Effects of weak amplitude-modulated microwave fields on calcium efflux from awake cat cerebral cortex

Calcium (45Ca2+) efflux was studied from preloaded cortex in cats immobilized under local anesthesia, and exposed to a 3.0-mW/cm2 450-MHz field, sinusoidally amplitude modulated at 16 Hz modulation depth 85%). Tissue dosimetry showed a field of 33 V/m in the interhemispheric fissure (rate of energy deposition 0.29 W/kg). Field exposure lasted 60 min. By comparison with controls, efflux curves from field exposed brains were disrupted by waves of increased 45Ca2+ efflux. These waves were irregular in amplitude and duration, but many exhibited periods of 20-30 min. They continued into the postexposure period. Binomial probability analysis indicates that the field-exposed efflux curves constitute a different population from controls at a confidence level of 0.96. In about 70% of cases, initiation of field exposure was followed by increased end-tidal CO2 excretion for about 5 min. However, hypercapnea induced by hypoventilation did not elicit increased 45Ca2+ efflux. Thus this increase with exposure does not appear to arise as a secondary effect of raised cerebral CO2 levels. Radioactivity measurements in cortical samples after superfusion showed 45Ca2+ penetration at about 1.7 mm/hr, consistent with diffusion of the ion in free solution.     

Distinct effects of various amino acids on 45Ca2+ fluxes in rat pancreatic islets.

The effects of three types of amino acids on 45Ca2+ fluxes in rat pancreatic islets have been compared. Alanine, a non-insulinotropic neutral amino acid, transported with Na+, increased 45Ca2+ efflux in the presence or in the absence of extracellular Ca2+, but not in the absence of Na+. Its effects in Na+-solutions were practically abolished by 7 mM-glucose. Alanine slightly stimulated 45Ca2+ influx (5 min uptake) only when Na+ was present. Two insulinotropic cationic amino acids (arginine and lysine) triggered similar changes in 45Ca2+ efflux. They accelerated the efflux in the presence of Ca2+ and inhibited the efflux in a Ca2+-free medium, whether glucose was present or not. In an Na+-free Ca2+-medium, arginine and lysine markedly accelerated 45Ca2+ efflux, but this effect was suppressed by 7 mM-glucose. Arginine stimulated 45Ca2+ influx irrespective of the presence or absence of glucose and Na+. Leucine, a neutral insulinotropic amino acid well metabolized by islet cells, inhibited 45Ca2+ efflux from the islets in a Ca2+-free medium; this effect was potentiated by glutamine. In the presence of Ca2+ and Na+, leucine was ineffective alone, but triggered a marked increase in 45Ca2+ efflux when combined with glutamine. In an Na+-free Ca2+-medium, leucine accelerated 45Ca2+ efflux to the same extent with or without glutamine. Leucine also stimulated 45Ca2+ influx in the presence or in the absence of Na+, but its effects were potentiated by glutamine only in the presence of Na+. The results show that amino acids of various types cause distinct changes in 45Ca2+ fluxes in pancreatic islets. Certain of these changes involve an Na+-mediated mobilization of cellular Ca2+ from sequestering sites where glucose appears to exert an opposite effect.     

Sodium-dependent calcium efflux from adrenal chromaffin cells following exocytosis. Possible role of secretory vesicle membranes.

Although cytosolic Ca2+ transients are known to influence the magnitude and duration of hormone and neurotransmitter release, the processes regulating the decay of such transients after cell stimulation are not well understood. Na(+)-dependent Ca2+ efflux across the secretory vesicle membrane, following its incorporation into the plasma membrane, may play a significant role in Ca2+ efflux after stimulation of secretion. We have measured an enhanced 45Ca2+ efflux from cultured bovine adrenal chromaffin cells following cell stimulation with depolarizing medium (75 mM K+) or nicotine (10 microM). Such stimulation also causes Ca2+ uptake via voltage-gated Ca2+ channels and secretion of catecholamines. Na+ replacement with any of several substitutes (N-methyl-glucamine, Li+, choline, or sucrose) during cell stimulation inhibited the enhanced 45Ca2+ efflux, indicating and Na(+)-dependent Ca2+ efflux process. Na+ deprivation did not inhibit 45Ca2+ uptake or catecholamine secretion evoked by elevated K+. Suppression of exocytotic incorporation of secretory vesicle membranes into the plasma membrane with hypertonic medium (620 mOsm) or by lowering temperature to 12 degrees C inhibited K(+)-stimulated 45Ca2+ efflux in Na(+)-containing medium but did not inhibit the stimulated 45Ca2+ uptake. Enhancement of exocytotic secretion with pertussis toxin resulted in an enhanced 45Ca2+ efflux without affecting calcium uptake. The combined results suggest that Na(+)-dependent Ca2+ efflux across secretory vesicle membranes, following their incorporation into the plasma membrane during exocytosis, plays a significant role in regulating calcium efflux and the decay of cytosolic Ca2+ in adrenal chromaffin cells and possibly in related secretory cells.     

Effects of influenza A virus on human neutrophil calcium metabolism

Bacterial superinfection in influenza A virus-related illness may in part be explained by virus-induced neutrophil dysfunction. We here provide evidence that this effect is related to abnormal calcium metabolism of virus-infected cells. Neutrophils exposed to influenza virus for 0.5 h at 37 degrees C showed depressed O2- generation and release of radiolabeled arachidonic acid upon stimulation with FMLP. The peak cytosolic Ca2+ level achieved by virus-infected neutrophils after FMLP stimulation was significantly depressed as is efflux of 45Ca2+. This deficient Ca2+ mobilization could not be attributed to alterations of inositol phosphate production or Ca2+ influx in response to FMLP, both of which were unaffected by prior virus infection. Given these findings, the immediate effects of influenza virus on neutrophil Ca2+ metabolism were examined. The virus itself caused a rise in cytosolic Ca2+ and an efflux of 45Ca2+ without any corresponding 45Ca2+ influx. Total cell Ca2+ however was not depleted as measured by atomic absorption. Influenza virus, therefore, causes neutrophil activation leading to significant perturbations in Ca2+ metabolism and later to impaired mobilization of Ca2+ stores. This system offers a model for phagocyte deactivation and an opportunity to define control mechanisms of signal transduction.     

Studies on the role of inositol trisphosphate in the regulation of insulin secretion from isolated rat islets of Langerhans.

Glucose (20 mM) and carbachol (1 mM) produced a rapid increase in [3H]inositol trisphosphate (InsP3) formation in isolated rat islets of Langerhans prelabelled with myo-[3H]inositol. The magnitude of the increase in InsP3 formation was similar when either agent was used alone and was additive when they were used together. In islets prelabelled with 45Ca2+ and treated with carbachol (1 mM), the rise in InsP3 correlated with a rapid, transient, release of 45Ca2+ from the cells, consistent with mobilization of 45Ca2+ from an intracellular pool. Under these conditions, however, insulin secretion was not increased. In contrast, islets prelabelled with 45Ca2+ and exposed to 20mM-glucose exhibited a delayed and decreased 45Ca2+ efflux, but released 7-8-fold more insulin than did those exposed to carbachol. Depletion of extracellular Ca2+ failed to modify the increase in InsP3 elicited by either glucose or carbachol, whereas it selectively inhibited the efflux of 45Ca2+ induced by glucose in preloaded islets. Under these conditions, however, glucose was still able to induce a small stimulation of the first phase of insulin secretion. These results demonstrate that polyphosphoinositide metabolism, Ca2+ mobilization and insulin release can all be dissociated in islet cells, and suggest that glucose and carbachol regulate these parameters by different mechanisms.     

Metabolic, cationic and secretory response to D-glucose in depolarized and Ca(2+)-deprived rat islets exposed to diazoxide

D-glucose stimulates insulin release from islets exposed to both diazoxide, to activate ATP-responsive K+ channels, and a high concentration of K+, to cause depolarization of the B-cell plasma membrane. Under these conditions, the insulinotropic action of D-glucose is claimed to occur despite unaltered cytosolic Ca2+ concentration, but no information is so far available on the changes in Ca2+ fluxes possibly caused by the hexose. In the present experiments, we investigated the effect of D-glucose upon 45Ca efflux from islets exposed to both diazoxide and high K+ concentrations. In the presence of diazoxide and at normal extracellular Ca2+ concentration, D-glucose (16.7 mmol/l) inhibited insulin release at 5 mmol/l K+, but stimulated insulin release of 90 mmol/l K+. In both cases, the hexose inhibited 45Ca outflow. In the presence of diazoxide, but absence of Ca2+, D-glucose (8.3 to 25.0 mmol/l) first caused a rapid decrease in insulin output followed by a progressive increase in secretory rate. This phenomenon was observed both at 5 mmol/l or higher concentrations (30, 60 and 90 mmol/l) of extracellular K+. It coincided with a monophasic decrease in 45Ca efflux and either a transient (at 5 mmol/l K+) or sustained (at 90 mmol/l K+) decrease in overall cytosolic Ca2+ concentration. The decrease in 45Ca efflux could be due to inhibition of Na(+)-Ca2+ countertransport with resulting localized Ca2+ accumulation in the cell web of insulin-producing cells. A comparable process may be involved in the secretory response to D-glucose in islets exposed to diazoxide and a high concentration of K+ in the presence of extracellular Ca2+.     

Bromo-eudistomin D, a novel inducer of calcium release from fragmented sarcoplasmic reticulum that causes contractions of skinned muscle fibers

Bromo-eudistomin D induced a contraction of the chemically skinned fibers from skeletal muscle at concentrations of 10 microM or more. This contractile response to bromo-eudistomin D was completely blocked by 10 mM procaine. The extravascular Ca2+ concentrations of the heavy fractions of the fragmented sarcoplasmic reticulum (HSR) were measured directly by a Ca2+ electrode to examine the effect of bromo-eudistomin D on the sarcoplasmic reticulum. After the HSR was loaded with Ca2+ by the ATP-dependent Ca2+ pump, the addition of 10 microM bromo-eudistomin D caused Ca2+ release that was followed by spontaneous Ca2+ reuptake. In the presence of 2 microM ruthenium red or 4 mM MgCl2, no Ca2+ release was induced by 20 microM bromo-eudistomin D. The rate of 45Ca2+ efflux from HSR, which had been passively preloaded with 45Ca2+, was accelerated 7 times by 10 microM bromo-eudistomin D. The concentration of bromo-eudistomin D for half-maximum effect on the apparent efflux rate was 1.5 microM, while that of caffeine was 0.6 mM. The bromo-eudistomin D-evoked efflux of 45Ca2+ was abolished by 2 microM ruthenium red or 0.5 mM MgCl2. Bromo-eudistomin D was found to be 400 times more potent than caffeine in its Ca2+-releasing action but was similar in its action in other respects. These results indicate that bromo-eudistomin D may induce Ca2+ release from the sarcoplasmic reticulum through physiologically relevant Ca2+ channels.     

Stimulatory effect of enkephalins on calcium efflux from bovine adrenal chromaffin cells in culture

The effects of leucine- and methionine-enkephalin, opiate peptides, on Ca2+ efflux from cultured bovine adrenal chromaffin cells were examined. These enkephalins stimulated the efflux of 45Ca2+ from cells in a concentration-dependent manner (10(-8) M-10(-6) M). Leucine-enkephalin did not increase the intracellular free Ca2+ level, 45Ca2+ uptake, catecholamine secretion, cAMP level or cGMP level. The peptide-stimulated 45Ca2+ efflux was not inhibited by incubation in Ca2+-free medium, but was inhibited by incubation in Na+-free medium. These results indicate that enkephalins stimulate extracellular Na+-dependent 45Ca2+ efflux from cultured bovine adrenal chromaffin cells, probably by stimulating membrane Na+/Ca2+ exchange.     

Ca2+ dependence of transverse tubule-mediated calcium release in skinned skeletal muscle fibers

Isometric force and 45Ca efflux from the sarcoplasmic reticulum were measured at 19 degrees C in frog skeletal muscle fibers skinned by microdissection. After Ca2+ loading, application of the ionophores monensin, an Na+(K+)/H+ exchanger, or gramicidin D, an H+ greater than K+ greater than Na+ channel-former, evoked rapid force development and stimulated release of approximately 30% of the accumulated 45Ca within 1 min, whereas CCCP (carbonyl cyanide pyruvate p-trichloromethoxyphenylhydrazone), a protonophore, and valinomycin, a neutral, K+-specific ionophore, did not. When monensin was present in all bathing solutions, i.e., before and during Ca2+ loading, subsequent application failed to elicit force development and to stimulate 45Ca efflux. 5 min pretreatment of the skinned fibers with 50 microM digitoxin, a permeant glycoside that specifically inhibits the Na+,K+ pump, inhibited monensin and gramicidin D stimulation of 45Ca efflux; similar pretreatment with 100 microM ouabain, an impermeant glycoside, was ineffective. Monensin stimulation of 45Ca efflux was abolished by brief pretreatment with 5 mM EGTA, which chelates myofilament-space calcium. These results suggest that: monensin and gramicidin D stimulate Ca2+ release from the sarcoplasmic reticulum that is mediated by depolarization of the transverse tubules, which seal off after sarcolemma removal and form closed compartments; a transverse tubule membrane potential (myofilament space-negative) is maintained and/or established by the operation of the Na+,K+ pump in the transverse tubule membranes and is sensitive to the permeant inhibitor digitoxin; the transverse tubule-mediated stimulation of 45Ca efflux appears to be entirely Ca2+ dependent.     

Ca2+ dependence of stimulated 45Ca efflux in skinned muscle fibers

Stimulation of sarcoplasmic reticulum Ca release by Mg reduction of caffeine was studied in situ, to characterize further the Ca2+ dependence observed previously with stimulation by Cl ion. 45Ca efflux and isometric force were measured simultaneously at 19 degrees C in frog skeletal muscle fibers skinned by microdissection; EGTA was added to chelate myofilament space Ca either before or after the stimulus. Both Mg2+ reduction (20 or 110 microM to 4 microM) and caffeine (5 mM) induced large force responses and 45Ca release, which were inhibited by pretreatment with 5 mM EGTA. In the case of Mg reduction, residual efflux stimulation was undetectable, and 45Ca efflux in EGTA at 4 microM Mg2+ was not significantly increased. Residual caffeine stimulation at 20 microM Mg2+ was substantial and was reduced further in increased EGTA (10 mM); at 600 microM Mg2+, residual stimulation in 5 mM EGTA was undetectable. Caffeine appears to initiate a small Ca2+-insensitive efflux that produces a large Ca2+-dependent efflux. Additional experiments suggested that caffeine also inhibited influx. The results suggest that stimulated efflux is mediated mainly or entirely by a channel controlled by an intrinsic Ca2+ receptor, which responds to local [Ca2+] in or near the channel. Receptor affinity for Ca2+ probably is influenced by Mg2+, but inhibition is weak unless local [Ca2+] is very low.     

Acetylphosphate-induced Ca2+-Ca2+ exchange that is mediated by (Ca2+,Mg2+)-ATPase in sarcoplasmic reticulum vesicles

Sarcoplasmic reticulum vesicles were preloaded with either 45Ca2+ or unlabeled Ca2+. 45Ca2+ efflux and influx were determined in the presence and absence of acetylphosphate. Phosphorylation of the membrane-bound (Ca2+,Mg2+)-ATPase by [32P]acetylphosphate was also determined. The rate of efflux with acetylphosphate was considerably higher than that without acetylphosphate. When the acetylphosphate concentration was greatly reduced by diluting the reaction mixture after the start of the reaction, the rate of the efflux decreased markedly. These results demonstrate the acceleration of 45Ca2+ efflux by acetylphosphate. This acetylphosphate-induced efflux required external Ca2+. The external Ca2+ concentration giving half-maximum activation of efflux was 3.8 microM. The Ca2+ concentration dependence of the efflux coincided with that of phosphorylation. When the acetylphosphate concentration was varied, the rate of acetylphosphate-induced efflux changed approximately in proportion to the phosphoenzyme concentration. These and other findings show that acetylphosphate-induced 45Ca2+ efflux represents Ca2+-Ca2+ exchange (between the external medium and the internal medium) mediated by the phosphoenzyme and further demonstrate the direct dissociation of Ca2+ from the Ca2+-bound phosphoenzyme to the external medium in Ca2+-Ca2+ exchange.     

Potential-dependent passive transport of calcium in myocardium sarcolemma vesicles

The effect of membrane potential on passive Ca2+ transport in isolated cardiac sarcolemmal vesicles was investigated. The membrane potentials were induced by creating potassium gradients across the vesicular membranes in the presence of valinomycin. The fluorescence changes in the voltage-sensitive dye, dis-C3(5), were consistent with the induction of potassium equilibrium potentials. The rate of 45Ca2+ efflux from inside-out vesicles was considerably greater at 0 than at -80 or +55 mV; prepolarization of the membrane to +90 mV did not enhance the 45Ca2+ efflux upon subsequent depolarization. The voltage-dependent 45Ca2+ efflux increased with a rise in internal Ca2+ concentration and exhibited a saturation effect. Furthermore, evaluation of the rate of 45Ca2+ efflux over a wide range of membrane potentials produced a profile similar to that of current-voltage relationships for single calcium channels in isolated cardiomyocytes. It is concluded that the voltage-dependent Ca2+ efflux from the vesicles occurs via Ca2+-channels.     

Size of the inositol 1,4,5-trisphosphate-sensitive calcium pool in guinea-pig hepatocytes.

Permeabilized hepatocytes accumulated 45Ca2+ into a non-mitochondrial pool when provided with ATP. 45Ca2+ efflux from this pool was revealed by removal of ATP with glucose and hexokinase or by inhibiting uptake with NaVO3. The effect of inositol 1,4,5-trisphosphate (IP3) on 45Ca2+ efflux from the pool was investigated. IP3 (5 microM) evoked a rapid increase in the rate of 45Ca2+ efflux. Kinetic analysis of the effect of IP3 indicated the existence of two distinct Ca2+ fractions within the pool; only one, accounting for about one-third of the ATP-dependent Ca2+ content of the pool, was responsive to IP3. The effect of IP3 on 45Ca2+ efflux from the non-mitochondrial pool does not require ATP, a finding that is inconsistent with a previous suggestion that this effect may be mediated by protein phosphorylation.     

Alpha 1-adrenergic receptor activation mobilizes cellular Ca2+ in a muscle cell line

Regulation of cellular Ca2+ movements by alpha 1-adrenergic receptors has been studied using 45Ca2+ flux techniques in monolayer cultures of intact BC3H-1 cells. Unidirectional 45Ca2+ efflux from BC3H-1 cells reveals multiphasic kinetics, with a major fraction of cellular Ca2+ residing in a slowly exchanging intracellular compartment. Stimulation of alpha 1-adrenergic receptors by the agonist phenylephrine substantially increases 45Ca2+ unidirectional efflux, accompanied by a far smaller increase in 45Ca2+ influx. The selective enhancement of 45Ca2+ unidirectional efflux upon alpha 1-adrenergic receptor activation results in a net 30-40% decline in total cell Ca2+ content, measured either by radioisotopic equilibrium techniques or by atomic absorption spectroscopy. The relatively large pool of Ca2+ responsive to alpha-adrenergic stimulation is not displaced by La3+ but can be depleted with the Ca2+ ionophore A-23187. These results indicate that alpha 1-adrenergic receptor activation predominantly mobilizes Ca2+ from intracellular stores, together with a much smaller increase in transmembrane Ca2+ permeability. This interpretation is supported by comparative 45Ca2+ flux studies using a sister clone of BC3H-1 cells possessing surface nicotinic acetylcholine receptors but no alpha 1-adrenergic receptors. Agonist stimulation of the cholinergic receptor opens a well characterized transmembrane ion permeability gate. Cholinergic receptor activation greatly enhances the observed 45Ca2+ unidirectional influx relative to efflux, leading to net elevation of cellular Ca2+ content as Ca2+ moves down its inwardly directed concentration gradient.     

Coupling of ATP synthesis to reversal of rat liver microsomal Ca2+-ATPase

The reversal of the rat liver microsomal Ca2+-ATPase transport cycle was studied. Microsomes were loaded with 45Ca2+ (approximately 30 nmol/mg of protein) in an ATP-dependent process, and the time dependency of the microsomal 45Ca2+ efflux was determined with various ADP and inorganic phosphate (Pi) concentrations. Pseudo-first-order rate constants (K'e) for 45Ca2+ efflux were determined. Although there was considerable 45Ca2+ efflux in the absence of added ADP or Pi, the addition of ADP or Pi alone had minimal effects upon the K'e; in contrast, a 2.5-fold increase in the K'e was observed in the presence of both ADP and Pi. The apparent Km values for ADP and Pi were 4 microM and 0.22 mM, respectively. Stimulation of 45Ca2+ efflux by ADP and Pi was associated with ATP synthesis. The calcium ionophore A23187 prevented ATP synthesis, which indicates that the Ca2+ gradient facilitates the coupling of ATP synthesis to Ca2+ efflux.     

Dibutyryl cyclic AMP effects on calcium metabolism by mouse mastocytoma cells

The ability of mouse mastocytoma cells to take up 45Ca2+ was measured in normal growth medium. As previously observed in physiological buffers with succinate and Pi, cells grown for 18h with N6,O2'-dibutyryladenosine 3',5' cyclic monophosphate (DB cyclic AMP) to inhibit growth took up more 45Ca2+ than untreated cells. However 45Ca2+ uptake by cells in growth medium was less sensitive to respiratory inhibitors or uncouplers than 45Ca2+ uptake in physiological buffer. Increased 45Ca2+ uptake by 18h cyclic nucleotide-treated cells was not a result of tighter mitochondrial coupling since mitochondria prepared from cyclic nucleotide-treated cells were less coupled than those from untreated cells. Nevertheless studies with uncouplers suggested that the bulk of the intracellular Ca2+ was associated with mitochondria. DB cyclic AMP-treated cells contained less total Ca2+ than untreated cells indicating that net Ca2+ efflux occurred during the 18h period of drug treatment. These observations suggest that Ca2+ fluxes increase in DB cyclic AMP-treated PY815 cells and that a net efflux of Ca2+ occurs during growth inhibition by the cyclic nucleotide.     

Interaction of asparagine-linked oligosaccharides with an immobilized rice (Oryza sativa) lectin column.

Inside-out plasma-membrane vesicles isolated from rat liver [Prpic, Green, Blackmore & Exton (1984) J. Biol. Chem. 259, 1382-1385] accumulated a substantial amount of 45Ca2+ when they were incubated in a medium whose ionic composition and pH mimicked those of cytosol and which contained MgATP. The Vmax of the initial 45Ca2+ uptake rate was 2.9 +/- 0.6 nmol/min per mg and the Km for Ca2+ was 0.50 +/- 0.08 microM. The ATP-dependent 45Ca2+ uptake by inside-out plasma-membrane vesicles was about 20 times more sensitive to saponin than was the ATP-dependent uptake by a microsomal preparation. The 45Ca2+ efflux from the inside-out vesicles, which is equivalent to the Ca2+ influx in intact cells, was increased when the free Ca2+ concentration in the medium was decreased. The Ca2+ antagonists La3+ and Co2+ inhibited the 45Ca2+ efflux from the vesicles. Neomycin stimulated the Ca2+ efflux in the presence of either a high or a low free Ca2+ concentration. These results confirm that polyvalent cations regulate Ca2+ fluxes through the plasma membrane.     

Calcium mobilization by cadmium or decreasing extracellular Na+ or pH in coronary endothelial cells

Replacing extracellular Na+ with choline transiently increased cytoplasmic free Ca2+ ([Ca2+]i) more than 5-fold in coronary endothelial cells. Removing external Na+ stimulated 45Ca2+ efflux approximately 4-fold and influx approximately 1.7-fold. The stimulation of efflux was independent of extracellular Ca2+ and the osmotic Na+ substitute. The release of stored Ca2+, rather than Ca2+ influx via Na(+)-Ca2+ exchange, probably causes the increase in [Ca2+]i and 45Ca2+ efflux. Cadmium or decreasing external, not intracellular, pH transiently increased [Ca2+]i. Cd2+ and some other divalent metals also stimulated 45Ca2+ efflux. The potency order of the metals that stimulated efflux was Cd2+ greater than CO2+ greater than Ni2+ greater than Fe2+ greater than Mn2+. Incubating the cells with Zn2+ prior to assaying efflux in the absence of Zn2+ strongly inhibited the stimulation of 45Ca2+ efflux by Cd2+, pH 6, and the removal of external Na+ without affecting the stimulation of efflux by ATP. These findings support the hypothesis that certain trace metals or decreasing external Na+ or pH trigger the release of stored Ca2+ by stimulating a cell surface "receptor."     

Effect of amplitude-modulated 147 MHz radiofrequency radiation on calcium ion efflux from avian brain tissue

Cerebral cortex tissue slices and cerebral hemispheres prepared from Gallus domesticus chicks were exposed to 147 MHz radiofrequency radiation, amplitude modulated at 16 Hz and applied at a power density of 0.75 mW/cm2, to determine the effect of such exposure of 45Ca2+ efflux from the avian brain tissue. Statistical analysis of these data demonstrates that such exposure has no significant effect on 45Ca2+ efflux.     

Platelet-derived growth factor stimulates non-mitochondrial Ca2+ uptake and inhibits mitogen-induced Ca2+ signaling in Swiss 3T3 fibroblasts

Changes in intracellular free Ca2+ concentration [( Ca2+]i) were used to study the interaction between mitogens in Swiss 3T3 fibroblasts. Platelet-derived growth factor (PDGF) produced an increase in [Ca2+]i and markedly decreased the increases in [Ca2+]i caused by subsequent addition of bradykinin and vasopressin. If the order of the additions was reversed the [Ca2+]i response to PDGF was not inhibited by bradykinin or vasopressin. Inhibition of protein kinase C by staurosporine or chronic treatment of the cells with phorbol 12-myristate 13-acetate prevented the inhibitory effect of PDGF on the [Ca2+]i response to vasopressin but not bradykinin. PDGF did not decrease the receptor binding of bradykinin and produced only a small decrease in the receptor binding of vasopressin. PDGF decreased the rise in [Ca2+]i caused by the Ca2+ ionophores 4-bromo-A23187 and ionomycin and by a membrane perturbing ether lipid, 1-octadecyl-2-methyl-rac-glycero-3-phosphocholine, both in the presence and absence of external Ca2+. There was no change in cell 45Ca2+ influx caused by PDGF, vasopressin, or bradykinin. 45Ca2+ efflux from cells exposed to PDGF and vasopressin mirrored the changes in [Ca2+]i caused by the agents, that is, PDGF added after vasopressin produced a further increase in 45Ca2+ efflux but vasopressin did not increase 45Ca2+ efflux after PDGF. PDGF but not vasopressin caused an increase in the uptake of 45Ca2+ into an inositol 1,4,5-trisphosphate-insensitive non-mitochondrial store in permeabilized cells. The results suggest that the decreased [Ca2+]i response to mitogens after PDGF represents an action of PDGF at a point beyond the release of intracellular Ca2+ and the influx of external Ca2+, caused by an increase in the rate of removal of cytoplasmic free Ca2+. This increased removal of cytoplasmic Ca2+ by PDGF is not due to the increased export of Ca2+ from the cell but results from increased Ca2+ uptake into non-mitochondrial stores.