alpha-Adrenergic augmentation of myogenic response in rat arterioles: role of protein kinase C

1993 ◽  
Vol 264 (2) ◽  
pp. H547-H552 ◽  
Author(s):  
J. Watanabe ◽  
M. Keitoku ◽  
K. Hangai ◽  
A. Karibe ◽  
T. Takishima
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Lumen Diameter ◽  
Myogenic Response ◽  
High Potassium ◽  
Kinase C ◽  
Pkc Inhibitors ◽  
Pkc Activation ◽  
Potassium 40

We tested the hypothesis that protein kinase C (PKC) activation plays a major role in alpha-adrenergic augmentation of the myogenic response in rat isolated arterioles. Lumen diameter measured was with a video-monitored microscopic system. Lumen diameter did not change (131 +/- 5 vs. 126 +/- 6 microns) despite an increase in lumen pressure from 40 to 100 mmHg. Phenylephrine (Phe; 3 x 10(-7) M) augmented the myogenic response, since lumen diameter decreased significantly from 117 +/- 8 to 101 +/- 8 microns. High potassium (40 mM) failed to augment the myogenic response, while constricting the vessels to nearly the same extent as did Phe. PKC inhibitors 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine dihydrochloride (H-7, 5 x 10(-5) M, n = 7) and staurosporine (3 x 10(-9) M, n = 7) abolished the Phe-induced augmentation. H-7 and staurosporine depressed the myogenic response even without Phe. PKC activators phorbol 12,13-dibutyrate (3 x 10(-9) M; n = 7) and 4 beta-phorbol 12-myristate 13-acetate (6 x 10(-8) M; n = 6) constricted the vessels by 11 +/- 2 and 18 +/- 3%, respectively. However, PKC activators failed to augment the myogenic response. These results suggest that PKC activation does not play a major role in alpha-adrenergic augmentation of the myogenic response in rat skeletal arterioles.

PDGF-induced mitogenic signaling is not mediated through protein kinase C and c-fos pathway in VSM cells

1993 ◽  
Vol 264 (1) ◽  
pp. C71-C79 ◽  
Author(s):  
R. V. Sharma ◽  
R. C. Bhalla
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Dna Synthesis ◽  
Thymidine Incorporation ◽  
Cytosolic Calcium ◽  
Platelet Derived Growth Factor ◽  
Kinase C ◽  
Pdgf Stimulation ◽  
Pkc Activation

This study examines the role of protein kinase C (PKC) in platelet-derived growth factor (PDGF)-induced vascular smooth muscle (VSM) cell proliferation and initial signaling events. A 24-h pretreatment of VSM cells with 200 nM phorbol 12-myristate 13-acetate (PMA) completely abolished immunologically reactive PKC activity. Depletion of PKC activity from VSM cells did not attenuate PDGF-stimulated [3H]thymidine incorporation compared with control cells. Similarly, acute activation of PKC by treatment with 200 nM PMA for 10 min had no effect on PDGF-mediated [3H]thymidine incorporation. Both PMA and PDGF increased c-fos induction to the same magnitude; however, treatment with PMA did not induce DNA synthesis in these cells. In PKC-depleted cells PDGF-mediated c-fos induction was reduced by 50-60%, while DNA synthesis in response to PDGF stimulation was not reduced. PKC depletion did not alter PDGF-stimulated increase in cytosolic calcium levels, 125I-PDGF binding, or receptor autophosphorylation. On the basis of these results, we conclude that PKC activation and c-fos induction do not play a significant role in PDGF-mediated mitogenesis in VSM cells.

Mechanism of negative lusitropic effect of alpha 1-adrenoceptor stimulation in cat papillary muscles

1996 ◽  
Vol 270 (2) ◽  
pp. H701-H709 ◽  
Author(s):  
M. Vila-Petroff ◽  
G. N. Perez ◽  
B. Alvarez ◽  
H. E. Cingolani ◽  
A. Mattiazzi
Keyword(s):  
Protein Kinase C ◽  
Relaxation Time ◽  
Protein Kinase ◽  
Maximal Velocity ◽  
Papillary Muscles ◽  
Similar Increase ◽  
Kinase C ◽  
Maximal Tension ◽  
Pkc Inhibitors ◽  
Pkc Activation

Experiments were performed in cat papillary muscles to explore the mechanisms by which alpha 1-adrenoceptor stimulation affects myocardial relaxation. Phenylephrine (PE; 10 microM) + atenolol (1 microM; n = 8 experiments) produced a negative lusitropic effect, i.e., a prolongation of half-relaxation time (t1/2; time to 50% relaxation) by 30 +/- 10% (P < 0.05) and a proportionally smaller increase in maximal velocity of relaxation (-T) than in maximal velocity of contraction (+T), which significantly increased the ratio +T/-T. A similar increase in contractility, produced by increasing calcium, failed to significantly change t1/2 and +T/-T. PE-induced negative lusitropic effect was significantly inhibited by two protein kinase C (PKC) inhibitors, staurosporine (0.1 microM) and chelerythrine (10 microM). PE also increased intracellular pH by 0.18 +/- 0.05 pH units (P < 0.05, n = 4), as measured by the fluorescent dye 2'-7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Intracellular alkalosis and the negative lusitropic effect of PE were prevented by the Na+/H+ exchanger inhibitor ethylisopropylamiloride (10 microM). No significant changes in calcium myofilament sensitivity and maximal tension were detected in trabeculae treated with PE either before or after chemical skinning. These results indicate that a Na+/H+ exchanger-induced intracellular alkalosis, possibly mediated by PKC activation, may fully account for the negative lusitropism of alpha 1-adrenoceptor stimulation.

Protein kinase C regulates endocytosis and recycling of E-cadherin

2002 ◽  
Vol 283 (2) ◽  
pp. C489-C499 ◽  
Author(s):  
Tam Luan Le ◽  
Shannon R. Joseph ◽  
Alpha S. Yap ◽  
Jennifer L. Stow
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Esters ◽  
Recycling Endosomes ◽  
Kinase C ◽  
Pkc Isozymes ◽  
Pkc Activation ◽  
Darby Canine Kidney ◽  
E Cadherin

E-cadherin is a major component of adherens junctions in epithelial cells. We showed previously that a pool of cell surface E-cadherin is constitutively internalized and recycled back to the surface. In the present study, we investigated the potential role of protein kinase C (PKC) in regulating the trafficking of surface E-cadherin in Madin-Darby canine kidney cells. Using surface biotinylation and immunofluorescence, we found that treatment of cells with phorbol esters increased the rate of endocytosis of E-cadherin, resulting in accumulation of E-cadherin in apically localized early or recycling endosomes. The recycling of E-cadherin back to the surface was also decreased in the presence of phorbol esters. Phorbol ester-induced endocytosis of E-cadherin was blocked by specific inhibitors, implicating novel PKC isozymes, such as PKC-ε in this pathway. PKC activation led to changes in the actin cytoskeleton facilitating E-cadherin endocytosis. Depolymerization of actin increased endocytosis of E-cadherin, whereas the PKC-induced uptake of E-cadherin was blocked by the actin stabilizer jasplakinolide. Our findings show that PKC regulates vital steps of E-cadherin trafficking, its endocytosis, and its recycling.

Phorbol 12,13-Diacetate-Induced Contraction of the Canine Basilar Artery: Role of Protein Kinase C

1991 ◽  
Vol 11 (1) ◽  
pp. 135-142 ◽  
Author(s):  
Makoto Sugawa ◽  
Tohru Koide ◽  
Shigetaka Naitoh ◽  
Michiaki Takato ◽  
Tohru Matsui ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Normal Medium ◽  
Kinase C ◽  
Prostaglandin F ◽  
Basilar Arteries ◽  
Canine Basilar Artery ◽  
Biochemical Mechanisms ◽  
Pkc Activation

The pharmacological and biochemical mechanisms of contractile responses to the protein kinase C (PKC) activator phorbol-12, 13-diacetate (PDA) were investigated in canine basilar arteries, In the normal medium, PDA elicited a strong, dose-related, and slow-developing sustained contraction, Among the constrictors examined, including serotonin, prostaglandin F2α and endothelin, only PDA yielded contractions in a 2 Ca2+ -free medium, In both media, the PDA-induced contractions were virtually inhibited by either staurosporine, H-7, or quinacrine, while neither neurotransmitter blockades nor R24571 (calmidazolium) exerted significant effects, In addition, it was shown that 8-bromocyclic GMP, but not 8-bromocyclic AMP, markedly curtailed the PDA-induced contractions, Biochemical analysis, furthermore, showed that PDA induced increased phosphorylations of 27- and 96-kDa and proteins other than the myosin light chain (MLC) 20-kDa protein, Thus, the present results open up a novel mechanism of sustained cerebral artery contractions, where PKC activation rather than Ca2+/calmodulin/MLC system plays a key role that is regulated both by phospholipase A 2 and by cyclic GMP.

scholarly journals Protein kinase A modulates Ca2+- and protein kinase C-dependent amylase release in permeabilized rat pancreatic acini

1992 ◽  
Vol 287 (2) ◽  
pp. 403-406 ◽  
Author(s):  
A J O'Sullivan ◽  
J D Jamieson
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Response Curve ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Kinase C ◽  
Pkc Activation ◽  
Kinase A

The role of protein kinase A (PKA) in the release of amylase from permeabilized pancreatic acini was investigated. Addition of cyclic AMP (cAMP) to permeabilized acini resulted in a potentiation of Ca(2+)-dependent amylase release, shifting the Ca2+ dose/response curve leftwards. As with protein kinase C (PKC) activation, this is due to an increase in the time of active discharge. The effect of cAMP was shown to be blocked by two inhibitors of PKA, H89 and the PKI-(5-24)-peptide. At low concentration, cAMP synergizes from phorbol 12-myristate 13-acetate (PMA), while at optimal concentrations cAMP and PMA are additive. PKA and PKC appear to work via similar, but not identical mechanisms.

Modulation of Na+ transport and epithelial sodium channel expression by protein kinase C in rat alveolar epithelial cells

2005 ◽  
Vol 83 (11) ◽  
pp. 977-987 ◽  
Author(s):  
Toshiyuki Yamagata ◽  
Yuko Yamagata ◽  
Chantal Massé ◽  
Marie-Claude Tessier ◽  
Emmanuelle Brochiero ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Sodium Channel ◽  
Epithelial Sodium Channel ◽  
Alveolar Epithelial Cells ◽  
Alveolar Epithelial ◽  
Kinase C ◽  
Pkc Inhibitors ◽  
Pkc Activation

Although the amiloride-sensitive epithelial sodium channel (ENaC) plays an important role in the modulation of alveolar liquid clearance, the precise mechanism of its regulation in alveolar epithelial cells is still under investigation. Protein kinase C (PKC) has been shown to alter ENaC expression and activity in renal epithelial cells, but much less is known about its role in alveolar epithelial cells. The objective of this study was to determine whether PKC activation modulates ENaC expression and transepithelial Na+ transport in cultured rat alveolar epithelial cells. Alveolar type II cells were isolated and cultured for 3 to 4 d before they were stimulated with phorbol 12-myristate 13-acetate (PMA 100 nmol/L) for 4 to 24 h. PMA treatment significantly decreased α, β, and γENaC expression in a time-dependent manner, whereas an inactive form of phorbol ester had no apparent effect. This inhibitory action was seen with only 5-min exposure to PMA, which suggested that PKC activation was very important for the reduction of αENaC expression. The PKC inhibitors bisindolylmaleimide at 2 µmol/L and Gö6976 at 2 µmol/L diminished the PMA-induced suppression of αENaC expression, while rottlerin at 1 µmol/L had no effect. PMA elicited a decrease in total and amiloride-sensitive current across alveolar epithelial cell monolayers. This decline in amiloride-sensitive current was not blocked by PKC inhibitors except for a partial inhibition with bisindolylmaleimide. PMA induced a decrease in rubidium uptake, indicating potential Na+-K+-ATPase inhibition. However, since ouabain-sensitive current in apically permeabilized epithelial cells was similar in PMA-treated and control cells, the inhibition was most probably related to reduced Na+ entry at the apical surface of the cells. We conclude that PKC activation modulates ENaC expression and probably ENaC activity in alveolar epithelial cells. Ca2+-dependent PKC is potentially involved in this response.Key words: alveolar epithelial cells, Na+ transport, Na+ channel, ENaC, protein kinase C, Na+-K+-ATPase, amiloride, gene expression.

Signal transduction of flumazenil-induced preconditioning in myocytes

2001 ◽  
Vol 280 (3) ◽  
pp. H1249-H1255 ◽  
Author(s):  
Zhenhai Yao ◽  
Bradley C. McPherson ◽  
Huiping Liu ◽  
Zuohui Shao ◽  
Changqing Li ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Cell Death ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Oxygen Radicals ◽  
Oxygen Radical ◽  
Kinase C ◽  
Radical Generation ◽  
Pkc Inhibitors

The objective of this study was to examine the role of oxygen radicals, protein kinase C (PKC), and ATP-sensitive K+(KATP) channels in mediating flumazenil-produced preconditioning. Chick cardiomyocyte death was quantified using propidium iodide, and oxygen radical generation was assessed using 2′,7′-dichlorofluorescin oxidation. Preconditioning was initiated with 10 min of ischemia followed by 10 min of reoxygenation. Alternatively, flumazenil was infused for 10 min and removed 10 min before ischemia. Flumazenil (10 μM) and preconditioning increased oxygen radicals [1,693 ± 101 ( n = 3) and 1,567 ± 98 ( n = 3), respectively, vs. 345 ± 53 ( n = 3) in control] and reduced cell death similarly [22 ± 3% ( n = 5) and 18 ± 2% ( n = 6), respectively, vs. controls 49 ± 5% ( n = 8)]. Protection and increased oxygen radicals by flumazenil were abolished by pretreatment with the antioxidant thiol reductant 2-mercaptopropionyl glycine (800 μM; 52 ± 10%, n = 6). Specific PKC inhibitors Go-6976 (0.1 μM) and chelerythrine (2 μM), given during ischemia and reoxygenation, blocked flumazenil-produced protection (47 ± 5%, n = 6). The PKC activator phorbol 12-myristate 13-acetate (0.2 μM), given during ischemia and reoxygenation, reduced cell death similarly to that with flumazenil [17 ± 4% ( n = 6) and 22 ± 3% ( n = 5)]. Finally, 5-hydroxydecanoate (1 mM), a selective mitochondrial KATP channel antagonist given during ischemia and reoxygenation, abolished the protection of flumazenil and phorbol 12-myristate 13-acetate. Thus flumazenil mimics preconditioning to reduce cell death in cardiomyocytes. Oxygen radicals activate mitochondrial KATP channels via PKC during the process.

Role of protein kinase C in the regulation of ATP-triggered intracellular Ca2+ oscillations in chicken granulosa cells

1996 ◽  
Vol 134 (6) ◽  
pp. 743-750 ◽  
Author(s):  
Paul Morley ◽  
Balu R Chakravarthy ◽  
Geoffrey AR Mealing ◽  
Benjamin K Tsang ◽  
James F Whitfield
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Granulosa Cells ◽  
National Research Council ◽  
Fold Increase ◽  
Oscillatory Activity ◽  
Kinase C ◽  
Intracellular Ca ◽  
Pkc Inhibitors

Morley P. Chakravarthy BR, Mealing GAR, Tsang BK, Whitfield JF. Role of protein kinase C in the regulation of ATP-triggered intracellular Ca2+ oscillations in chicken granulosa cells. Eur J Endocrinol 1996;134:743–50. ISSN 0804–4643 These studies were designed to investigate the role of protein kinase C (PKC) in the regulation of ATP-triggered intracellular Ca2+ ([Ca2+]i) oscillations in chicken granulosa cells. Granulosa cells were obtained from the two largest preovulatory follicles (F1 and F2) of hens and [Ca2+]i was measured in cells loaded with the Ca2+-responsive fluorescent dye fura-2. Adenosine triphosphate (100 μmol/l) triggered an immediate, large [Ca2+]i spike that was followed by oscillations that returned to the resting level between spikes. The ATP (100 μmol/l) also stimulated a 1.70 ± 0.1-fold increase in membrane-associated PKC activity over control levels. The frequency of the ATP-triggered [Ca2+]i oscillations was reduced in a concentration-dependent (1–10 nmol/l) manner by treating the cells for 2 min with a PKC activator, 12-O-tetradecanoyl phorbol-13-acetate (TPA). A higher TPA concentration (100 nmol/l) completely prevented ATP from triggering the initial [Ca2+]i spike and oscillations. Adding TPA during the ATP-triggered [Ca2+]i oscillations immediately stopped the oscillatory activity. Interestingly, PKC inhibitors failed to amplify the ATP-triggered [Ca2+]i oscillations. Instead, adding the PKC inhibitors staurosporine (20 nmol/l), calphostin C (200 nmol/l) or 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H7; 100 μmol/l), either before or during the ATP (100μmol/l)-triggered [Ca2+]i response, also completely blocked the [Ca2+]i oscillations. Therefore, ATP-triggered [Ca2+]i oscillations in chicken granulosa cells appear to be regulated by a negative feedback loop requiring PKC, because the [Ca2+]i oscillations were prevented by either full activation or inhibition of PKC activity. Paul Morley, Institute for Biological Sciences, National Research Council of Canada, Building M54, Montreal Road, Ottawa, Ontario, K1A OR6, Canada

Acute regulation of Na/H exchanger NHE3 activity by protein kinase C: role of NHE3 phosphorylation

1999 ◽  
Vol 276 (5) ◽  
pp. C1205-C1217 ◽  
Author(s):  
Michael R. Wiederkehr ◽  
Hui Zhao ◽  
Orson W. Moe
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Functional Response ◽  
Kinase C ◽  
Hormonal Modulation ◽  
Functional Regulation ◽  
Pkc Activation

Acute hormonal modulation of NHE3 activity is partly mediated by kinases, including protein kinase C (PKC). We examined the role of NHE3 phosphorylation in regulating its activity in response to PKC activation by phorbol 12-myristate 13-acetate (PMA). In pooled NHE-deficient fibroblasts transfected with NHE3, PMA increased NHE3 activity and phosphorylation. When six potential PKC target serines were mutated, NHE3 phosphorylation was drastically reduced and PMA failed to regulate NHE3 phosphorylation or function. To examine whether NHE3 phosphorylation is sufficient for functional regulation by PKC, we exploited the heterogeneous response of NHE3 activity to PMA in individual clones of transfectants. Clones with stimulatory, inhibitory, or null responses to PMA were observed. Despite the diverse functional response, changes in NHE3 phosphorylation as revealed by tryptic phosphopeptide maps were similar in all clones. We conclude that although phosphorylation appears to be necessary, it is insufficient to mediate PKC regulation of NHE3 function and factors extrinsic to the NHE3 protein must be involved.

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