol levels (23%). Comparable increases (27%) in the cellular concentrations of phosphatidylinositol were observed in response to luteinizing hormone. In pulse-chase experiments employing 32PO4 - or [3H]inositol-prelabeled cells, luteinizing hormone did not alter phospholipid degradation. In addition, luteinizing hormone did not stimulate degradation of polyphosphoinositides. These results demonstrate that: (a) luteinizing hormone has selective effects on phospholipid metabolism in rat granulosa cells which involve phosphatidic acid, phosphatidylinositol, and the polyphosphoinositides, (b) luteinizing hormone increases net levels of phosphatidylinositol and presumably phosphatidic acid and the polyphosphoinositides, and (c) luteinizing hormone does not increase phospholipid degradation. Our findings suggest that luteinizing hormone provokes increases in de novo synthesis of phosphatidylinositol in rat granulosa cells. These changes in phospholipid metabolism may be important for steroidogenesis and other enzymatic processes during treatment with luteinizing hormone.     

Effects of prostaglandins and luteinizing hormone-releasing hormone on phosphatidic acid-phosphatidylinositol labeling in rat granulosa cells

In cultures of rat granulosa cells, luteinizing hormone-releasing hormone (LHRH) increases 32P incorporation into both phosphatidylinositol (PI) and phosphatidic acid (PA). After 20 min, the level of radioactivity was three- to four-fold (p less than 0.01) above control in the PI and PA fractions, respectively. The stimulatory effect of LHRH on 32P incorporation was limited to PI and PA. Similar to the effects of LHRH, a rapid and marked increase of 32P incorporation into both PI and PA is observed upon addition of prostaglandin F2 alpha (PGF2 alpha) (10(-5)M) to rat granulosa cells. Incorporation of radioactivity into PA was already increased (p less than 0.05) by 2 min following PGF2 alpha addition, while the increase in 32P-labeled PI became significant (p less than 0.01) by 5 min. In contrast to PGF2 alpha, the labeling of PI and PA following the addition of PGE2 (10(-5)M) was not significantly different from control levels during the entire 10 min of incubation. The sensitivity of the increased PA-PI labeling induced by LHRH and PGF2 alpha is compared in another experiment. After 20 min incubation 10(-6)M LHRH increased PI and PA labeling by six- and four-fold, respectively. Although the effect of PGF2 alpha is less than that of LHRH, 10(-5)M PGF2 alpha significantly (p less than 0.01) increased PI and PA labeling by three- and two-fold, respectively. By contrast, 10(-6)M PGE2 failed to affect 32P incorporation into the various phospholipid fractions, but a small enhancement (p less than 0.05) of PI and PA labeling was observed only at 10(-5)M PGE2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mechanism of action of TRH: involvement of the phosphatidylinositol (PI) response in the action of TRH in rat anterior pituitary cells

In order to further investigate the molecular basis for the action of TRH, experiments were performed to examine the ability of rat anterior pituitary cells to incorporate [32P]orthophosphate into phospholipids in response to the neurohormone. Addition of 0.1 microM TRH rapidly stimulates the phosphatidylinositol (PI) response. The incorporation of radioactivity into PA and PI is increased as early as in 2 and 5 min after TRH addition, respectively, and reaches 40 and 140% above control, respectively, at 20 min. By contrast, 0.1 microM somatostatin is inactive. TRH exerts its stimulatory effect at an ED50 value of ca. 10 nM. The present results suggest that the PI response is an early event associated with the action of TRH in the anterior pituitary gland.     

Phosphoinositide hydrolysis increase by angiotensin-(1--7) in neonatal rat brain

Angiotensin (Ang)-(1–7) is an endogenous peptide hormone of the renin–angiotensin system which exerts diverse biological actions, some of them counterregulate Ang II effects. In the present study potential effect of Ang-(1–7) on phosphoinositide (PI) turnover was evaluated in neonatal rat brain. Cerebral cortex prisms of seven-day-old rats were preloaded with [³H]myoinositol, incubated with additions during 30 min and later [³H]inositol-phosphates (IPs) accumulation quantified. It was observed that PI hydrolysis enhanced 30% to 60% in the presence of 0.01 nM to 100 nM Ang-(1–7). Neither 10 nM [D-Ala⁷]Ang-(1–7), an Ang-(1–7) specific antagonist, nor 10 nM losartan, an angiotensin II type 1 (AT₁) receptor antagonist, blocked the effect of 0.1 nM Ang-(1–7) on PI metabolism. The effect of 0.1 nM Ang-(1–7) on PI hydrolysis was not reduced but it was even significantly increased in the simultaneous presence of [D-Ala⁷]Ang-(1–7) or losartan. PI turnover enhancement achieved with 0.1 nM Ang-(1–7) decreased roughly 30% in the presence of 10 nM PD 123319, an angiotensin II type 2 (AT₂) receptor antagonist. The antagonists alone also enhanced PI turnover. Present findings showing an increase in PI turnover by Ang-(1–7) represent a novel action for this peptide and suggest that it exerts a function in this signaling system in neonatal rat brain, an effect involving, at least partially, angiotensin AT₂ receptors.     

Effects of lithium on angiotensin-stimulated phosphatidylinositol turnover and aldosterone production in adrenal glomerulosa cells: a possible causal relationship

Turnover of 32P-labelled phosphatidylinositol (PI) was examined in isolated adrenal glomerulosa cells. Increased incorporation of [32P]phosphate into PI in response to angiotensin II was completely prevented by Li+. A simultaneous accumulation of 32P activity in phosphatidic acid (PA) was also observed. Angiotensin II increased the breakdown of PI despite the presence of Li+. These results suggest that Li is a suitable tool to interrupt the accelerated PI cycle in angiotensin-stimulated cells. Aldosterone production of superfused cells was inhibited by Li+ when the cells were stimulated with angiotensin II. On the other hand, Li+ did not inhibit the aldosterone response of the cells to ACTH, a hormone which acts via cyclic AMP and does not enhance PI turnover in these cells. On the basis of these results, we assume that the inhibitory effect of Li+ on aldosterone production is related to its effect on PI turnover.     

Ca2+-dependent and Ca2+-independent mechanisms for phosphatidylinositol hydrolysis and 32P-labeling during cholinergic stimulation of the rat submaxillary gland in vitro

Ca²⁺ was required for carbachol-induced decreases in phosphatidylinositol (PI) and increases in phosphatidic acid (PA) concentrations during incubation of rat submaxillary gland fragments, but was not required for increases in [³²P]P_i incorporation into these phospholipids. Like carbachol, A23187 provoked a Ca²⁺-dependent decrease in PI mass. These results suggest concomitant operation of two separate mechanisms for stimulating PI hydrolysis and ³²P labeling of PA and PI during carbachol action: one mechanism is not dependent on external Ca²⁺ and is manifested by rapid labeling in a relatively small PA-PI pool; the other mechanism is dependent on Ca²⁺ and involves a large PA-PI pool which appears to have a relatively slow renewal (labeling) rate.     

Phosphatidic acid is a specific activator of phosphatidylinositol-4-phosphate kinase.

The lipid dependence of phosphatidylinositol-4-phosphate (PIP) kinase purified from bovine brain membranes was investigated. In the assay used, PIP-Triton X-100 micelles containing the lipid to be tested were presented to the enzyme. Under these conditions, phosphatidic acid (PA) stimulated the enzyme activity in a concentration-dependent manner up to 20-fold when an equal molar ratio of PA to PIP was attained. Stimulation by PA was highly specific; other lipids including lyso-PA and dicetylphosphate had a relatively small effect. The activation by PA was completely suppressed by phosphatidylinositol 4,5-bisphosphate (PIP2). To investigate the effect of PA on PIP kinase activity in natural membranes, endogenous PA was generated in rat brain synaptosomal plasma membranes by incubation with phospholipase D. Subsequent phosphorylation with [gamma-32P]ATP yielded an enhanced labeling of PIP2 but not of PIP in these membranes. These results suggest that PIP kinase activity may be under control of PA levels in membranes. This may have important implications for the regulation of cellular responses by agonist-induced phosphoinositide turnover.     

A new model system for studying the phosphatidylinositol cycle

An early manifestation of the response of WRK-1 rat mammary tumor cells to vasopressin is an increase in incorporation of (³²P)P_i into phospholipids. Incorporation into all classes of phospholipids is stimulated; however, incorporation into phosphatidylinositol (PI) is increased to the greatest degree (3- to 10-fold as compared with 1.3- to 2-fold for the other phosholipids). Furthermore, increased incorporation into PI is accompanied by an increased rate of PI turnover; turnover rates of the other phospholipids are unaffected by vasopressin.     

Muscarinic receptor-stimulated Ca2+ signaling and inositol lipid metabolism in avian salt gland cells

Activation of muscarinic cholinergic receptors was studied by measuring agonist-stimulated inositol lipid turnover and changes in [Ca²⁺]_i in dissociated salt gland secretory cells. Carbachol stimulation of quin2-loaded cells results in a sustained 4-fold increase in [Ca²⁺]_i, while incorporation of [³²P]P_i into phosphatidylinositol (PI) and phosphatidate are similarly increased. [³H]Inositol phosphates, measured in the presence of Li⁺, increased 13-fold. The stimulated increment in [Ca²⁺]_i required extracellular Ca²⁺, whereas [³H]inositol phosphate accumulation was independent of external Ca²⁺. Dose-response curves for carbachol-induced increments in [Ca²⁺]_i, PI labeling, and labeled inositol phosphate release are similar, with EC₅₀ values of 6, 4.5 and 8 μM, respectively. Dissociation constants for atropine vs. the quin2 and phospholipid responses are 0.59 ± 0.3 nM and 0.48 ± 0.28 nM, respectively. These cells thus provide a model system for the study of non-exocytotic secretion as a consequence of stimulated inositol lipid turnover.     

Calcium-independent phosphatidylinositol response in gonadotropin-releasing-hormone-stimulated pituitary cells.

This paper describes the effect of gonadotropin-releasing hormone (GnRH, gonadoliberin) on phospholipid metabolism in cultured rat pituitary cells. The cells were incubated with [32P]Pi to label endogenous phospholipids (10-60 min) and then stimulated with GnRH for up to 60 min. Cellular phospholipids were separated by two-dimensional t.l.c. and the radioactivity was determined. Phosphatidylinositol (PI), a minor constituent of cellular phospholipids (7.7%), was the major labelled phospholipid, accounting for 45% of the total radioactivity, at early periods after pulse labelling. On the other hand, phosphatidylcholine, the major cellular phospholipid (37%), was labelled only to 32% of the total radioactivity. The remaining label was distributed among phosphatidylethanolamine (4.2%), cardiolipin (3.4%), phosphatidic acid (PA, 2.5%), and phosphatidylserine (1.8%). GnRH doubled 32P labelling of PA and PI significantly at 1 and 5 min of incubation respectively in the presence or absence of extracellular Ca2+. Labelling of other phospholipids was not affected by GnRH treatment. The half-maximal stimulating dose (ED50) for PI labelling and lutropin release was 0.75 nM and 0.5 nM respectively, and the stimulatory effect was blocked by the potent GnRH antagonist [D-Glp1,pClPhe2,D-Trp3,6]GnRH. GnRH-stimulated PA and PI labelling could not be demonstrated after 1 and 45 min of incubation respectively, or when the prelabelling was conducted for 60 min rather than 10 min. These results suggest heterogeneous compartmentalization of gonadotroph PA and PI pools and that increased PI turnover might be a transducing signal for Ca2+ gating that follows gonadotroph GnRH-receptor activation.     

Phosphatidic Acid and Phosphoinositide Turnover in Myelin and Its Stimulation by Acetylcholine

Brain slices obtained from the forebrains of adult female rats were incubated with [32P]phosphate and [3H]glycerol for 60 min, and lipids extracted and analyzed by TLC. The 32P in brain slice lipids was primarily in polyphosphoinositides, phosphatidylinositol (PI), and phosphatidate (PA). Distribution of the 32P-labeled lipids in isolated myelin was biased toward PA, 38%, relative to 16% in whole tissue slice lipids. About 33% of the total labeled PA in brain slices was accounted for by that in myelin. On a per milligram protein basis, PA labeling in myelin is about 2.5-fold greater than that of whole brain slice. Since incorporation of [3H]glycerol (indicative of synthesis by the de novo synthetic pathway) was at very low levels, we conclude that [32P]phosphate entered into myelin PA primarily through a pathway involving phospholipase C activity. Much of the production of PA relates to hydrolysis of phosphoinositides, yielding diacylglycerol which is then phosphorylated within myelin. The distribution of label among the inositol-containing lipids suggests that only a fraction of the myelin polyphosphoinositides serve as substrate for rapid diglyceride production. In the presence of 10 mM acetylcholine (ACh) there was a 20-60% stimulation of [32P]phosphate incorporation into PA and PI of brain slice lipids and purified myelin. Stimulation by ACh was blocked by atropine. The observed increase in the 32P/3H ratio, relative to controls, indicated that for both total lipids and myelin lipids there was selective stimulation of a phospholipase C-dependent cycle relative to de novo biosynthesis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diacylglycerol rather than Ca2+ mediates GnRH inhibition of FSH induced steroidogenesis in ovarian granulosa cells.

Treatment of cultured granulosa cells with PLC or GnRH stimulated the rapid generation of DAG and phosphoinositide turnover. The PKC activators PLC (3 mU/ml) and TPA (10(-7)M) or the decapeptide GnRH (10(-6)M) elicited similar inhibitory responses on FSH or cAMP stimulated granulosa cell steroidogenesis. Mobilization of intracellular Ca2+ with A23187 (10(-8)M) was followed by a slight increase in the steroidogenic activity of cultured granulosa cells, whereas elevation of extracellular K+ (50 mM) largely augmented the steroid biosynthetic activity of the granulosa cells. These results suggest that the inhibitory effect of GnRH on granulosa cell steroidogenesis is mediated by generation of DAG, rather than by increases in intracellular Ca2+ concentrations.     

Compartmentation of phosphorylated precursors of phospholipid biosynthesis in cultured neuroblastoma cells

The continuous turnover of membrane phospholipids requires a steady supply of biosynthetic precursors. We evaluated the effects of decreasing extracellular Na+ concentration on phospholipid metabolism in cultured neuroblastoma (N1E 115) cells. Incubating cultures with 145 to 0 mM NaCl caused a concentration-dependent inhibition of [32P]phosphate uptake into the water-soluble intracellular pool and incorporation into phospholipid. Phospholipid classes were differentially affected; [32P]phosphate incorporated into phosphati-dylethanolamine (PE) and phosphatidylcholine (PC) was consistently less than into phosphatidylinositol (PI) and phosphatidylserine (PS). This could not be attributed to decreased phospholipid synthesis since under identical conditions, there was no effect on arachidonic acid or ethanolamine incorporation, and choline utilization for PC synthesis was increased. The effect of Na+ was highly specific since reducing phosphate uptake to a similar extent by incubating cultures in a phosphate-deficient medium containing Na+ did not alter the relative distribution of [32P]phosphate in phospholipid. Of several cations tested only Li+ could partially (50%) replace Na+. Incubation in the presence of ouabain or amiloride had no effect on [32P]phosphate incorporation into phospholipid. The differential effects of low Na+ on [32P]phosphate incorporation into PI relative to PC and PE suggests preferential compartmentation of [32P]phosphate into ATP in pools used for phosphatidic acid synthesis and relatively less in ATP pools used for synthesis of phosphocholine and phosphoethanolamine, precursors of PC and PE, respectively. This suggestion of heterogeneous and distinct pools of ATP for phospholipid biosynthesis, and of potential modulation by Na+ ion, has important implications for understanding intracellular regulation of metabolism.     

Control of phosphatidylinositol turnover in adrenal glomerulosa cells

The purpose of the present experiments was to compare the effects on phosphatidylinositol metabolism of agents stimulating aldosterone secretion. Glomerulosa cells, isolated from rat adrenals, were incubated in the presence of one of the following stimuli: angiotensin II, elevated potassium concentration, corticotropin, dibutyryl cyclic AMP and prostaglandin E2. Of all these substances, only angiotensin II stimulated the incorporation of [32P]phosphate into phosphatidylinositol. The effect was already detected 2.5 min and was still maintained 60 min after the onset of stimulation. A slight enhancement of the incorporation into other phospholipids was observed in the first minutes of stimulation. Cycloheximide abolished the effect of angiotensin II on aldosterone production, but not on phosphatidylinositol synthesis. In cells prelabelled with [32P]phosphate, radioactivity in phosphatidylinositol relative to that in other phospholipids decreased in response to angiotensin II within 5 min. This indicates that angiotensin II induces a specific breakdown of phosphatidylinositol. Corticotropin failed to enhance the incorporation of [32P]phosphate into phosphatidylinositol and other phospholipids in isolated fasciculate-reticularis cells. The results suggests that although both angiotensin II and potassium are presumed to act through changes in calcium metabolism, angiotensin alone generates the calcium signal by increased phosphatidylinositol turnover.