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Query: CAS:113-59-7 (CPT)
 3,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The signal transduction cascade between the activation of the somatostatin (SOM) receptor and modulation of transmitter release was study using Acetylcholine (Ach) release measurements and patch clamp recordings of Ca2+ current from acutely dissociated St 40 ciliary ganglion neurons. As in intact synapses, somal ACh release was blocked by 100 nM SOM or 100 microM dibutyril cGMP, and the SOM-mediated inhibition could be reversed by 10 microM 1-NAME (a selective inhibitor of nitric oxide synthase, NOS) or 100 microM Rp-8p-CPT-cGMPs (a selective inhibitor of a cGMP protein dependent kinase, PKG). In whole cell recordings, SOM inhibition of Ca2+ current rapidly relaxes to control levels but is sustained in perforated patch recordings which decreases cell dialysis. Inhibition of NOS or PKG in perforated patch recordings, however caused SOM effects to become transient again. We hypothesize that PKG alters the characteristics of the membrane-delimited G protein inhibition of Ca2+ current. Therefore SOM receptors trigger a membrane-delimited signal transduction cascade that is modulated by soluble messengers, converging on voltage activated Ca2+ channels. When both pathways are active together, SOM causes a sustained inhibition of neuronal Ca2+ current leading to a decrease in transmitter release.
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PMID:Membrane delimited and intracellular soluble pathways in the somatostatin modulation of ACh release. 863 27

The effects of cyclic GMP (cGMP) and activation of cGMP-dependent protein kinase (PKG) on the phosphorylation of the inositol 1,4, 5-trisphosphate (IP3) receptor were examined in intact rat aorta using the technique of back phosphorylation. Aorta treated with the nitric oxide donors, S-nitroso-N-acetylpenicillamine and sodium nitroprusside, or the selective PKG activator, 8-(4-para-chlorophenylthio)-cGMP (8-CPT-cGMP), demonstrated increased IP3 receptor phosphorylation in situ, which was both time- and concentration-dependent with a stoichiometry of 0.5 mol of phosphate/mol of receptor above control. Treatment of aorta with the adenyl cyclase activator, forskolin, also demonstrated increased phosphorylation of the IP3 receptor on the PKG site, although the selective cAMP-dependent protein kinase activator, 8-(4-para-chlorophenylthio)-cAMP (8-CPT-cAMP), did not increase the phosphorylation of the IP3 receptor. Moreover, the PKG selective inhibitor, KT 5823, inhibited both sodium nitroprusside and forskolin-induced IP3 receptor phosphorylation more potently than the selective cAMP-dependent protein kinase inhibitor, KT 5720, suggesting that PKG mediates the increase in IP3 receptor phosphorylation by both cyclic nucleotides in intact aorta. These results provide further support for the notion that PKG is activated by both cAMP and cGMP in intact vascular smooth muscle and that PKG performs a critical role in cyclic nucleotide-dependent relaxation of blood vessels.
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PMID:Phosphorylation of the inositol 1,4,5-trisphosphate receptor. Cyclic GMP-dependent protein kinase mediates cAMP and cGMP dependent phosphorylation in the intact rat aorta. 870 97

Nitric oxide (NO) donors were recently shown to produce biphasic contractile effects in cardiac tissue, with augmentation at low NO levels and depression at high NO levels. We examined the subcellular mechanisms involved in the opposing effects of NO on cardiac contraction and investigated whether NO modulates contraction exclusively via guanylyl cyclase (GC) activation or whether some contribution occurs via cGMP/PKG-independent mechanisms, in indo 1-loaded adult cardiac myocytes. Whereas a high concentration of the NO donor S-nitroso-N-acetylpenicillamine (SNAP, 100 micromol/L) significantly attenuated contraction amplitude by 24.4+/-4.5% (without changing the Ca2+ transient or total cAMP), a low concentration of SNAP (1 micromol/L) significantly increased contraction amplitude (38+/-10%), Ca2+ transient (26+/-10%), and cAMP levels (from 6.2 to 8.5 pmol/mg of protein). The negative contractile response of 100 micromol/L SNAP was completely abolished in the presence of the specific blocker of PKG KT 5823 (1 micromol/L); the positive contractile response of 1 micromol/L SNAP persisted, despite the presence of the selective inhibitor of GC 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 micromol/L) alone, but was completely abolished in the presence of ODQ plus the specific inhibitory cAMP analog Rp-8-CPT-cAMPS (100 micromol/L), as well as by the NO scavenger oxyhemoglobin. Parallel experiments in cell suspensions showed significant increases in adenylyl cyclase (AC) activity at low concentrations (0.1 to 1 micromol/L) of SNAP (AC, 18% to 20% above basal activity). We conclude that NO can regulate both AC and GC in cardiac myocytes. High levels of NO induce large increases in cGMP and a negative inotropic effect mediated by a PKG-dependent reduction in myofilament responsiveness to Ca2+. Low levels of NO increase cAMP, at least in part, by a novel cGMP-independent activation of AC and induce a positive contractile response.
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PMID:Activation of distinct cAMP-dependent and cGMP-dependent pathways by nitric oxide in cardiac myocytes. 1032 39

The hypothesis that cAMP-dependent protein kinase (protein kinase A; PKA) is in an active state in small arteries possessing a myogenic tone was investigated in pressurized rat tail small arteries. At a pressure of 80 mmHg, these vessels constricted to 71.6 +/- 1.0% (n = 32) of the diameter of the fully relaxed state. The PKA inhibitors Rp-8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphothioate (Rp-CPT-cAMPS) and N-(2-([3-(4-bromophenyl)-2-propenyl]amino)-ethyl)-5- isoquinolinesulfonamide HCl (H-89) constricted these vessels dose dependently. For example, 300 microM Rp-CPT-cAMPS and 9 microM H-89 reduced vessel diameter by 11.0 +/- 1.2% (n = 8) and 14.3 +/- 3.6% (n = 5), respectively. The cGMP-dependent protein kinase (protein kinase G; PKG) inhibitor Rp-8-bromo-beta-phenyl-1,N(2)-etheno-guanosine 3', 5'-cyclic monophosphothioate (Rp-8-Br-PET-cGMPS) did not alter vessel diameter up to a concentration of 10 microM. Neither endothelium removal nor inhibition of neural transmission affected the action of Rp-CPT-cAMPS. The effect of 300 microM Rp-CPT-cAMPS was reduced by 82% after pretreatment of the vessel with 100 nM iberiotoxin, a blocker of calcium-activated potassium (K(Ca)) channels. However, the effect of 300 microM Rp-CPT-cAMPS was not altered after pretreatment with 1 mM 4-aminopyridine, a blocker of delayed rectifier potassium channels, or 10 microM ryanodine, a blocker of ryanodine receptor-generated calcium sparks. In inside-out patch-clamp experiments on cells isolated from rat tail small arteries, 10 U/ml of the catalytic subunit of PKA together with 100 microM MgATP increased K(Ca) channel activity 30.1 +/- 9. 8-fold (n = 9). Additionally, neither inhibition of PKA or PKG nor moderate activation of PKA or PKG altered the vessel response to a pressure step from 80 to 120 mmHg. These results suggest that in rat tail small arteries possessing a myogenic tone 1) PKA is in an active state modulating the level of the myogenic tone, and 2) K(Ca) channels mediate, at least partly, this effect of PKA.
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PMID:cAMP-dependent protein kinase is in an active state in rat small arteries possessing a myogenic tone. 1048 37

The effect of a selective cyclic guanocine 3',5'-monophosphate (cGMP)-dependent protein kinase Ialpha inhibitor, Rp-8-[(4-chlorophenyl)thio]-cGMPS triethylamine (Rp-8-p-CPT-CGMPS), on either N-methyl-D-aspartate (NMDA)- or N-ethyl-2-(1-ethyl-2-hydroxy-2-nitrosohydrazino)ethanamine (NOC-12, a nitric oxide (NO) donor)-produced thermal hyperalgesia was examined in the rat. Intrathecal administration of NMDA (15 pg/10 microl) or NOC-12 (10, 20 and 30 microg/10 microl) produced a marked curtailment of the tail-flick latency. Maximal NMDA- or NOC-12-produced facilitation of the tail-flick reflex was significantly and dose-dependently blocked by intrathecal pretreatment with Rp-8-p-CPT-CGMPS (7.5, 15 and 30 microg/10 microl). Rp-8-p-CPT-CGMPS given alone did not markedly alter baseline tail-flick latency. These results suggest that the activation of cGMP-dependent protein kinase Ialpha is required for NMDA- or NO-produced facilitation of thermal hyperalgesia at the spinal cord level.
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PMID:Activation of cGMP-dependent protein kinase Ialpha is required for N-methyl-D-aspartate- or nitric oxide-produced spinal thermal hyperalgesia. 1076 67

cAMP-dependent vasodilators are used to treat a variety of cardiovascular disorders; however, the signal transduction pathways and effector mechanisms stimulated by these agents are not fully understood. In the present study we demonstrate that cAMP-stimulating agents enhance the activity of the large-conductance, calcium-activated potassium (BK(Ca)) channel in single myocytes from coronary arteries by "cross-activation" of the cGMP-dependent protein kinase (protein kinase G, PKG). Single-channel patch-clamp data revealed that 10 micromol/L isoproterenol, forskolin, or dopamine opens BK(Ca) channels in coronary myocytes and that this effect is attenuated by inhibitors of PKG (KT5823; Rp-8-pCPT-cGMPS), but not by inhibiting the cAMP-dependent protein kinase (protein kinase A, PKA). In addition, a membrane-permeable analog, CPT-cAMP, also opened BK(Ca) channels in these myocytes, and this effect was reversed by KT5823. Direct biochemical measurement confirmed that dopamine or forskolin stimulates PKG activity in coronary arteries but does not elevate cGMP. Finally, the stimulatory effect of cAMP on BK(Ca) channels was reconstituted in a cell-free, inside-out patch by addition of purified PKG activated by either cGMP or cAMP. In contrast, channel gating was unaffected by exposure to the purified catalytic subunit of PKA. In summary, findings from on-cell and cell-free patch-clamp experiments provide direct evidence that cAMP-dependent vasodilators open BK(Ca) channels in coronary myocytes by cross-activation of PKG (but not via PKA). Biochemical assay confirmed this cross-activation mechanism of cAMP action in these arteries. This signaling pathway is a novel mechanism for regulation of potassium channel activity in vascular smooth muscle and other cells.
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PMID:cAMP-dependent vasodilators cross-activate the cGMP-dependent protein kinase to stimulate BK(Ca) channel activity in coronary artery smooth muscle cells. 1078 13

We hypothesized that nitric oxide (NO) plays an important role in mediating the anti-adrenergic effect of adenosine on atrioventricular (AV) nodal conduction. In guinea-pig hearts instrumented for measurement of AV nodal conduction time (atrium-to-His bundle, A-H, interval), the NO synthase (NOS) inhibitor, l-NMMA (100 microm), reversibly inhibited 80% (P=0.009, n=6) of adenosine's anti-adrenergic action on the positive dromotropic effect of isoproterenol (0.01 microm). In parallel studies carried out in rabbit AV nodal myocytes, intracellular mechanisms whereby NO mediates the inhibitory effect of adenosine on isoproterenol-induced A-H interval shortening were studied. Adenosine (3 microm) inhibited isoproterenol-stimulated (0.1 microm) I(Ca,L)(beta -I(Ca,L)) by 46+/-6% (P<0.001, n=17). Consistent with isolated heart data, the NOS inhibitors, l -NMMA (100 microm) and L-NNA (500 microm) attenuated the effect of adenosine on beta -I(Ca,L)by 69+/-8% (P<0.001, n=16) and 69+/-7% (P<0.001, n=10), respectively. An inhibitor of NO-stimulated guanylyl cyclase LY83538 (40 microm) reduced the inhibitory effect of adenosine on beta -I(Ca,L)by 97+/-6% (P=0.004, n=15). Similarly, the non-specific inhibitor of cAMP-phosphodiesterases IBMX (50 microm) decreased the anti-adrenergic effect of adenosine by 60% (P=0.02, n=6), whereas the extracellular application of the non-hydrolyzeable cAMP analog 8-Br-cAMP (500 microm) prevented this action of adenosine. Activation of cGMP-dependent protein kinase (PKG) by CPT-cGMP (300 microm) diminished beta -I(Ca,L), but to a significantly smaller degree (16+/-4%, P=0.025, n=12) than that caused by adenosine. NO mediates the anti-adrenergic effect of adenosine on AV nodal conduction by a mechanism predominately involving activation of cGMP-dependent cAMP-phosphodiesterase and to a lesser extent activation of PKG.
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PMID:Antagonism of the positive dromotropic effect of isoproterenol by adenosine: role of nitric oxide, cGMP-dependent cAMP-phosphodiesterase and protein kinase G. 1096 24

Effects of cGMP on voltage-gated currents in the somatic membrane of isolated newt olfactory receptor cells were investigated using the whole-cell mode of the patch-clamp technique. Under voltage clamp, membrane depolarization generated time- and voltage-dependent current responses, a transient inward current and a sustained outward current. When cGMP or a membrane permeant analog of cGMP, 8-p-chlorophenylthio-cGMP (CPT-cGMP), was applied to the recorded cell, the amplitude of the transient inward current increased markedly, but that of the sustained outward current did not change significantly. When each current was isolated by pharmacological agents, 0.1 mM CPT-cGMP increased the peak amplitude of a Na(+) current (I(Na)) by approximately 40%, a T-type Ca(2+) current (I(Ca,T)) by approximately 40%, and an L-type Ca(2+)current (I(Ca,L)) by approximately 10%; however it did not change significantly the amplitude of a delayed rectifier K(+) current (I(K)). A selective cGMP-dependent protein kinase inhibitor, KT5823, blocked the enhancement by cGMP of I(Na) and I(Ca,T), suggesting that cGMP increases these currents via cGMP-dependent phosphorylation. Under current-clamp conditions, application of CPT-cGMP lowered the current threshold of action potentials induced by current injection, and increased the maximum spike frequency in response to strong stimuli. We suggest that cGMP may lower the threshold in olfactory perception by decreasing the current threshold to generate spikes, and also prevent the saturation of odor signals by increasing the maximum spike frequency.
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PMID:Modulation by cGMP of the voltage-gated currents in newt olfactory receptor cells. 1124 73

1. The spasmolytic and anti-spasmogenic activity of beta-adrenoceptor agonists on airways smooth muscle is thought to involve activation of the cyclic AMP/cyclic AMP-dependent protein kinase (PKA) cascade. Here we have tested the hypothesis that PKA mediates the anti-spasmogenic activity of isoprenaline and other cyclic AMP-elevating agents in guinea-pig isolated trachea by utilizing a number of cell permeant cyclic AMP analogues that act as competitive 'antagonists' of PKA. 2. Anion-exchange chromatography of guinea-pig tracheae resolved two peaks of PKA activity that corresponded to the type I ( approximately 5%) and type II ( approximately 93%) isoenzymes. 3. Pre-treatment of tracheae with zardaverine (30 microM), vasoactive intestinal peptide (VIP) (1 microM) and the non-selective activator of PKA, Sp-8-CPT-cAMPS (10 microM), produced a non-parallel rightwards shift in the concentration-response curves that described acetylcholine (ACh)-induced tension generation. The type II-selective PKA inhibitor, Rp-8-CPT-cAMPS (300 microM), abolished this effect. 4. Pre-treatment of tracheae with Sp-8-Br-PET-cGMPS (30 microM) produced a non-parallel rightwards shift of the concentration-response curves that described ACh-induced tension generation. The selective cyclic GMP-dependent protein kinase (PKG) inhibitor, Rp-8-pCPT-cGMPS (300 microM), abolished this effect. 5. Pre-treatment of tracheae with isoprenaline (1 microM) produced a 10 fold shift to the right of the ACh concentration-response curve by a mechanism that was unaffected by Rp-8-Br-cAMPS (300 microM, selective inhibitor of type I PKA), Rp-8-CPT-cAMPS (300 microM) and Rp-8-pCPT-cGMPS (300 microM). 6. We conclude that the anti-spasmogenic activity of Sp-8-CPT-cAMPS, zardaverine and VIP in guinea-pig trachea is attributable to activation of the cyclic AMP/PKA cascade whereas isoprenaline suppresses ACh-induced contractions by a mechanism(s) that is independent of PKA and PKG.
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PMID:Evidence that the anti-spasmogenic effect of the beta-adrenoceptor agonist, isoprenaline, on guinea-pig trachealis is not mediated by cyclic AMP-dependent protein kinase. 1149 3

Both nitric oxide (NO) and natriuretic peptides produce apoptosis of vascular smooth muscle cells. However, there is evidence that NO induces endothelial cell proliferation, which suggests that there is a difference in the response of endothelial cells to natriuretic peptides. The purpose of this study was to investigate the effect of atrial natriuretic peptide (ANP) on human endothelial cell survival. ANP within the physiological concentration (10(-11) mol/l) induced a 52% increase in the number of human coronary arterial endothelial cells and a 63% increase in human umbilical vein endothelial cells at a low concentration of serum. The increase in cell numbers was blocked by pretreatment with RP8-CPT-cGMP (RP8), a cGMP-dependent protein kinase inhibitor, with wortmannin, an Akt/PKB inhibitor, and with PD-98059, an ERK1/2 inhibitor. In a Transwell migration test, ANP also increased the cell migration, and RP8, wortmannin, and PD-98059 blocked this increase. A wound healing assay was performed to examine the effects of ANP on regeneration in vitro. ANP increased both cell numbers and migration, but the effects were blocked by the above three kinase inhibitors. ANP increased the expression of phospho-Akt and of phospho-ERK1/2 within 1.5 h. These results suggest that ANP can potentiate endothelial regeneration by cGMP-dependent protein kinase stimulation and subsequent Akt and ERK1/2 activations.
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PMID:Physiological concentration of atrial natriuretic peptide induces endothelial regeneration in vitro. 1250 72


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