Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.21 (CPT)
 4,580 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Regulation of dihydropyridine (nifedipine)-sensitive calcium influx was studied in rabbit culture proximal tubule cells using the fura 2 fluorescence ratio technique. "Osmo-mechanically induced" swelling of cells by exposure to hypotonic medium (220 mosmol/kgH2O) caused a rapid rise in intracellular calcium that was predominantly due to influx of calcium via both dihydropyridine-sensitive (nifedipine-sensitive) and -insensitive calcium influx pathways. The dihydropyridine-sensitive pathway was regulated, in part, by the phosphatidylinositol signaling pathway. Inhibition of phospholipase C by treatment with 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (NCDC), inhibition of protein kinase C (PKC) by staurosporine, or long-term (24 h) treatment with phorbol 12-myristate 13-acetate (PMA) to downregulate PKC abolished most of the osmo-induced, dihydropyridine-sensitive calcium influx signal. Short-term (seconds) PMA treatment to activate PKC produced a marked stimulation of both dihydropyridine-sensitive and -insensitive calcium influx in isotonic (2- to 3-fold stimulation) and hypotonic (5-fold stimulation) conditions. In contrast, elevation of adenosine 3',5'-cyclic monophosphate (cAMP) by treatment with forskolin or inhibition of protein kinase A (PKA) by treatment with the cAMP analog, Rp-8-CPT-cAMPS (the Rp diastereoisomer of adenosine 3',5'-cyclic monophosphothionate), had little or no influence on calcium influx, including dihydropyridine-sensitive calcium influx. It is concluded that osmo-mechanical stress activates a dihyropyridine-sensitive calcium influx pathway that is predominantly regulated via the phosphatidylinositol signaling pathway and PKC and not through the cAMP/PKA signaling pathway.
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PMID:Osmo-mechanically sensitive phosphatidylinositol signaling regulates a Ca2+ influx channel in renal epithelial cells. 924 99

Recent studies have shown that beta 2-adrenergic receptor (beta 2-AR)-stimulated increases in the intracellular Ca2+ (Cai) transient and contraction in cardiac myocytes are dissociated from the increase in adenosine 3',5'-cyclic monophosphate (cAMP) level and are not accompanied by an increase in phospholamban phosphorylation, an acceleration in relaxation, or a reduction in myofilament Ca2+ response. Thus we hypothesized that the beta 2-AR modulation of cardiac excitation-contraction (EC) coupling may be mediated by either a cAMP-independent mechanism or a compartmentalized cAMP pathway. To directly distinguish between these two possibilities, the responses of the L-type Ca2+ current (ICa), Cai transient, and contraction to beta 2-AR as well as to beta 1-AR stimulation were examined in rat ventricular myocytes in the presence or absence of specific inhibitory cAMP analogs, Rp diastereomers of adenosine 3',5'-cyclic monophosphothioate (Rp-cAMPS) and 8-(4-chlorophenylthio)-cAMP (Rp-CPT-cAMPS). As expected, the positive inotropic effect induced by an adenylyl cyclase activator, forskolin (2 x 10(-7) M), or a beta 1-AR agonist, norepinephrine (5 x 10(-8) M) plus prazosin (10(-6) M), was completely blocked by Rp-CPT-cAMPS. More importantly, the responses of ICa, Cai transient, and contraction to beta 2-AR stimulation by zinterol (10(-5) M) or isoproterenol plus a selective beta 1-AR antagonist, CGP-20712A, were also entirely abolished by Rp-cAMPS (in the patch-pipette solution) or Rp-CPT-cAMPS (in the bath solution). In pertussis toxin-treated cells, although the response of cAMP was not altered, the beta 2-AR-stimulated increase in contraction amplitude was markedly enhanced and accompanied by a hastened relaxation, resulting in a tight association between cAMP and contraction. These results indicate that beta 2-AR modulation of cardiac excitation-contraction coupling requires cAMP. The dissociation of beta 2-AR-stimulated cAMP production and regulation of myofilament and sarcoplasmic reticulum functions is attributable to a functional compartmentation of the cAMP-dependent signaling due to an activation of beta 2-AR-coupled Gi and/or G(o).
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PMID:Localized cAMP-dependent signaling mediates beta 2-adrenergic modulation of cardiac excitation-contraction coupling. 932 56

1. The patch-clamp technique was used in conjunction with the fluorescent Ca2+ indicator indo-1 to measure simultaneously cytosolic Ca2+ concentration ([Ca2+]i) and membrane potential in single rat corticotrophs identified with the reverse haemolytic plaque assay. 2. Application of the adrenocorticotropin (ACTH) secretagogue, corticotropin-releasing hormone (CRH), triggered a sustained [Ca2+]i elevation and membrane depolarization. 3. The CRH action was mediated via the cAMP-dependent protein kinase cascade. Both the CRH-induced depolarization and [Ca2+]i elevation could be mimicked by extracellular application of the adenylate cyclase activator forskolin or the membrane-permeable cAMP analogue, 8-(4-chlorophenylthio)-adenosine-3',5'-cyclic monophosphate (8-CPT-cAMP). Intracellular adenosine cyclic 3',5'-(Rp)-phosphothioate (Rp-cAMPS), a protein kinase A inhibitor, abolished the CRH effects. 4. Voltage-clamp studies suggest that the CRH-triggered depolarization was due to the reduction of background K+ conductances. The CRH-sensitive current was Ca2+ independent and was insensitive to the K+ channel blockers tetraethylammonium (TEA) or 4-aminopyridine (4-AP), but could be partially inhibited by Ba2+. 5. The CRH-triggered steady-state depolarization stimulated extracellular Ca2+ entry via voltage-gated Ca2+ channels and raised [Ca2+]i. CRH failed to stimulate [Ca2+]i rise in cells that were voltage clamped at their resting potential. Removal of extracellular Ca2+ or inhibition of Ca2+ channels by Ni2+ abolished the [Ca2+]i rise. 6. Voltage-clamp studies of voltage-gated Ca2+ channels using Ba2+ as charge carrier show that approximately 90% of the channels were available for activation at the resting potential. CRH did not enhance the voltage-gated Ca2+ channels.
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PMID:Mechanism underlying corticotropin-releasing hormone (CRH) triggered cytosolic Ca2+ rise in identified rat corticotrophs. 936 11

von Willebrand factor (vWF) is stored and released from endothelial secretory granules called Weibel-Palade (WP) bodies. Acute release can be induced by thrombin, histamine, and other mediators of thrombosis or inflammation. Their effect is thought to be mediated by an increase in intracellular free calcium ([Ca2+]i). Purine nucleotides such as adenosine triphosphate (ATP) and adenosine diphosphate (ADP) are released from platelet dense granules and from ischemic tissues and are important regulators of platelet function and vascular tone. In the present study, we investigated whether they could also induce exocytosis from cultured endothelial cells. ATP (1 to 100 micromol/L) induced a dose-related increase in vWF release, with a 2.3-fold maximal increase after 30 minutes. Similar responses were observed with ADP. ATP induced calcium mobilization from intracellular stores, an effect mimicked by 2-methylthio-ATP, a selective agonist for P2y receptors. However, 2-methylthio-ATP-induced vWF release was only 43% of the ATP response. ATP-induced vWF release was also associated with a twofold increase in cellular cyclic adenosine monophosphate (cAMP) content, and was potentiated by 3-isobutyl-1-methylxanthine ([IBMX] added to increase cAMP levels by blocking cellular phosphodiesterases) and 8-bromo-cAMP and inhibited by more than 50% by Rp-8-CPT-cAMPS, a competitive protein kinase A inhibitor. Adenosine but not 2-methylthio-ATP mimicked the ATP-induced increase in cAMP. ATP-induced vWF release was partly inhibited by adenosine deaminase, which degrades adenosine generated from ATP in the incubation medium. Adenosine (1 to 100 micromol/L) failed to induce vWF release, but potentiated the secretory response to 2-methylthio-ATP and thrombin without modifying the calcium response to these agents. Our results suggest that ATP/ADP can induce vWF release from endothelial cells via dual activation of P2y and adenosine A2 receptors. ATP/ADP-induced exocytosis could be involved in the regulation of thrombus formation and ischemia-reperfusion injuries. Further, we provide evidence that a receptor-mediated increase in cellular cAMP can potentiate the secretory response to calcium-mobilizing agents.
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PMID:Purine nucleotides induce regulated secretion of von Willebrand factor: involvement of cytosolic Ca2+ and cyclic adenosine monophosphate-dependent signaling in endothelial exocytosis. 941 75

Current data suggest that apoptosis controls neutrophil numbers in tissues. We analyzed roles for and the sites of action for the cAMP-dependent protein kinases (cAPKs) in apoptosis induced in human neutrophils by in vitro storage, cycloheximide (CHX) exposure, and anti-Fas exposure. Treatment with 8-chlorophenylthio-cAMP (8-CPT-cAMP) prolonged the time required for 50% of the cells to exhibit apoptotic morphology (t50) from 16.3 to 41.8 h (in vitro culture), from 2.4 to 7.8 h (CHX), and from 4.8 to 6.5 h (anti-Fas). CHX +/- 8-CPT-cAMP did not significantly alter resting intracellular calcium levels and H-89, a selective inhibitor of cAPK, had no effect on apoptosis in the absence of the analogue. In contrast, site-selective cAMP analogues that specifically activated the type I cAPK, but not type II cAPK, synergistically attenuated apoptosis. Exposure to 8-CPT-cAMP delayed, in parallel, the activity of caspase-3 (CPP-32beta), whereas mitogen-activated protein kinase kinase (MAPKK) inhibitor, PD98059, had no effect on CHX-induced apoptosis +/- 8-CPT-cAMP. Together these results indicate that type I cAPK activation is necessary and sufficient to mediate cAMP-induced delay in human neutrophil apoptosis induced by several mechanisms and suggest that one of the major sites of cAPK action is upstream of caspase-3 (CPP-32beta) activation.
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PMID:Type I cAMP-dependent protein kinase delays apoptosis in human neutrophils at a site upstream of caspase-3. 950 73

This study aimed to characterize the cellular pathways along which nitric oxide (NO) influences the secretion of renin from the kidney. Using the isolated perfused rat kidney model, we found that the NO donor sodium nitroprusside (SNP) (1-30 mumol/l) induced a prompt, concentration-dependent fourfold increase of basal renin secretion. The membrane-permeable cGMP analogs 8-bromo-cGMP and 8-(4-chlorophenylthio)-cGMP (8-pCPT-cGMP; each 5-50 mumol/l) inhibited basal renin secretion and attenuated the stimulation of renin secretion by SNP. Conversely, the renin stimulatory effect of SNP was enhanced in the presence of the G kinase inhibitor Rp-8-CPT-cGMPS (10 mumol/l). The renin stimulatory effect of SNP was amplified in nominally calcium-free perfusate and was abolished in the presence of angiotensin II (1 nmol/l). Renin secretion stimulated by SNP was clearly attenuated by the A kinase inhibitor Rp-8-CPT-cAMPS (25 mumol/l). These findings indicate that the renin stimulatory effect of NO donors in renal juxtaglomerular cells cannot be explained by activation of G kinase and is also less likely to be causally related to the regulation of renin secretion by calcium. Because A kinase activity is required for the stimulation of renin secretion by SNP, it appears as if the renin stimulatory effect is causally related to the cAMP pathway controlling renin secretion.
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PMID:Stimulation of renin secretion by NO donors is related to the cAMP pathway. 957 95

Adenylyl cyclase (AC) modulation of vesicular cycling was visualized at cultured cerebellar granule cell synapses using the sequential uptake of antibodies directed against the intraluminal domain of synaptotagmin I. Vesicle recycling due to spontaneous transmitter release in the absence of action potentials was increased by the AC/protein kinase A (PKA) activators forskolin and CPT-cAMP. These effects were blocked by the PKA inhibitor Rp-cAMPs. Cyclic AMP elevation also induced new cycling at previously silent sites. Activation of L-AP4-sensitive mGluR reduced the cAMP/PKA enhancement at preexisting synapses downstream of both AC and calcium channels. Modulation of the turnover and the number of vesicular release sites provide one mechanism that may underlie cAMP-dependent cerebellar long-term potentiation.
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PMID:Visualization of cyclic AMP-regulated presynaptic activity at cerebellar granule cells. 958 68

The mechanism of malonyl-CoA-independent acute control of hepatic carnitine palmitoyltransferase I (CPT-I) activity was investigated. In a first series of experiments, the possible involvement of the cytoskeleton in the control of CPT-I activity was studied. The results of these investigations can be summarized as follows. (i) Very mild treatment of permeabilized hepatocytes with trypsin produced around 50% stimulation of CPT-I activity. This effect was absent in cells that had been pretreated with okadaic acid (OA) and seemed to be due to the action of trypsin on cell component(s) distinct from CPT-I. (ii) Incubation of intact hepatocytes with 3, 3'-iminodipropionitrile, a disruptor of intermediate filaments, increased CPT-I activity in a non-additive manner with respect to OA. Taxol, a stabilizer of the cytoskeleton, prevented the OA- and 3, 3'-iminodipropionitrile-induced stimulation of CPT-I. (iii) CPT-I activity in isolated mitochondria was depressed in a dose-dependent fashion by the addition of a total cytoskeleton fraction and a cytokeratin-enriched cytoskeletal fraction, the latter being 3 times more potent than the former. In a second series of experiments, the possible link between Ca2+/calmodulin-dependent protein kinase II (Ca2+/CM-PKII) and the cytoskeleton was studied in the context of CPT-I regulation. The data of these experiments indicate that (i) purified Ca2+/CM-PKII activated CPT-I in permeabilized hepatocytes but not in isolated mitochondria, (ii) purified Ca2+/CM-PKII abrogated the inhibition of CPT-I of isolated mitochondria induced by a cytokeratin-enriched fraction, and (iii) the Ca2+/CM-PKII inhibitor KN-62 prevented the OA-induced phosphorylation of cytokeratins in intact hepatocytes. Results thus support a novel mechanism of short-term control of hepatic CPT-I activity which may rely on the cascade Ca2+/CM-PKII activation --> cytokeratin phosphorylation --> CPT-I de-inhibition.
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PMID:Malonyl-CoA-independent acute control of hepatic carnitine palmitoyltransferase I activity. Role of Ca2+/calmodulin-dependent protein kinase II and cytoskeletal components. 970 78

Regulation of expression of functional voltage-gated ion channels for inward currents was studied in Schwann cells in organotypic cultures of dorsal root ganglia from E19 mouse embryos maintained in serum-free medium. Of the Schwann cells that did not contact axons, 46.5% expressed T-type Ca2+ conductances (ICaT). Two days or more after excision of the ganglia, and consequent disappearance of neurites, ICaT were detectable in only 10.9% of the cells, and the marker 04 disappeared. On Schwann cells deprived of neurons, T- (but not L-) type Ca2+ conductances were re-induced by weakly hydrolysable analogues of cAMP, and by forskolin (an activator of adenylyl cyclase) after long-term treatment (4 days). With CPT cAMP (0.1-2 mM), 8Br cAMP, db cAMP or forskolin (0.01 or 0.1 mM), the proportion of cells with ICaT was not significantly different from the proportion in the cultures with neurons. These agents also induced expression in some cells of tetrodotoxin-resistant Na+ currents, which were rarely induced by neurons, but 04 was not re-induced by cAMP analogue treatments that re-induced ICaT. Inward currents (Ba2+ or Na+) were partly restored (P < 0.05) on Schwann cells cultured for 6-7 days beneath a filter bearing cultured neurons. In contrast, addition of neuron-conditioned medium was ineffective. The results suggest that neurons activate, via diffusible and degradable factors, a subset of Schwann cell cAMP pathways leading to expression of IcaT, and activate additional non-cAMP pathways that lead to expression of 04.
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PMID:Functional Ca2+ and Na+ channels on mouse Schwann cells cultured in serum-free medium: regulation by a diffusible factor from neurons and by cAMP. 975 Nov 51

In AtT20 mouse corticotroph tumour cells large conductance Ca2+-activated K+-channels (BK-channels) have an essential role in the early glucocorticoid inhibition of adrenocorticotrophin (ACTH) secretion evoked by corticotrophin-releasing factor. The present study examined whether or not BK-channels are also pivotal to glucocorticoid inhibition of normal rat anterior pituitary cells. A membrane-permeant, non-metabolizable cyclic AMP analogue, 8-(4-Chlorophenylthio)adenosine-3',5'-cyclic-monophosphate (CPT-cAMP) was used as the primary secretagogue stimulus, as this mimics the increase of intracellular cyclic AMP caused by corticotrophin-releasing factor, but is not subject to the complex Ca2+-dependent regulation of cyclic AMP metabolism that is evident in corticotroph cells. Experiments in AtT20 cells showed that ACTH secretion stimulated by 1 mM CPT-cAMP was suppressed to 34+/-1.5% (n = 12) of the control stimulus by a maximal dose of 100 nM dexamethasone. The ACTH secretion evoked by the combination of 1 mM CPT-cAMP with either 5 microm (-)BayK8644 (L-type Ca2+-channel activator) or 5 mM TEA (K+-channel blocker) was respectively 69.1+/-7.6% and 69.3+/-11.8% of control after 2 h preincubation with 100 nM dexamethasone (P<0.05 vs CPT-cAMP). The ACTH response elicited by 5 microM (-)BayK8644 and 5 mM TEA given together was completely resistant to inhibition by 100 nM dexamethasone. Furthermore, TEA and (-)BayK8644 given together synergistically stimulated ACTH release in combination with 0.1 mM or 1 mM CPT-cAMP, and these ACTH responses were not inhibited by 100 nM dexamethasone. In primary cultures of rat anterior pituitary cells, TEA (up to 20 mM), charybdotoxin (30 nM) or apamin (100 nM) failed to modify the glucocorticoid inhibition of 0.1 mM CPT-cAMP-induced ACTH release. The combination of 5 mM TEA and 5 microM (-)BayK8644 elicited a small but significant increase in ACTH secretion but did not modify the inhibition of 0.3 mM CPT-cAMP-induced ACTH secretion by 100 nM dexamethasone. In primary cultures of rat anterior pituitary cells, depolarization of the membrane potential with 40 mM KCl enhanced the ACTH response to CPT-cAMP and markedly reduced the maximal inhibitory effect of dexamethasone to 55+/-1.2% as well as that of corticosterone to 33+/-2.1% vs 100+/-2.5% and 100+/-1.9% inhibition respectively, when 0.1 mM CPT-cAMP was used alone. Introduction of 5 microM (-)BayK8644 with 40 mM KCl in this system had no additional effect on glucocorticoid inhibition. No glucocorticoid inhibition of ACTH release to any of the stimuli applied was observed in cells pretreated with the mRNA synthesis inhibitor, 5,6-dichloro-furanosyl-benzimidazole riboside (DRB) (0.1 mM) or the protein synthesis blocker, puromycin (0.1 mM). In summary, early glucocorticoid inhibition of stimulated ACTH release by cultured rat anterior pituitary cells was dependent on the synthesis of new mRNA and protein. Depolarization of the membrane potential potentiated CPT-cAMP-induced ACTH secretion in AtT20 cells as well as cultured rat corticotrophs and this was associated with a resistance to the early inhibitory effect of glucocorticoids. Glucocorticoid inhibition in rat anterior pituitary corticotrophs was unaltered by TEA, charybdotoxin as well as apamin, and hence it is unlikely to involve predominantly BK-or SK-type Ca2+-activated K+-channels. These results support the thesis that a prime target of glucocorticoid feedback inhibition in anterior pituitary corticotrophs is the membrane potential and indicate that glucocorticoid-induced proteins regulate the activities of several distinct plasma membrane ion channels.
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PMID:Depolarization counteracts glucocorticoid inhibition of adenohypophysical corticotroph cells. 975 91


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