Translocation of protein kinase C is associated with inhibition of 5-HT uptake by cultured endothelial cells

1989 ◽  
Vol 257 (4) ◽  
pp. L253-L258 ◽  
Author(s):  
C. L. Myers ◽  
J. S. Lazo ◽  
B. R. Pitt
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Dependent Manner ◽  
Trypan Blue Exclusion ◽  
Concentration Dependent Manner ◽  
Membrane Function ◽  
Membrane Activity ◽  
Kinase C ◽  
5 Ht Uptake

Acute exposure (30 min-1 h) of bovine pulmonary artery endothelial cells in culture (BPAEC) to phorbol myristate acetate (PMA 10(-10)-10(-6) M) resulted in concentration-dependent decrement in serotonin (5-HT) uptake. Neither cell viability (trypan blue exclusion or release of deoxyglucose) nor activity of another plasma membrane function, angiotensin-converting enzyme activity, were affected. A decrease in 5-HT uptake was also noted with phorbol 12,13 dibutyrate and mezerein, but not with 4-alpha-phorbol 12,13 didecanoate, which does not stimulate protein kinase C (PKC). Inhibition of 5-HT uptake by PMA (160 nM) was reversed in a concentration-dependent manner by pretreatment of cells with the PKC inhibitor staurosporine (3-100 nM). BPAEC were treated with PMA (160 nM) for 1 h, and activities of PKC in cytosolic and membrane compartments were determined. PMA did not significantly affect total cellular PKC activity but resulted in a translocation of activity from cytosol to membrane (control membrane activity 67 +/- 4%; PMA-treated membrane activity 97 +/- 1% of total cellular PKC). Thus we propose that translocation of PKC from cytosol to membrane results in inhibition of 5-HT uptake by BPAEC.

Endothelin activation of phospholipase D: dual modulation by protein kinase C and Ca2+

1993 ◽  
Vol 264 (5) ◽  
pp. F845-F853
Author(s):  
M. M. Friedlaender ◽  
D. Jain ◽  
Z. Ahmed ◽  
D. Hart ◽  
R. L. Barnett ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase D ◽  
Intracellular Signaling ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Eta Receptor ◽  
Ca2 Channels ◽  
Stimulation Of

Previous work from this laboratory has identified an endothelin (ET) type A (ETA) receptor on cultured rat renal medullary interstitial cells (RMIC), coupled to phosphatidylinositol-specific phospholipase C (PI-PLC), dihydropyridine-insensitive receptor-operated Ca2+ channels, and phospholipase A2. The current studies explored a role for ET stimulation of phosphatidylcholine-specific phospholipase D (PC-PLD) in intracellular signaling of this cell type. ET stimulated PLD activation, as measured by phosphatidic acid (PA) or phosphatidylethanol (PEt) accumulation, in a time- and concentration-dependent manner. Inhibition of diacylglycerol (DAG) kinase by ethylene glycol dioctanoate or 6-(2)4-[(4-fluorophenyl)-phenylmethylene]-1-piperadinyl]ethy l-7-methyl-5H - thiaxolo-[3,2-alpyrimidin]-5-one (R 59022) failed to blunt PA accumulation, indicating that PLD, and not DAG, was the source of PA. Inhibition of PA phosphohydrolase (PAP) by propranolol increased late accumulation of PA, suggesting that the prevailing metabolic flow was in the direction of PA to DAG. Phorbol 12-myristate 13-acetate (PMA) augmented ET-evoked PEt accumulation, whereas downregulation of protein kinase C (PKC) obviated agonist-induced PEt production. PMA augmentation of PLD activity proceeded independent of cytosolic free Ca2+ concentration. Ca2+ derived from either intracellular or extracellular sources enhanced ET-related PEt accumulation but was without effect in PKC-downregulated cells. Collectively, these observations indicate that ET stimulates PLD production in RMIC. PKC is the major regulator of this process, with Ca2+ playing a secondary, modulatory role. In addition, these data suggest that PC-PLD is coupled to the ETA receptor.

Carvedilol-induced elevation in cytosolic free Ca2+ level and apoptosis in SIRC corneal epithelial cells

2009 ◽  
Vol 29 (6) ◽  
pp. 477-487
Author(s):  
Pochuen Shieh ◽  
Chih-Hung Lee ◽  
Ng Ling Yi ◽  
Chung-Ren Jan
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Phospholipase C ◽  
Dependent Manner ◽  
Corneal Epithelial Cells ◽  
Concentration Dependent Manner ◽  
Corneal Epithelial ◽  
Kinase C

The effect of the cardiovascular drug carvedilol on cytosolic free Ca2+ concentrations ([Ca 2+]i) and viability was examined in Statens Seruminstitut rabbit cornea (SIRC) corneal epithelial cells. [Ca2+]i and cell viability were measured using the fluorescent dyes fura-2 and 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] (WST-1), respectively. Carvedilol at concentrations between 1 and 30 μM increased [Ca 2+]i in a concentration-dependent manner. The Ca2+ signal was reduced partly by removing extracellular Ca2+. Carvedilol induced Mn2+ quench of fura-2 fluorescence implicating Ca2+ influx. The Ca2+ influx was inhibited by suppression of protein kinase C activity. In Ca2+-free medium, after pretreatment with 1 μM thapsigargin (an endoplasmic reticulum Ca 2+ pump inhibitor), carvedilol-induced [Ca2+]i rise was reduced; and conversely, carvedilol pretreatment inhibited a major part of thapsigargin-induced [Ca 2+]i rise. Addition of the phospholipase C inhibitor 1-[6-[[17 beta-3-methoxyestra-1,3,5(10)-trien-17-yl]amino] hexyl]-1H-pyrrole-2,5-dione (U73122; 2 μM) did not change carvedilol-induced [Ca2+]i rise. At concentrations between 5 and 70 μM, carvedilol killed cells in a concentration-dependent manner. The cytotoxic effect of 20 μM carvedilol was not reversed by prechelating cytosolic Ca2+ with BAPTA/AM. Apoptosis was induced by 5—70 μM carvedilol. Collectively, in SIRC corneal epithelial cells, carvedilol-induced [Ca2+]i rises by causing Ca2+ release from the endoplasmic reticulum in a phospholipase C-independent manner, and Ca 2+ influx via protein kinase C-regulated Ca2+ channels. Carvedilol-caused cytotoxicity was mediated by Ca2+-independent apoptosis in a concentration-dependent manner.

scholarly journals G-protein βγ subunits antagonize protein kinase C-dependent phosphorylation and inhibition of phospholipase C-β1

1997 ◽  
Vol 326 (3) ◽  
pp. 701-707 ◽  
Author(s):  
Irene LITOSCH
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
G Protein ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Negative Feedback Regulation ◽  
Kinase C ◽  
Stimulation Of

Protein kinase C (PKC) isoforms phosphorylated phospholipase C-β1 (PLC-β1) in vitro as follows: PKCα ≫ PKCϵ; not PKCζ. PLC-β3 was not phosphorylated by PKCα. G-protein βγ subunits inhibited the PKCα phosphorylation of PLC-β1 in a concentration-dependent manner. Half-maximal inhibition occurred with 500 nM βγ. G-protein βγ subunits also antagonized the PKCα-mediated inhibition of PLC-β1 enzymic activity. PKCα, in turn, inhibited the stimulation of PLC-β1 activity by βγ. There was little effect of PKCα on the stimulation of PLC-β1 by αq/11–guanosine 5′[γ-thio]triphosphate (GTP[S]). These findings demonstrate that G protein βγ subunits antagonize PKCα regulation of PLC-β1. Thus βγ subunits might have a role in modulating the negative feedback regulation of this signalling system by PKC.

The role of protein kinase C in LHRH-induced LH and FSH release and LHRH self-priming in rat anterior pituitary glands in vitro

1988 ◽  
Vol 116 (2) ◽  
pp. 231-239 ◽  
Author(s):  
M. S. Johnson ◽  
R. Mitchell ◽  
G. Fink
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Esters ◽  
Polymyxin B ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Pituitary Glands

ABSTRACT We have investigated the role of protein kinase C (PKC) in LHRH-induced LH and FSH secretion and LHRH priming. Hemipituitary glands from pro-oestrous rats were incubated with agents known to affect PKC and with or without LHRH, during which time the secretion of gonadotrophins was measured. Phorbol esters and phospholipase C, activators of PKC, released LH and FSH in a concentration-dependent manner and potentiated the LHRH-induced secretion of gonadotrophins in parallel with their ability to release these hormones alone. Inhibitors of PKC had either no effect on LH release (1-(5-isoquinolinesulphonyl)-2-methylpiperazine hydrochloride) or they augmented LHRH-induced gonadotrophin release (polymyxin B and 8-(N,N-diethylamino) octyl-3,4,5-trimethoxybenzoate). Neither the activators nor the inhibitors of PKC, when present with LHRH, caused any change in LHRH priming, even though the activators alone produced a release of gonadotrophins that showed a temporal pattern similar to that produced by LHRH priming. The profiles of effects on LH and FSH secretion were always qualitatively similar. These results show that PKC may be involved in general regulation of gonadotrophin release but that it is not important in acute responses to LHRH nor in LHRH self-priming. J. Endocr. (1988) 116, 231–239

scholarly journals Modulation of Calmodulin and Protein Kinase C Activities by Pencillium Mycotoxins

1999 ◽  
Vol 18 (2) ◽  
pp. 91-96 ◽  
Author(s):  
I. Pala ◽  
A. Srinivasan ◽  
P. J. S. Vig ◽  
D. Desaiah
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Complex Formation ◽  
Mammalian Brain ◽  
Dependent Manner ◽  
Active Conformation ◽  
Cellular Functions ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Ic50 Values

Calmodulin (CaM), a calcium-binding protein, is found in high concentrations in mammalian brain where it plays a pivotal role in a large number of cellular functions. Protein kinase C (PKC), a multifunctional cytosolic enzyme, in the presence of both Ca2+ and phospholipids, transduce extracellular signals into intracellu-lar events. Both CaM and PKC are partially involved in maintaining Ca2+ homeostasis in the cell. Any fluctuations in the intracel-lular Ca2+ can modulate cellular functions and may contribute to neuronal dysfunction. Hence, the present investigation was initiated to study the effects of some selected penicillium (naturally occurring tremorgenic) mycotoxins like secalonic acid, citreoviridin, and verruculogen on CaM activity, active conformation of CaM and PKC activity. Stimulation of CaM-deflcient bovine brain 3′-5′ phosphodieste rase (PDE) indicated CaM activity. The modification of CaM active conformation was studied by the binding of fluorescent probe N-phenyl-1-napthylamine (NPN) to CaM. Alterations in the fluorescence of dansyl-CaM was used to study the effect of these compounds on complex formation between CaM and PDE. Rat brain cytosolic PKC was studied using 32P-ATP as a measure of altered protein phosphorylation. The concentrations of mycotoxins used were in the range of 10 to 50 μM. All three mycotoxins inhibited CaM-stimulated PDE activity in a concentration-dependent manner. Citreoviridin and secalonic acid inhibited NPN fluorescence and Ca2+-dependent complex formation of dansyl-CaM and PDE. The IC50 values for NPN fluorescence of citreoviridin and secalonic acid were 13 μM and 19 μM respectively. However, verruculogen showed little effect on NPN fluorescence and the Ca2+-dependent complex formation of dansyl-CaM and PDE. These mycotoxins also inhibited PKC activity in a concentration-dependent manner with IC50 values of 19.8, 25.7, and 38.4 μM for secalonic acid, citreoviridin, and verruculogen, respectively. The results of our study suggest that these mycotoxins at very low concentrations are interacting with CaM and PKC. Such an effect could lead to impairment of neurotransmission and result in neurotoxicity.

scholarly journals The diacylglycerols dioctanoylglycerol and oleoylacetylglycerol enhance prostaglandin synthesis by inhibition of the lysophosphatide acyltransferase

1987 ◽  
Vol 247 (3) ◽  
pp. 773-777 ◽  
Author(s):  
M Goppelt-Strübe ◽  
H J Pfannkuche ◽  
D Gemsa ◽  
K Resch
Keyword(s):  
Arachidonic Acid ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Direct Interaction ◽  
Prostaglandin E ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Key Enzyme ◽  
Free Arachidonic Acid

Prostanoids are synthesized by resident macrophages upon stimulation with diacylglycerols. Oleoylacetylglycerol and dioctanoylglycerol induced prostaglandin E and thromboxane synthesis in a time- and concentration-dependent manner. Both diacylglycerols inhibited the lysophosphatide acyltransferase, which is the key enzyme in the reacylation of arachidonic acid. By this mechanism the pool of free arachidonic acid available for prostanoid synthesis is increased. Both diacylglycerols were able to inhibit the membrane-bound lysophosphatide acyltransferase by a direct interaction independent of protein kinase C. Thus lysophosphatide acyltransferase could be shown to be a new target of these diacylglycerols, known as activators of protein kinase C.

Resveratrol Attenuates Adenosine Diphosphate-Induced Platelet Activation by Reducing Protein Kinase C Activity

2008 ◽  
Vol 36 (03) ◽  
pp. 603-613 ◽  
Author(s):  
Yu-Min Yang ◽  
Xing-Xiang Wang ◽  
Jun-Zhu Chen ◽  
Shi-Jun Wang ◽  
Hu Hu ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Platelet Aggregation ◽  
Protein Kinase ◽  
Platelet Activation ◽  
Thrombus Formation ◽  
Adenosine Diphosphate ◽  
Dependent Manner ◽  
Chinese Medicinal Herb ◽  
Concentration Dependent Manner ◽  
Kinase C

Inappropriate platelet activation is the key point of thrombogenesis. The aim of the present study was to investigate the effects of resveratrol (RESV), a compound extracted from the Chinese medicinal herb Polygonum cuspidatum sieb et Zucc, on the platelet activation induced by adenosine diphosphate (ADP) and its possible mechanism. The percentage of platelet aggregation and surface P-selectin-positive platelets, and the activity of protein kinase C (PKC) of platelet were observed with platelet aggregometer, flow cytometry and phosphorimaging system, respectively. RESV at 25, 50 and 100 μM showed anti-platelet aggregation and inhibition of surface P-selectin-positive platelets in a concentration-dependent manner. RESV (50 μM) inhibited the activity of PKC in the membrane fraction of platelets and decreased the percentage of membrane associated PKC activity in total PKC activity. Moreover, DL-erythro-1,3-Dihydroxy-2-aminooctadecane, an elective protein kinase C inhibitor (PKCI), and RESV had additive effects of inhibiting the percentage of platelet aggregation and surface P-selectin-positive platelets. It is suggested that RESV may inhibit platelet aggregation, the percentage of surface P-selectin-positive platelets and subsequent thrombus formation. The mechanisms may be partly relative to the decrease of the activity of PKC of platelets.

PGE2 regulates cAMP production in cultured rabbit CCD cells: evidence for dual inhibitory mechanisms

1992 ◽  
Vol 263 (6) ◽  
pp. C1208-C1215 ◽  
Author(s):  
T. D. Noland ◽  
C. E. Carter ◽  
H. R. Jacobson ◽  
M. D. Breyer
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Collecting Duct ◽  
Dependent Manner ◽  
Cortical Collecting Duct ◽  
Camp Production ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Inhibitory Mechanisms ◽  
Protein Kinase C Inhibitor

In cultured cortical collecting duct (CCD) cells, exogenous prostaglandin E2 (PGE2) inhibited arginine vasopressin (AVP)-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) production in a concentration-dependent manner. Although pertussis toxin (PT, 500 ng/ml) alone did not reverse the PGE2-dependent inhibition, PT and staurosporine, a protein kinase C inhibitor, together partially reversed the effect of exogenous PGE2. In contrast, PT completely reversed the inhibition of AVP-dependent cAMP production by sulprostone. These data suggest that exogenous PGE2 can inhibit AVP-stimulated cAMP production and that the inhibitory effects of PGE2 are mediated by staurosporine- and PT-sensitive component(s). Short-term (15-240 min) incubation with phorbol 12-myristate 13-acetate (PMA, 10(-7) M) inhibited PGE2-stimulated cAMP production. Long-term (20 h) incubation with PMA augmented PGE2-stimulated cAMP production. These data provide evidence for the maintenance of a PT-sensitive PGE2-dependent inhibitory pathway of cAMP production in cultured CCD cells. In addition, data are presented that support an inhibitory role for protein kinase C in the effects of PGE2 on the metabolism of cAMP in these cells.

Interactions between inhibitory and excitatory modulatory signals in single submucosal neurons

1994 ◽  
Vol 267 (5) ◽  
pp. C1359-C1365 ◽  
Author(s):  
C. Barajas-Lopez
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Substance P ◽  
Adenylyl Cyclase ◽  
Somatostatin Receptors ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Cyclic Monophosphate ◽  
Kinase C ◽  
Excitatory Responses

Intracellular recordings were made in submucosal neurons from the guinea pig ileum to study the actions of norepinephrine and somatostatin on slow depolarizations induced by 2-chloroadenosine (CADO) and substance P. Local application (by pressure) of CADO and substance P induced a slow depolarization that occurred concomitantly with an increase in input membrane resistance. Norepinephrine, UK-14304 (alpha 2-adrenoceptor agonist), and somatostatin blocked the excitatory responses induced by CADO in a concentration-dependent manner. The alpha 2-adrenoceptor antagonists idazoxan and yohimbine antagonized these inhibitory effects of UK-14304 and norepinephrine. UK-14304 also decreased depolarizations induced by forskolin, but not those induced by the adenosine 3',5'-cyclic monophosphate analogue 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate. Slow depolarizations induced by substance P were blocked neither by UK-14304 nor by somatostatin. It was previously shown that staurosporine (an inhibitor of various protein kinases) and KT-5720 (an inhibitor of protein kinase A) inhibited slow depolarizations induced by CADO. Here, substance P depolarizations were inhibited by staurosporine and calphostin C (a blocker of protein kinase C) but not by KT-5720. In conclusion, activation of alpha 2-adrenoceptors and somatostatin receptors selectively blocks excitatory responses induced by CADO, most likely by inhibition of adenylyl cyclase and via pertussis toxin-sensitive G proteins. Slow depolarizations induced by substance P are independent of adenylyl cyclase activation and involve activation of protein kinase C.

Peptidergic Modulation of an Insect Na+ Current: Role of Protein Kinase A and Protein Kinase C

2001 ◽  
Vol 85 (1) ◽  
pp. 374-383 ◽  
Author(s):  
Dieter Wicher
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Time Constant ◽  
Periplaneta Americana ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Kinase A

The modulation of voltage-gated Na+ currents in isolated somata of dorsal unpaired median (DUM) neurons of the cockroach Periplaneta americana was investigated using the patch-clamp technique. The neuropeptide Neurohormone D (NHD), which belongs to the family of adipokinetic hormones, reversibly reduced the Na+ current in concentration-dependent manner (1 pM to 10 nM). At 10 nM, NHD caused an attenuation of the maximum of current-voltage ( I-V) relation for peak currents by 23 ± 6%. An analysis of NHD action on current kinetics in terms of the Hodgkin-Huxley formalism revealed that NHD reduces the time constant of inactivation, whereas steady-state activation and inactivation as well as the time constant of activation were not affected. In addition, NHD prolonged the recovery from inactivation. The cAMP analogue 8-bromo-cAMP, forskolin, and the catalytic subunit of protein kinase A mimicked the action of NHD. Furthermore, preincubation of cells with the protein kinase A inhibitor KT 5720 abolished the action of NHD. Thus NHD seems to modify the Na+ current via channel phosphorylation by protein kinase A. Activation of protein kinase C by oleoylacetylglycerol (OAG) also reduced the Na+ current, but it did not occlude the action of NHD. On the other hand, inhibition of protein kinase C by chelerythrine or Gö 6976 did not essentially impair the NHD effects.

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