Phenylethylamine induces an increase in cytosolic Ca2+ in yeast

Beta-phenylethylamine (PEA) induced an increase in cytosolic free calcium ion concentration ([Ca2+]c) in Saccharomyces cerevisiae cells monitored with transgenic aequorin, a Ca2+-dependent photoprotein. The PEA-induced [Ca2+]c increase was dependent on the concentrations of PEA applied, and the Ca2+ mostly originated from an extracellular source. Preceding the Ca2+ influx, H2O2 was generated in the cells by the addition of PEA. Externally added H2O2 also induced a [Ca2+]c increase. These results suggest that PEA induces the [Ca2+]c increase via H2O2 generation. The PEA-induced [Ca2+]c increase occurred in the mid1 mutant with a slightly smaller peak than in the wild-type strain, indicating that Mid1, a stretch-activated nonselective cation channel, may not be mainly involved in the PEA-induced Ca2+ influx. When PEA was applied, the MATa mid1 mutant was rescued from alpha-factor-induced death in a Ca2+-limited medium, suggesting that the PEA-induced [Ca2+]c increase can reinforce calcium signaling in the mating pheromone response pathway.     

Cytochalasin induces an increase in cytosolic free calcium in murine B lymphocytes

Cytochalasin promotes the progression of anti-immunoglobulin-treated B lymphocytes to S phase. However, the intracellular events induced by cytochalasin which may mediate signaling for progression have not been elucidated. In this study, the effect of cytochalasin on the level of intracellular free calcium in murine splenic B lymphocytes was assessed by using the fluorescent calcium indicator Indo-1. Cytochalasins A, B, D, and E induced a rapid and sustained elevation of intracellular free calcium. The calcium response to cytochalasin derived largely from the influx of extracellular calcium, although a small, transient elevation in intracellular calcium persisted when the suspension medium was made calcium-free with EGTA, implicating an intracellular source for a portion of the calcium response. Single cell fluorescence studies revealed that cytochalasin elicited a calcium response in most splenic B cells in suspension, indicating that this phenomenon is not restricted to a subpopulation of responding B cells. Phorbol esters inhibited the B cell calcium response to cytochalasin, and an established response to cytochalasin was rapidly and completely reversed by subsequently administered phorbol ester. T cells that lack the cytochalasin pathway showed a markedly diminished calcium response that was only apparent at higher cytochalasin concentration. However, B cells from xid-defective [CBA/N X DBA/2]F1 males, which fail to respond to anti-immunoglobulin plus cytochalasin, showed a calcium response to cytochalasin similar to that of phenotypically normal F1 females. These data, along with the finding that the rise in intracellular calcium occurred in naive B cells as well as B cells previously treated with anti-immunoglobulin, suggest that there is no clear association between the calcium response induced by cytochalasin and the ability of cytochalasin to stimulate progression to S phase. However, this effect of cytochalasin may suggest a connection between actin filaments and calcium influx in B cells.     

Increased intracellular Ca2+ induces Ca2+ influx in human T lymphocytes.

One current hypothesis for the initiation of Ca2+ entry into nonelectrically excitable cells proposes that Ca2+ entry is linked to the state of filling of intracellular Ca2+ stores. In the human T lymphocyte cell line Jurkat, stimulation of the antigen receptor leads to release of Ca2+ from internal stores and influx of extracellular Ca2+. Similarly, treatment of Jurkat cells with the tumor promoter thapsigargin induced release of Ca2+ from internal stores and also resulted in influx of extracellular Ca2+. Initiation of Ca2+ entry by thapsigargin was blocked by chelation of Ca2+ released from the internal storage pool. The Ca2+ entry pathway also could be initiated by an increase in the intracellular concentration of Ca2+ after photolysis of the Ca(2+)-cage, nitr-5. Thus, three separate treatments that caused an increase in the intracellular concentration of Ca2+ initiated Ca2+ influx in Jurkat cells. In all cases, Ca(2+)-initiated Ca2+ influx was blocked by treatment with any of three phenothiazines or W-7, suggesting that it is mediated by calmodulin. These data suggest that release of Ca2+ from internal stores is not linked capacitatively to Ca2+ entry but that initiation is linked instead by Ca2+ itself, perhaps via calmodulin.     

Phospholipase A2 activation in chemotactic peptide-stimulated HL60 granulocytes: synergism between diacylglycerol and Ca2+ in a protein kinase C-independent mechanism

In dimethylsulfoxide-differentiated HL60 granulocytes, the chemotactic peptide N-formyl-Met-Leu-Phe (FMLP) augments arachidonic acid (AA) release via phospholipase A2 activity induced by the Ca2+-ionophore, A23187. Evidence indicates that this augmentation is mediated by diacylglycerols formed endogenously during FMLP receptor activation: The augmentation is mimicked by the synthetic diglyceride 1-oleoyl-2-acetyl-glycerol (OAG) and the tumor promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate; Pertussis toxin inhibits FMLP-induced augmentation but not OAG-induced augmentation: At suboptimal concentrations FMLP and OAG act cooperatively to augment ionophore A23187-induced AA release but not at optimal concentrations. These data indicate that phospholipase A2 activation in FMLP-stimulated HL60 granulocytes involves cooperative interactions between diacylglycerol formed endogenously and Ca2+. Interestingly, this effect of diacylglycerol appears not to be mediated by protein kinase C, since a specific protein kinase C inhibitor, 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H7) does not inhibit receptor-mediated release of AA by stimulated HL60 granulocytes.     

Polyunsaturated free fatty acids stimulate an increase in cytosolic Ca2+ by mobilizing the inositol 1,4,5-trisphosphate-sensitive Ca2+ pool in T cells through a mechanism independent of phosphoinositide turnover

Polyunsaturated free fatty acids (PUFAs) of both w-3 and w-6 series, induce a rapid increase of cytosolic free Ca2+ concentration ([Ca2+]i) in a leukemic T-cell line (JURKAT), measured by the fluorescent indicator fura-2. The early increase in [Ca2+]i was transient, falling to a sustained level which returned to base line after 10-15 min. In Ca2+-free medium, PUFAs still caused an early increase in [Ca2+]i but rapidly returned to basal. Depletion of endoplasmic reticular Ca2+ pool by addition of OKT3 (antibodies to CD3 of the T3-antigen receptor complex) to JURKAT cells (in Ca2+-free medium) abolished the PUFAs-mediated [Ca2+]i increase and vice versa. By using saponin-permeabilized JURKAT cells, the intracellular free Ca2+ released by PUFAs was found to be the non-mitochondrial, ATP-dependent sequestered Ca2+ pool which is sensitive to inositol 1,4,5-trisphosphate. However, PUFAs do not induce any apparent increase in inositol phosphates in JURKAT cells. No Ca2+ influx was detected in JURKAT cells when stimulated with PUFAs. A correlation was observed between both the carbon chain length and the number of double bonds with the ability to mobilize cytosolic free [Ca2+]i in the w-3 PUFAs. These results demonstrate that PUFAs stimulate the release of Ca2+ from the inositol 1,4,5-trisphosphate-sensitive Ca2+ pool in the endoplasmic reticulum of JURKAT cells via a mechanism independent of inositol lipid hydrolysis.     

Antimicrobial and immunostimulatory peptide,KLK, induces an increase in cytosolic Ca2+ concentration by mobilizing Ca2+ from intracellular stores

The cationic antimicrobial immunomodulatory peptide, KLK (KLKL₅KLK), exerts profound membrane interacting properties, impacting on ultrastructure and fluidity. KLK–membrane interactions that lead to these alterations require the ability of the peptide to move into an α‐helical conformation. We show that KLK induces an increase of the intracellular Ca²⁺ concentration in human T24 cells. The effect of KLK is buffer‐sensitive, as it is detected when HBSS buffer is used, but not with PBS. This, together with the lack of effect of the middle leucine‐to‐proline‐substituted peptide derivative [KPK (KLKLLPLLKLK)], indicates that it is the conformational propensity rather than the net positive charge that contributes to the effect of KLK on intracellular Ca²⁺ level of T24 cells. We show that, although KLK slightly stimulates Ca²⁺ influx into the cell, the bulk increase of Ca²⁺ levels is due to KLK‐induced depletion of intracellular Ca²⁺ stores. Finally, we demonstrate a KLK‐induced switch of PS (phosphatidylserine) from the inner to the outer plasma membrane leaflet that contributes to the onset of early apoptotic changes in these cells.     

Effect of perturbing diacylglycerol metabolism on cytosolic free Ca2+ oscillations induced in single hepatocytes.

Single rat hepatocytes microinjected with aequorin show free Ca oscillations when stimulated with Ca(2+)-mobilizing hormones. We show here that an inhibitor of diacylglycerol kinase (R 59 022) and an analogue of native diacylglycerol (diC8) inhibit free Ca oscillations induced by phenylephrine and vasopressin. These results agree with a negative feedback effect of protein kinase C on free Ca oscillations.     

NAD+ levels control Ca2+ store replenishment and mitogen-induced increase of cytosolic Ca2+ by Cyclic ADP-ribose-dependent TRPM2 channel gating in human T lymphocytes

Intracellular NAD(+) levels ([NAD(+)](i)) are important in regulating human T lymphocyte survival, cytokine secretion, and the capacity to respond to antigenic stimuli. NAD(+)-derived Ca(2+)-mobilizing second messengers, produced by CD38, play a pivotal role in T cell activation. Here we demonstrate that [NAD(+)](i) modifications in T lymphocytes affect intracellular Ca(2+) homeostasis both in terms of mitogen-induced [Ca(2+)](i) increase and of endoplasmic reticulum Ca(2+) store replenishment. Lowering [NAD(+)](i) by FK866-mediated nicotinamide phosphoribosyltransferase inhibition decreased the mitogen-induced [Ca(2+)](i) rise in Jurkat cells and in activated T lymphocytes. Accordingly, the Ca(2+) content of thapsigargin-sensitive Ca(2+) stores was greatly reduced in these cells in the presence of FK866. When NAD(+) levels were increased by supplementing peripheral blood lymphocytes with the NAD(+) precursors nicotinamide, nicotinic acid, or nicotinamide mononucleotide, the Ca(2+) content of thapsigargin-sensitive Ca(2+) stores as well as cell responsiveness to mitogens in terms of [Ca(2+)](i) elevation were up-regulated. The use of specific siRNA showed that the changes of Ca(2+) homeostasis induced by NAD(+) precursors are mediated by CD38 and the consequent ADPR-mediated TRPM2 gating. Finally, the presence of NAD(+) precursors up-regulated important T cell functions, such as proliferation and IL-2 release in response to mitogens.     

Receptor-mediated activation of electropermeabilized neutrophils. Evidence for a Ca2+- and protein kinase C-independent signaling pathway

Electrically permeabilized neutrophils were used to study the mechanism of activation of the respiratory burst by the chemotactic agent formyl-methionyl-leucyl-phenylalanine (fMLP). Permeabilization was assessed by flow cytometry, radioisotope trapping, and by the requirement for exogenous NADPH for oxygen consumption. A respiratory burst could be elicited by fMLP, phorbol ester, or diacylglycerol in permeabilized cells suspended in EGTA-buffered medium with 100 nM free Ca2+. The fMLP response persisted even in cells depleted of intracellular Ca2+ stores by pretreatment with ionomycin. Therefore, a change in cytosolic free Ca2+ ([Ca2+]i) is not required for receptor-mediated stimulation of the respiratory burst. The responses induced by phorbol ester and diacylglycerol were largely inhibited by H7, a protein kinase C antagonist. In contrast, the stimulation of oxygen consumption by fMLP was unaffected by H7. These results suggest that a third signaling pathway, distinct from changes in [Ca2+]i and activation of protein kinase C, is involved in the response of neutrophils to chemoattractants.     

Stimulus-response coupling in insulin-secreting HIT cells. Effects of secretagogues on cytosolic Ca2+, diacylglycerol, and protein kinase C activity

The hamster islet B cell line HIT retains the ability to secret insulin in response to glucose and several receptor agonists. We used HIT cells to study the initial signaling events in glucose or receptor agonist-stimulated insulin secretion. Glucose stimulated insulin release from HIT cells in a dose-dependent manner with a half-maximal effect seen already at 1 mM. Insulin release was also stimulated by carbachol in a glucose-dependent manner. Glucose depolarized the HIT cell membrane potential as assessed with the fluorescent probe bisoxonol and raised intracellular Ca2+ as revealed by fura-2 measurements. Using a Mn2+ fura-2 quenching technique, we could show that the rise in intracellular Ca2+ was due to Ca2+ influx following opening of voltage-gated Ca2+ channels. Glucose is thought to increase the diacylglycerol (DAG) content of insulin-secreting cells. However, although HIT cells respond to glucose in terms of insulin secretion, membrane depolarization, and Ca2+ rise, the hexose was unable to increase the proportion of protein kinase C activity associated with membranes. In contrast, the membrane-associated protein kinase C activity increased in HIT cells exposed to the two receptor agonists carbachol and bombesin. Bombesin was shown to generate DAG with the expected fatty acid composition of activators of phospholipase C. Glucose, in contrast, only caused minor increases in DAG containing myristic and palmitic acid without affecting total DAG mass. The failure to detect stimulation of protein kinase C by glucose could be due to both the limited amount and to the different fatty acid composition of the metabolically generated DAG. The latter was in part supported by experiments performed on protein kinase C partially purified from HIT cells. Indeed, 1,2-dipalmitoylglycerol, presumed to be the main DAG species generated by glucose, was only one-third as active as 1,2-dioleoylglycerol and 1-stearoyl-2-arachidonylglycerol in stimulating the isolated enzyme at physiological Ca2+ concentration. It is therefore unlikely that DAG and protein kinase C play a major role in glucose-stimulated insulin secretion.     

Intestinal secretagogues increase cytosolic free Ca2+ concentration and K+ conductance in a human intestinal epithelial cell line

Summary A human intestinal epithelial cell line (Intestine 407) is known to retain receptors for intestinal secretagogues such as acetylcholine (ACh), histamine, serotonin (5-HT) and vasoactive intestinal peptide (VIP). The cells were also found to possess separate receptors for secretin and ATP, the stimulation of which elicited transient hyperpolarizations coupled to decreased membrane resistances. These responses were reversed in polarity at the K⁺ equilibrium potential. The hyperpolarizing responses to six agonists were reversibly inhibited by quinine or quinidine. By means of Ca²⁺-selective microelectrodes, increases in the cytosolic free Ca²⁺ concentration were observed in response to individual secretagogues. The time course of Ca²⁺ responses coincided with that of hyperpolarizing responses. The responses to ACh and 5-HT were abolished by a reduction in the extracellular Ca²⁺ concentration down to pCa 7 or by application of Co²⁺. Thus, in Intestine 407 cells, not only the intestinal secretagogues, which are believed to act via increased cytosolic Ca²⁺ (ACh, 5-HT and histamine), but also those which elevate cyclic AMP (VIP, secretin and ATP) induce increases in cytosolic Ca²⁺, thereby activating the K⁺ conductance. It is likely that the origin of increased cytosolic Ca²⁺ is mainly extracellular for ACh- and 5-HT-induced responses, whereas histamine, VIP, secretin and ATP mobilize Ca²⁺ from the internal compartment.     

Mechanisms of nitric-oxide-induced increase of free cytosolic Ca2+ concentration in Nicotiana plumbaginifolia cells

In this study, we investigated a role for nitric oxide (NO) in mediating the elevation of the free cytosolic Ca(2+) concentration ([Ca(2+)](cyt)) in plants using Nicotiana plumbaginifolia cells expressing the Ca(2+) reporter apoaequorin. Hyperosmotic stress induced a fast increase of [Ca(2+)](cyt) which was strongly reduced by pretreating cell suspensions with the NO scavenger carboxy PTIO, indicating that NO mediates [Ca(2+)](cyt) changes in plant cells challenged by abiotic stress. Accordingly, treatment of transgenic N. plumbaginifolia cells with the NO donor diethylamine NONOate was followed by a transient increase of [Ca(2+)](cyt) sensitive to plasma membrane Ca(2+) channel inhibitors and antagonist of cyclic ADP ribose. We provided evidence that NO might activate plasma membrane Ca(2+) channels by inducing a rapid and transient plasma membrane depolarization. Furthermore, NO-induced elevation of [Ca(2+)](cyt) was suppressed by the kinase inhibitor staurosporine, suggesting that NO enhances [Ca(2+)](cyt) by promoting phosphorylation-dependent events. This result was further supported by the demonstration that the NO donor induced the activation of a 42-kDa protein kinase which belongs to SnRK2 families and corresponds to Nicotiana tabacum osmotic-stress-activated protein kinase (NtOSAK). Interestingly, NtOSAK was activated in response to hyperosmotic stress through a NO-dependent process, supporting the hypothesis that NO also promotes protein kinase activation during physiological processes.     

Inositol phosphate formation in fMet-Leu-Phe-stimulated human neutrophils does not require an increase in the cytosolic free Ca2+ concentration.

The accumulation of inositol phosphates in myo-[3H]inositol-labelled human neutrophils stimulated with the chemotactic peptide fMet-Leu-Phe was measured. The challenge with the chemotactic peptide caused the generation of inositol monophosphate (InsP), inositol bisphosphate (InsP2) and inositol trisphosphate (InsP3). The formation of the three inositol phosphates followed a differential time course: InsP3 accumulated very rapidly and transiently, whereas InsP increased steadily for more than 2 min. Inositol phosphate formation was only partially decreased by procedures which prevented the fMet-Leu-Phe-dependent increase of cytosolic free Ca2+ concentration.     

Stimulation of a T helper cell class 2 clone with immobilized anti-T cell receptor antibody activates a Ca2+ and protein kinase C-independent lethal signaling pathway.

Stimulation of an IL-2-dependent variant of the Th2 clone D10.G4.1 with antibodies (Ab) specific for CD3 epsilon or the TCR-alpha beta caused either activation of the clone to secrete the autocrine lymphokine IL-4, or lethal activation in which the cells secreted high quantities of IL-4 but then died within 2 days. High densities of immobilized Ab delivered a lethal signal, whereas soluble forms of Ab and low densities of immobilized Ab caused productive activation in which cell viability was maintained. Lethal activation was not prevented by accessory cells, IL-1, or IL-2, or by co-cross-linkage of CD4 and TCR. The lethal signal was not mediated via a soluble effector from the activated cells. Lethal signaling was insensitive to cyclosporin A or dexamethasone. Studies with activators of protein kinase C (PKC), and PKC inhibitors, indicated that direct activation of PKC was not sufficient for lethal signaling. Nor could direct activation of PKC prevent the lethal signal. The lethal signal was not caused by Ca2+ mobilization mediated by Ca2+ ionophore and there was no evidence of apoptosis. The combination of a PKC activator and Ca2+ ionophore was not lethal, thereby showing that together these events are not sufficient. That these signal pathways were not necessary for lethal activation was evidenced by their inability to lower the density of immobilized anti-CD3 required to cause cell death. In this model, ligation of the TCR specifically activates a Ca2+/PKC-independent lethal signal transduction pathway.     

Secretin induces rapid increases in inositol trisphosphate, cytosolic Ca2+ and diacylglycerol as well as cyclic AMP in rat pancreatic acini.

Previous studies have shown that the dose-response relationship for secretin-stimulated cyclic AMP accumulation is different from that for secretin-stimulated enzyme secretion in the rat exocrine pancreas. Here we show that secretin concentrations of 10(-10) M and higher stimulated a rise in cyclic AMP levels, with maximum effect on cyclic AMP accumulation being achieved already with 10(-8) M-secretin. However, at this concentration of secretin, enzyme secretion rates were approximately half-maximal. Unexpectedly, at concentrations of secretin greater than 10(-8) M there was evidence suggestive of phosphatidylinositol bisphosphate hydrolysis with rapid increases in inositol trisphosphate, cytosolic free calcium and diacylglycerol content of rat pancreatic acini. Furthermore, there was a dose-response relationship among secretin concentration (in the range 10(-8) M-2 X 10(-6) M), increases in inositol trisphosphate and increases in cytosolic free calcium ([Ca2+]i). Contrary to what has been previously believed, these results clearly indicate that in rat pancreatic acini secretin not only stimulates cyclic AMP accumulation but also raises inositol trisphosphate, [Ca2+]i and diacylglycerol. Thus, two second messenger systems may play a role in the regulation of secretin-induced amylase release.     

Tributyltin increases cytosolic free Ca2+ concentration in thymocytes by mobilizing intracellular Ca2+, activating a Ca2+ entry pathway, and inhibiting Ca2+ efflux.

The immunotoxic environmental pollutant tri-n-butyltin (TBT) kills thymocytes by apoptosis through a mechanism that requires an increase in intracellular Ca2+ concentration. The addition of TBT (EC50 = 2 microM) to fura-2-loaded rat thymocytes resulted in a rapid and sustained increase in the cytosolic free Ca2+ concentration ([Ca2+]i) to greater than 1 microM. In nominally Ca(2+)-free medium, TBT slightly but consistently increased thymocyte [Ca2+]i by about 0.11 microM. The subsequent restoration of CaCl2 to the medium resulted in a sustained overshoot in [Ca2+]i; similarly, the addition of MnCl2 produced a rapid decrease in the intracellular fura-2 fluorescence in thymocytes exposed to TBT. The rates of Ca2+ and Mn2+ entry stimulated by TBT were essentially identical to the rates stimulated by 2,5-di-(tert.-butyl)-1,4-benzohydroquinone (tBuBHQ), which has previously been shown to empty the agonist-sensitive endoplasmic reticular Ca2+ store and to stimulate subsequent Ca2+ influx by a capacitative mechanism. The addition of excess [ethylenebis(oxyethylenenitrilo)]tetraacetic acid to thymocytes produced a rapid return to basal [Ca2+]i after tBuBHQ treatment but a similar rapid return to basal [Ca2+]i was not observed after TBT treatment. In addition, TBT produced a marked inhibition of both Ca2+ efflux from the cells and the plasma membrane Ca(2+)-ATPase activity. Also, TBT treatment resulted in a rapid decrease in thymocyte ATP level. Taken together, our results show that TBT increases [Ca2+]i in thymocytes by the combination of intracellular Ca2+ mobilization, stimulation of Ca2+ entry, and inhibition of the Ca2+ efflux process. Furthermore, the ability of TBT to apparently mobilize the tBuBHQ-sensitive intracellular Ca2+ store followed by Ca2+ and Mn2+ entry suggests that the TBT-induced [Ca2+]i increase involves a capacitative type of Ca2+ entry.     

The Ca2+ antagonists binding cytosolic protein has properties of the Ca2+ channel.

In our previous work (Krizanová et al. 1989) we have described a protein from rabbit skeletal muscle cytosolic fraction, which is able to bind dihydropyridines and phenothiazines. In the present work conclusive evidence is provided for the ability of the phospholipid-reconstituted cytosolic protein to transport calcium. The calcium transport was stimulated by BAY K 8644 and inhibited in the presence of PN 200-110. Our observations were confirmed also by electrophysiological measurements on planar lipid bilayers. The possibility that the cytosolic fraction was contaminated with membranes could be definitely ruled out. Nevertheless, the nature of the protein under study is still in the frame of guess.     

Activation of protein kinase C in human neutrophils attenuates agonist-stimulated rises in cytosolic free Ca2+ concentration by inhibiting bivalent-cation influx and intracellular Ca2+ release in addition to stimulating Ca2+ efflux.

Stimulation of fura-2-loaded human neutrophils with formylmethionyl-leucyl-phenylalanine (FMLP) or ionomycin elevated the cytosolic free Ca2+ concentration, [Ca2+], to a maintained elevated level. Activation of protein kinase C (C-kinase) with phorbol 12-myristate 13-acetate, 4 beta-phorbol 12,13-didecanoate or dioctanoylglycerol caused decreases in [Ca2+]i from this level. 4 alpha-Phorbol didecanoate, which does not activate C-kinase, had no effect. These results confirm previous reports that C-kinase activation decreases neutrophil [Ca2+]i by stimulating removal of Ca2+ from the cytosol. Further experiments showed that activation of C-kinase attenuated the component of the FMLP-stimulated [Ca2+]i rise that was dependent on external Ca2+. C-kinase activation also inhibited FMLP-stimulated entry of the quenching cation, Mn2+, used as an indicator of bivalent-cation entry. In contrast, C-kinase activation caused only a partial inhibition of FMLP-stimulated release of Ca2+ from intracellular stores. 4 alpha-Phorbol didecanoate was ineffective in inhibiting Ca2+ entry, Mn2+ entry and intracellular Ca2+ release. Addition of FMLP also stimulated a decrease in the ionomycin-elevated [Ca2+]i, and this effect was blocked by staurosporine, a protein kinase inhibitor. These results show that, in addition to stimulating Ca2+ efflux, C-kinase activation in neutrophils inhibits FMLP-stimulated entry of bivalent cations, and partially inhibits intracellular release of Ca2+. Further, FMLP itself can modulate [Ca2+]i by activation of C-kinase.