thapsigargin and tetrandrine

Tetrandrine inhibits tumor cell proliferation and demonstrates chemoprevention in cancer models. Speculation on the association between its effects on K+ and Ca2+ channels and cancer chemoprevention has been made. Thapsigargin also affects K+ and Ca2+ conductance. Thapsigargin, however, is a weak tumor promoter in the two-stage model of mouse skin carcinogenesis, yet it can induce apoptosis in androgen-independent prostatic cancer cells. I have postulated that arachidonic acid release from cells in culture is associated with cancer chemoprevention. The effects of tetrandrine and thapsigargin on arachidonic acid release from human colon carcinoma and rat liver cells and prostacyclin production by rat liver cells are compared in the current studies.. Tetrandrine and thapsigargin stimulate arachidonic acid release from human colon carcinoma and rat liver cells and prostacyclin production in rat liver cells. The stimulation by tetrandrine is not affected by incubation with actinomycin D, 100 mM KCl, the [Ca2+]i chelator, 1,2-bis (o-amino-5-fluorophenoxy) ethane-N,N,N',N',-tetraacetic acid tetraacetoxymethylester (BAPTA/AM) or in the absence of extracellular Ca2+. In contrast, stimulation by thapsigargin is inhibited by incubation with actinomycin D, 100 mM KCl, BAPTA/AM or in the absence of extracellular Ca2+.. Both tetrandrine and thapsigargin stimulate arachidonic acid release, but based on the different results obtained in the presence of actinomycin D, the [Ca2+]i chelator, 100 mM KCl and in the absence of extracellular Ca2+, the mechanisms leading to this release and pathways leading to apoptosis and/or cancer chemoprevention may be different. Stimulations by tetrandrine may be mediated by activation of a secretory phospholipase A2, whereas thapsigargin's stimulations may be mediated by the cytoplasmic Ca2+-dependent phospholipase A2.

Topics: Alkaloids; Animals; Arachidonic Acid; Benzylisoquinolines; Calcium Channel Blockers; Carcinogens; Cells, Cultured; HT29 Cells; Humans; Liver; Rats; Thapsigargin; Tumor Cells, Cultured

Tumour necrosis factor-alpha is believed to have a deleterious role in the pathophysiology of brain injury. Tetrandrine has protective effect on neuronal cells, however, the mechanisms involved in its action have not been clearly established. The aim of this study was to investigate the role of tetrandrine on calcium-dependent tumour necrosis factor-alpha production in glia-neurone mixed cultures. Glia-neurone mixed cultures were treated by addition of Ca2+ regulating agents for a period of 6 hr. Tetrandrine or/and TMB-8 were added 30 min. before the stimulation. The supernatant tumour necrosis factor-alpha levels were quantified by enzyme-linked immunosorbent assay. Exposure of lipopolysaccharide 10 and 100 ng/ml caused significant increase in tumour necrosis factor-alpha production respectively, with no alteration in cultures treated with 1 ng/ml lipopolysaccharide. Glia-neurone mixed cultures exhibited a marked elevation in tumour necrosis factor-alpha production after exposure to CaCl2, KCl, thapsigargin, BHQ and norepinephrine in the presence of lipopolysaccharide at 1 ng/ml respectively. Tetrandrine 0.3, 1, and 3 microM concentration-dependently reduced tumour necrosis factor-alpha production evoked by CaCl2 or KCl. Tetrandrine preincubation had no significant effect on the response to Ca2+-ATPase inhibitor thapsigargin or BHQ. Norepinephrine-induced tumour necrosis factor-alpha production was significantly reduced by tetrandrine and almost abolished by combination of tetrandrine and intracellular Ca2+ release inhibitor TMB-8. These results suggested that tetrandrine at a concentration of 0.3, 1, or 3 microM inhibited tumour necrosis factor-alpha production induced by Ca2+ entry in glia-neurone mixed cultures.

Topics: Alkaloids; Animals; Benzylisoquinolines; Calcium; Calcium Channel Blockers; Calcium Chloride; Cells, Cultured; Hydroquinones; Lipopolysaccharides; Neuroglia; Neurons; Norepinephrine; Potassium Chloride; Rats; Rats, Sprague-Dawley; Thapsigargin; Tumor Necrosis Factor-alpha

The aim of this study was to characterize the effects of hypotonicity on the activity of large-conductance Ca(2+)-activated K+ (BK(Ca)) channels in human retinal pigment epithelial (RPE R-50) cells. Effects of hypotonicity on ion currents were investigated with the aid of the patch-clamp technique. A regulatory volume decrease in response to a hypotonic solution (200 mOsm/L) was observed that could be blunted by paxilline. In whole-cell current recordings, a hypotonic solution (200 mOsm/L) reversibly increased the amplitude of K+ outward currents (I(K)). The increase of I(K) could be reversed by iberiotoxin (200 nM), paxilline (1 microM), or tetrandrine (5 microM), but not by glibenclamide (10 microM), disulphonic acid (DIDS) (100 microM), or dequalinium dichloride (10 microM). In RPE R-50 cells pretreated with thapsigargin, aristolochic acid, or pertussis toxin, the increased amplitude of I(K) in response to hypotonicity was unaltered. In cell-attached patches, an increase in BK(Ca)-channel activity was observed during hypotonicity-induced cell swelling. The enhanced channel activity elicited under this condition was mainly mediated by an increase in the number of long-lived openings. These findings support the evidence for the coupling of volume swelling to the functional activity of BK(Ca) channels.

Topics: Alkaloids; Aristolochic Acids; Benzylisoquinolines; Calcium; Calcium Channel Blockers; Cell Membrane Permeability; Cell Size; Cells, Cultured; Humans; Hypotonic Solutions; Indoles; Lactones; Membrane Potentials; Ophthalmic Solutions; Osmolar Concentration; Patch-Clamp Techniques; Peptides; Pertussis Toxin; Pigment Epithelium of Eye; Potassium; Potassium Channel Blockers; Potassium Channels, Calcium-Activated; Signal Transduction; Thapsigargin

To study the effects of tetrandrine, a Chinese herbal medicine, on the action potential (AP), contraction as well as sarcoplasmic reticulum (SR) calcium uptake of myocardium in guinea-pigs and dogs.. Changes in AP, dV/dt, peak tension (PT) and dT/dt of myocardial cells were studied using the technique of glass electrode. Changes of the calcium uptake rate by sarcoplasmic reticulum and release of inorganic phosphate from sarcoplasmic reticulum were assessed with biochemical techniques.. Tetrandrine exerts a concentration-dependent and frequency-dependent negative inotropic effect and shortens action potential duration. Tetrandrine depresses both dT(E)/dt and dT(L)/dt as well as the tension of myocardium, and reduces dV/dt and amplitude only in the slow action potential, thus implying that tetrandrine blocks the slow calcium channel. In addition, compared with thapsigargin, a specific inhibitor of Ca(2+)-ATPase on SR, tetrandrine more apparently suppresses the contraction of the myocardium.. Tetrandrine is a wide-range calcium antagonist of plant origin. Not only it blocks the voltage-operated calcium channels as other authors reported, but also may play an important role in affecting the function of Ca(2+)-ATPase and calcium release channels on SR. From this study, we also suggest that the calcium channel appears to be more critical than SR for the contraction of myocardium.

Topics: Action Potentials; Alkaloids; Animals; Benzylisoquinolines; Calcium; Coronary Vessels; Dogs; Dose-Response Relationship, Drug; Female; Guinea Pigs; Male; Myocardial Contraction; Myocardium; Phosphates; Sarcoplasmic Reticulum; Thapsigargin

Ca2+-activated K+ currents (I(K(Ca)) can contribute to action potential repolarization and after-hyperpolarization in GH3 cells. In this study, we examined how the activation of I(K(Ca) at the cellular level could be functionally coupled to Ca2+ influx through L-type Ca2+ channels. A 30-msec Ca2+ influx step to 0 mV was found to exhibit substantial contribution of Ca2+ influx through the activation of I(Ca,L) to the activation of I(K(Ca)). A bell-shaped relationship between the conditioning potentials and the integrated I(K(Ca)) was observed, suggesting that the magnitude of integrated I(Ca,L) correlates well with that of integrated I(K(Ca)) in the same cell. A linear relationship of integrated I(Ca,L) and integrated I(K(Ca)) was found with a coupling ratio of 69+/-7. The value of the coupling ratio was unaffected by the presence of Bay K 8644 or nimodipine, although these compounds could effectively affect the amplitudes of both I(K(Ca)) and I(Ca,L). However, tetrandrine could decrease the coupling ratio. Paxilline or intracellular Ca2+ buffer with EGTA decreased the coupling ratio, while apamin had no effect on it. Interestingly, phorbol 12-myristate 13-acetate also reduced the coupling ratio significantly, whereas thapsigargin increased this value. Thus, the present study indicates that the activation of I(K(Ca)) during brief Ca2+ influx, which is inhibited by paxilline, is coupled to Ca2+ influx primarily through the L-type channels. The selective modulation of I(K(Ca)) by second messengers or Ca2+ release from internal stores may affect the coupling efficiency and hence cellular excitability.

Topics: Alkaloids; Animals; Benzylisoquinolines; Calcium; Calcium Channel Blockers; Calcium Channels, L-Type; Cell Line; Egtazic Acid; Electrophysiology; Enzyme Inhibitors; Indoles; Pituitary Gland; Potassium Channel Blockers; Potassium Channels, Calcium-Activated; Prolactin; Tetradecanoylphorbol Acetate; Thapsigargin

The effects of tetrandrine, a blocker of voltage-dependent Ca(2+) channels, on ionic currents were investigated in an endothelial cell line (HUV-EC-C) originally derived from human umbilical vein. In whole-cell configuration, tetrandrine (0.5-50 microM) reversibly decreased the amplitude of K(+) outward currents. The IC(50) value of tetrandrine-induced decrease in outward current was 5 microM. The K(+) outward current in response to depolarizing voltage pulses was also inhibited by iberiotoxin (200 nM), yet not by glibenclamide (10 microM) or apamin (200 nM). The reduced amplitude of outward current by tetrandrine can be reversed by the further addition of Evans' blue (30 microM) or niflumic acid (30 microM). Thus, the tetrandrine-sensitive component of outward current is believed to be Ca(2+)-activated K(+) current. Pretreatment with thapsigargin (1 microM) or sodium nitroprusside (10 microM) for 5 h did not prevent tetrandrine-mediated inhibition of outward current. In outside-out configuration, bath application of tetrandrine (5 microM) did not change the single-channel conductance but significantly reduced the opening probability of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels. The tetrandrine-mediated decrease in the channel activity was independent on internal Ca(2+) concentration. Tetrandrine (5 microM) can also shift the activation curve of BK(Ca) channels to more positive potentials by approximately 20 mV. The change in the kinetic behavior of BK(Ca) channels caused by tetrandrine is due to a decrease in mean open time and an increase in mean closed time. The present study provides substantial evidence that tetrandrine is capable of suppressing the activity of BK(Ca) channels in endothelial cells. The direct inhibition of these channels by tetrandrine should contribute to its effect on the functional activities of endothelial cells.

Topics: Alkaloids; Apamin; Benzylisoquinolines; Calcium; Calcium Channel Blockers; Cell Line; Dose-Response Relationship, Drug; Drug Interactions; Endothelium, Vascular; Evans Blue; Glyburide; Humans; Membrane Potentials; Niflumic Acid; Nitroprusside; Peptides; Potassium Channels; Thapsigargin; Umbilical Veins

The effects of tyrosine kinase inhibitors and non-selective cation channel blockers on capacitative Ca(2+) entry were examined in the presence of methoxyverapamil in rat ileal smooth muscles. In Ca(2+)-free solution, carbachol or caffeine produced a rapid contraction mediated by Ca(2+) release from the stores (Ca(2+)-release response), and then led to Ca(2+) depletion of the stores. Subsequently, reintroduction of Ca(2+) caused a transient contraction due to capacitative Ca(2+) entry. Tyrosine kinase inhibitors, genistein and tyrphostin 47 but not herbimycin A, suppressed the responses to Ca(2+)-reintroduction much greater than Ca(2+)-release responses to carbachol or caffeine. Similar inhibitory effects on the responses to Ca(2+)-reintroduction were obtained with daidzein and tyrphostin A1, respective inactive analogue of genistein and tyrphostins. After continuous depletion of the stores with thapsigargin, Ca(2+)-reintroduction produced a sustained contraction, which was inhibited by these agents to different extents, but not by herbimycin A. In beta-escin-treated skinned muscles, genistein slightly reduced Ca(2+)-induced contraction. In fura-2-loaded tissues, SK&F 96365 inhibited contractile and [Ca(2+)](i) responses to Ca(2+)-reintroduction but minimally affected Ca(2+)-release responses. Tetrandrine suppressed both responses to Ca(2+)-reintroduction and to Ca(2+)-release. These results suggest that genistein and tyrphostin 47 inhibit capacitative Ca(2+) entry through an inhibition of Ca(2+) entry channels rather than tyrosine kinase. SK&F 96365, but not tetrandrine, seems to selectively inhibit the contractile responses to capacitative Ca(2+) entry in rat ileal smooth muscles.

Topics: Alkaloids; Animals; Benzylisoquinolines; Caffeine; Calcium; Calcium Channels; Enzyme Inhibitors; Genistein; Ileum; Imidazoles; Male; Muscle Contraction; Muscle, Smooth; Protein-Tyrosine Kinases; Rats; Rats, Wistar; Thapsigargin; Tyrphostins

The effects of tetrandrine, a Ca2+ antagonist of bis-benzylisoquinoline alkaloid origin, on endothelium-dependent and -independent vascular responsiveness were investigated in perfused rat mesenteric artery. In endothelium-intact preparations pre-contracted with 3 microM phenylephrine and fully relaxed by 0.3 microM acetylcholine tetrandrine caused a rapid transient contraction. In endothelium-denuded preparations, tetrandrine caused only vasorelaxation of phenylephrine-contraction. The biphasic effect of tetrandrine in acetylcholine-relaxed preparations could also be mimicked by sequential applications of atropine/tetrandrine or N(G)-nitro-L-arginine-methylester (L-NAME)/tetrandrine, but atropine or L-NAME alone caused only vasoconstriction. This tetrandrine-induced transient vasoconstriction was also observed in preparations relaxed with ATP, histamine or thapsigargin (TSG), but not those relaxed with A23187, sodium nitroprusside or nifedipine. The present results suggest that tetrandrine, in addition to its known inhibitory effects on vascular smooth muscle by virtue of its Ca2+ antagonistic actions, also inhibits NO production by the endothelial cells possibly by blockade of Ca2+ release-activated Ca2+ channels.

Topics: Acetylcholine; Adenosine Triphosphate; Alkaloids; Animals; Atropine; Benzylisoquinolines; Calcium Channel Blockers; Calcium Channels; Drugs, Chinese Herbal; Endothelium, Vascular; Histamine; In Vitro Techniques; Male; Mesenteric Arteries; NG-Nitroarginine Methyl Ester; Nitric Oxide; Perfusion; Phenylephrine; Rats; Rats, Sprague-Dawley; Thapsigargin; Vasoconstriction; Vasodilation

The effect of tetrandrine on the electrically induced elevation of cytosolic Ca2+ concentration, [Ca2+]i, in the single isolated rat cardiomyocyte was studied with a fluorometric ratio method using fura-2 acetomethylester (fura-2/AM) was Ca2+ indicator. Tetrandrine (3-100 microM) concentration and time dependently inhibited the amplitude of the [Ca2+]i transient without any significant effect on the resting level of [Ca2+]i. At high concentrations (60-100 microM), tetrandrine also prolonged the time to reach the peak (t1.0) and the time to decline the 20% of the peak level (t0.2) of the electrically induced [Ca2+]i transient. The effect of tetrandrine was fast in onset and fully reversible upon washout. Tetrandrine (10 microM) partially inhibited the elevation of [Ca2+]i in response to KCl-induced depolarization. Verapamil and diltiazem mimicked the effects of tetrandrine given at low concentrations, but not at high concentrations. At high concentrations, tetrandrine reduced the magnitude of the caffeine-induced [Ca2+]i transient. Tetrandrine (100 microM) administered after thapsigargin, which itself decreased the amplitude and prolonged the duration of the electrically induced [Ca2+]i transient, further decreased the amplitude of the [Ca2+]i elevation. After ryanodine, which itself decreased the amplitude of the [Ca2+]i transient, 100 microM tetrandrine not only further reduced the amplitude, but also prolonged the duration of the electrically induced [Ca2+]i transient. These results provide evidence that in addition to its inhibitory effect on Ca2+ influx at the sarcolemma at the therapeutically relevant concentrations, tetrandrine at high concentrations may inhibit Ca2+ uptake into the sarcoplasmic reticulum.

Topics: Alkaloids; Animals; Benzylisoquinolines; Caffeine; Calcium; Calcium Channel Blockers; Diltiazem; Electric Stimulation; Heart; In Vitro Techniques; Potassium Chloride; Rats; Rats, Sprague-Dawley; Ryanodine; Sodium; Thapsigargin; Verapamil

The effects of tetrandrine (TET), a Ca2+ antagonist of Chinese herbal origin, and hernandezine (HER), a structural analogue of TET, on Ca2+ mobilization were studied in rat glioma C6 cells. TET and HER alone did not affect the resting cytoplasmic Ca2+ concentration ([Ca2+]i). TET and HER inhibited the peak and sustained elevation of [Ca2+]i induced by bombesin and thapsigargin (TG), a microsomal Ca2+ ATPase inhibitor, in a dose-dependent manner. The doses of TET or HER needed to abolish the sustained and peak increase in [Ca2+]i induced by bombesin and TG were 30 microM and 300 microM, respectively. TET and HER did not increase inositol 1,4,5-trisphosphate (IP3) accumulation by themselves but inhibited IP3 accumulation elevated by bombesin. In permeabilized C6 cells, the addition of IP3 and TG released Ca2+ from intracellular stores. Pretreatment with TET or HER abolished Ca2+ release from intracellular stores induced by bombesin and TG. In the absence of extracellular Ca2+, the addition of 3 mM Ca2+ to extracellular medium slightly increased [Ca2+]i, which indicated Ca2+ entry due to leakage of Ca2+ at the plasma membrane but not Ca2+ influx through Ca2+ channels. TET and HER did not affect this leakage entry of Ca2+. The present results suggest that TET and HER inhibit Ca2+ release from intracellular stores as well as Ca2+ entry from extracellular medium evoked by bombesin and TG. In addition, TET and HER inhibit IP3 accumulation induced by bombesin in rat glioma C6 cells.

Topics: Alkaloids; Animals; Antineoplastic Agents, Phytogenic; Benzylisoquinolines; Bombesin; Calcium; Calcium Channel Blockers; Cell Division; Cell Membrane; Cell Membrane Permeability; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Enzyme Inhibitors; Glioma; Inositol 1,4,5-Trisphosphate; Rats; Thapsigargin; Tumor Cells, Cultured

1. The Ca(2+)-antagonism of tetrandrine (TET) on the Ca2+ mobilization in various types of cells were reviewed. Inositol trisphosphate (IP3)-generating drugs were used as Ca(2+)-mobilizing agonists and the effects were compared with those produced by using the microsomal Ca(2+)-ATPase inhibitor thapsigargin (TG), which is a tool for analysing Ca2+ store-regulated Ca2+ entry (capacitative Ca2+ entry). 2. In rat phaeochromocytoma PC12 cells, 100 mumol/L TET abolished high K+ (30 mmol/L)-induced sustained increases in cytoplasmic Ca2+ concentrations ([Ca2+]i) and partially inhibited bradykinin (1 mumol/L)- or TG (100 nmol/L)-induced Ca2+ entry. 3. In NIH/3T3 fibroblasts and rat parotid acinar cells, 100 mumol/L TET abolished Ca2+ entry induced by bombesin (1 mumol/L) and carbachol (100 mumol/L), respectively, or TG (100 nmol/L). However, in the human leukaemia T cell line Jurkat, 100 mumol/L TET did not inhibit Ca2+ entry evoked by either the anti-CD3 antibody OKT3 (10 mg/L) or TG (100 nmol/L). 4. In rat glioma C6 cells, the effects of TET on Ca2+ mobilization were further examined. At a high concentration, TET (300 mumol/L) alone did not affect [Ca2+]i in C6 cells. Tetrandrine inhibited the peak and sustained increases in [Ca2+]i induced by bombesin and TG in a dose-dependent manner. Although TET or TG did not produce increases in IP3, TET did inhibit increases in IP3 produced by bombesin. 5. Our results suggest that the action of TET on Ca2+ entry is dependent on cell types and that TET inhibits both Ca2+ entry from the extracellular medium and Ca2+ release from intracellular stores in rat glioma C6 cells.

Topics: Alkaloids; Animals; Benzylisoquinolines; Bradykinin; Calcium; Calcium Channel Blockers; Cell Line; Enzyme Inhibitors; PC12 Cells; Rats; Thapsigargin

Tetrandrine, an alkaloid extracted from the Chinese medicinal herb Radix stephania tetrandrae, has traditionally been used to treat hypertension. In the present study, the effect of tetrandrine on vascular smooth muscle was investigated by using the rat tail artery as a model of a resistance vessel. Tetrandrine relaxes the tension in tail artery helical strips produced by depolarization with 60 mM KCl. Further studies show that tetrandrine inhibits the KCl-induced intracellular Ca++ increase and L-type voltage-dependent Ca++ channel currents, suggesting that tetrandrine relaxes the vessel via inhibition of Ca++ influx through Ca++ channels. Tetrandrine also inhibits norepinephrine (NE)-induced vasocontraction in the presence of extracellular Ca++. It does not, however, inhibit NE-induced vasocontraction in the absence of extracellular Ca++. Tetrandrine also inhibits the NE-induced intracellular Ca++ increase in the presence of extracellular Ca++ and has no effect on the NE-induced intracellular Ca++ increase in the absence of extracellular Ca++. This suggests that tetrandrine also blocks NE-induced Ca++ influx but not NE-induced Ca++ release from the intracellular Ca++ stores. Furthermore, tetrandrine inhibits thapsigargin-induced intracellular Ca++ concentration increase, suggesting that, in addition to blocking Ca++ influx, tetrandrine also may interfere with the interaction between thapsigargin and Ca++ adenosine triphosphatase.

Topics: Alkaloids; Animals; Benzylisoquinolines; Calcium; Calcium Channel Blockers; In Vitro Techniques; Male; Muscle, Smooth, Vascular; Norepinephrine; Potassium Chloride; Rats; Rats, Sprague-Dawley; Terpenes; Thapsigargin; Vasoconstriction

We examined the effects of tetrandrine (TET) on Ca2+ mobilization in various types of cells using inositol trisphosphate-generating drugs and compared it with those using the microsomal Ca(2+)-ATPase inhibitor thapsigargin (TG) which is a tool for analyzing Ca2+ store-regulated Ca2+ entry (capacitative Ca2+ entry). In rat pheochromocytoma PC12 cells, 100 microM TET abolished high K+ (30 mM)-induced sustained increase in [Ca2+]i and partially inhibited bradykinin (1 microM)-induced or TG (100 nM)-induced Ca2+ entry. In NIH/3T3 fibroblasts, 100 microM TET abolished Ca2+ entry induced by bombesin (1 microM) or TG (100 nM). In rat glioma C6 cells, the addition of 100 microM TET reduced the sustained elevation of [Ca2+]i induced by endothelin 1 (10 nM) or TG (100 nM) declining to the resting level. In rat parotid acinar cells, 100 microM TET abolished a sustained increase in [Ca2+]i induced by carbachol (100 microM) or TG (100 nM). In human leukemia T-cell line Jurkat, 100 microM TET did not inhibit Ca2+ entry evoked by the anti-CD3 antibody OKT3 (10 micrograms/ml) or TG (100 nM). The present results suggest that the action of TET on Ca2+ entry is dependent on cell types.

Topics: Alkaloids; Animals; Benzylisoquinolines; Bradykinin; Calcium Channel Blockers; Calcium-Transporting ATPases; Cell Line; Enzyme Inhibitors; Fibroblasts; Humans; Inosine Triphosphate; PC12 Cells; Potassium; Rats; Terpenes; Thapsigargin

1. Tetrandrine (TET, a Ca2+ antagonist of Chinese herbal origin) and thapsigargin (TSG, an endoplasmic reticulum Ca2+ pump inhibitor) concentration-dependently mobilized Ca2+ from intracellular stores of HL-60 cells, with EC50 values of 20 microM and 0.8 nM, respectively. After intracellular Ca2+ release by 30 nM TSG, there was no more discharge of Ca2+ by TET (100 microM), and vice versa. 2. Pretreatments with 100 nM rauwolscine (alpha 2-adrenoceptor antagonist), 100 nM prazosin (alpha 1-adrenoceptor antagonist), 10 nM phorbol myristate acetate (PMA, a protein kinase C activator) or 100 nM staurosporine (a protein kinase C inhibitor) had no effect on 100 microM TET-induced intracellular Ca2+ release. 3. After intracellular Ca2+ release by 30 nM TSG in Ca(2+)-free medium, readmission of Ca2+ caused a substantial and sustained extracellular Ca2+ entry. The latter was almost completely inhibited by 100 microM TET (IC50 of 20 microM) added just before Ca2+ readmission. In Ca(2+)-containing medium, 30 nM TSG caused a sustained phase of cytosolic Ca2+ elevation, which could be abolished by 100 microM TET. TET was also demonstrated to retard basal entry of extracellular Mn2+ and completely inhibit TSG-stimulated extracellular Mn2+ entry. 4. TSG-induced extracellular Ca2+ entry was insensitive to the L-type Ca2+ channel blocker, nifedipine (1 microM), but was completely inhibited by the non-selective Ca2+ channel blocker La3+ (300 microM). Depolarization with 100 mM KCl did not raise the cytosolic Ca2+ level. 5. These data suggest that (a) TET and TSG mobilized the same Ca2+ pool and TET-induced intracellular Ca2+ release was independent of protein kinase C activity and ox-adrenoceptor activation,and (b) TET blocked the voltage-insensitive Ca2+ entry pathway activated by TSG. These dual effects on HL-60 cells were also observed with hernandezine (HER), a TET-like compound and in another cell type, murine B lymphoma M12.4 cells.

Topics: Alkaloids; Benzylisoquinolines; Calcium; Calcium Channel Blockers; Cytosol; Humans; Leukemia, Promyelocytic, Acute; Nifedipine; Protein Kinase C; Receptors, Adrenergic, alpha; Terpenes; Thapsigargin; Tumor Cells, Cultured