Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

The effects of pertussis toxin, forskolin, and cAMP analogues on the antinociceptive action of nicotine were examined to investigate the possible involvement of adenylate cyclase and G-proteins in nicotine's antinociceptive effect. Intrathecal injection of pertussis toxin (0.25 and 0.50 micrograms) in mice inhibited nicotine-induced antinociception in the tail-flick test. The effect of the toxin was dose and time dependent. Forskolin, a potent adenylate cyclase activator, and 8-(-4-chlorophenylthio) adenosine-3':5' monophosphate, cyclic (8-CPT-cAMP), a cAMP analogue, inhibited the antinociceptive effects of nicotine in a dose-dependent manner. EGTA reversal of 8-CPT-cAMP's inhibitory effects suggests that calcium may to be involved. These data implicate the possible involvement of a G-protein and a second messenger system (activation of a cAMP-dependent protein kinase and increase in cyclic AMP levels) in nicotine-induced analgesia in mice.
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PMID:Nicotine-induced antinociception in mice: role of G-proteins and adenylate cyclase. 802 3

Effects of cAMP on insulin-stimulated mitogen-activated protein (MAP) kinase pathway were examined using rat hepatoma H4EII cells. MAP kinase was rapidly activated and reached a peak 3 min after the stimulation by insulin. Forskolin (1 microM) and 8(4-chlorophenylthio)cAMP (8-CPT-cAMP) (0.1 mM) inhibited the insulin-stimulated MAP kinase activity. Pretreatment of the cells with H-8 (50 microM), a cAMP-dependent protein kinase inhibitor, enhanced the insulin-stimulated MAP kinase activity and partially restored the inhibitory effect of cAMP. Furthermore, insulin-induced phosphorylation of MAP kinase was inhibited by 8-CPT-cAMP, and the inhibition was restored by H-8. 8-CPT-cAMP did not inhibit the autophosphorylation of insulin receptor. These data indicate that elevation of intracellular cAMP blocks the insulin-stimulated MAP kinase pathway downstream of insulin receptor.
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PMID:cAMP inhibits the insulin-stimulated mitogen-activated protein kinase pathway in rat hepatoma H4EII cells. 804 24

Fura 2 fluorescence measurements were carried out on microperfused rat cortical collecting ducts (CCD) to investigate the effect of adenosine 3',5'-cyclic monophosphate (cAMP) and adenylate cyclase-stimulating hormones on free cytosolic calcium ([Ca2+]i). Forskolin, 3-isobutyl-1-methylxanthine, and 8-(4-chlorophenylthio)-cAMP (CPT-cAMP) all triggered marked and sustained [Ca2+]i variations. Maximal increases elicited by 100 microM CPT-cAMP amounted to 101 +/- 11 nM (mean +/- SE, n = 18). This effect was mostly dependent on the presence of basolateral calcium and totally independent of luminal calcium. It remained unchanged in CCD perfused with sodium-free luminal fluid (82 +/- 10 nM, n = 5), pretreated with 1 mM bath ouabain (113 +/- 20, n = 4), or superfused with sodium-free bath in the presence of ouabain (82 +/- 22, n = 5). The V2 agonist 1-desamino-8-D-arginine vasopressin (DDAVP, 10 nM) increased [Ca2+]i by 57 +/- 5 nM (n = 27), a value 40% lower than that achieved with 10 nM AVP (141 +/- 7, n = 34) but similar to that observed with AVP + a V1a antagonist (57 +/- 6, n = 6). Significant effects could also be obtained with 200 pM DDAVP (31 +/- 6, n = 8) and arginine vasopressin (AVP) (72 +/- 6, n = 16). Rat calcitonin also raised [Ca2+]i by 43 +/- 10 (n = 8) and 66 +/- 8 nM (n = 17) at 1 and 10 nM, respectively, and its effect was not additive to that of CPT-cAMP. Calcitonin and DDAVP effects, like those of CPT-cAMP and forskolin, were nearly abolished in Ca(2+)-free bath, but AVP action on intracellular release persisted. These results show that, in rat CCD, cAMP effects on [Ca2+]i mainly result from basolateral calcium entry. In contrast to rabbit CCD the mechanism is independent on Na reabsorption and basolateral Na+/Ca2+ exchange. Calcitonin and DDAVP effects on [Ca2+]i are probably secondary to increased cAMP production.
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PMID:cAMP-dependent effects of vasopressin and calcitonin on cytosolic calcium in rat CCD. 809 49

1. Voltage-activated calcium currents participate in shaping the firing pattern of neurons. Calcium currents also have a role in signal transduction. In the retina, little is known of the regulation of calcium entry into neurons via voltage-activated channels. In the present series of experiments we used standard whole cell and perforated patch clamp techniques to study the ability of the neurotransmitter dopamine (DA) to modulate voltage-dependent calcium currents in isolated turtle retinal ganglion cells. 2. Two types of calcium current have been described in these cells, one transient and the other sustained. Here we focused our studies primarily on the sustained current (ICa). Exogenous DA reduced ICa in some cells (59%), facilitated ICa in others (17%), or had no effect on the remainder (24%). Regardless of the action of DA, there was no effect on the voltage dependence of ICa. In addition, the effects were all reversible. The average magnitude of decrease was 43%, whereas that of increase was 75%. 3. The application of a specific D1 receptor agonist, SKF38393, mimicked the effect of DA. This was also true for a membrane permeable cyclic AMP (cAMP) analogue (8-CPT-cAMP). Inhibition of protein kinase A (PKA) activity by a specific inhibitor, IP20-amide, injected into cells prevented the modulatory effects of DA on ICa. 4. Immunocytochemical studies demonstrated that DA stimulation of the retina significantly increased the level of cAMP immunoreactivity in peripheral ganglion cells, whereas those cells in central retina were less affected. Forskolin induced a general elevation of cytoplasmic cAMP staining in all ganglion cells. 5. Current clamp experiments were carried out to determine the role of the calcium currents in action potential generation. Both the sustained and transient currents participated in the shaping of current-induced firing patterns of isolated cells. Depolarizing current-induced spiking of ganglion cells was found to be highly modified by dopamine. 6. These results support the notion that endogenous DA modulates the conductance of voltage-dependent calcium channels in turtle retinal ganglion cells and that this modulation is mediated by a D1 dopamine receptor-cAMP-PKA pathway. The direct result of this modulation is an alteration in the signaling properties of certain cells.
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PMID:Calcium currents in turtle retinal ganglion cells. II. Dopamine modulation via a cyclic AMP-dependent mechanism. 817 36

The role of carnitine palmitoyltransferase I (CPT-I) in the control of ketogenesis was studied in primary cultures of rat astrocytes. Ketone bodies were the major product of [14C]palmitate oxidation by cultured astrocytes, whereas CO2 made a minor contribution to the total oxidation products. Using tetradecylglycidate as a specific, cell-permeable inhibitor of CPT-I, a flux control coefficient of 0.77 +/- 0.07 was calculated for CPT-I over the flux of [14C]palmitate to ketone bodies. CPT-I from astrocytes was sensitive to malonyl-CoA (IC50 = 3.4 +/- 0.8 microM) and cross-reacted on western blots with an antibody raised against liver CPT-I. On the other hand, astrocytes expressed significant acetyl-CoA carboxylase (ACC) activity, and consequently they contained considerable amounts of malonyl-CoA. Western blot analysis of ACC isoforms showed that ACC in astrocytes--like in neurons, liver, and white adipose tissue--mostly comprised the 265-kDa isoform, whereas the 280-kDa isoform--which was highly expressed in skeletal muscle--showed much lower abundance. Forskolin was used as a tool to study the modulation of the ketogenic pathway in astrocytes. Thus, forskolin decreased in parallel ACC activity and intracellular malonyl-CoA levels, whereas it stimulated CPT-I activity and [14C]palmitate oxidation to both ketone bodies and CO2. Results show that in cultured astrocytes (a) CPT-I exerts a very high degree of control over ketogenesis from palmitate, (b) the ACC/malonyl-CoA/CPT-I system is similar to that of liver, and (c) the ACC/malonyl-CoA/CPT-I system is subject to regulation by cyclic AMP.
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PMID:Role of carnitine palmitoyltransferase I in the control of ketogenesis in primary cultures of rat astrocytes. 975 Nov 93

Incubating 3T3-L1 adipocytes with forskolin, which increases intracellular cAMP by activating adenylate cyclase, mimicked rapamycin by attenuating the effect of insulin on stimulating the phosphorylation of four (S/T)P sites in PHAS-I, a downstream target of the mammalian target of rapamycin (mTOR) signaling pathway. To investigate the hypothesis that increasing cAMP inhibits mTOR, the protein kinase activity of mTOR was measured in an immune complex assay with recombinant PHAS-I as substrate. Both forskolin and 8-(4-chlorophenylthio)adenosine 3'-5'-monophosphate (CPT-cAMP) prevented the activation of mTOR by insulin in adipocytes, but neither agent affected mTOR activity when added directly to the immunopurified protein. In contrast, the cAMP phosphodiesterase inhibitor, theophylline, inhibited mTOR activity not only when added to intact adipocytes but also when added to immunopurified mTOR in vitro, demonstrating that certain methylxanthines are able to inhibit mTOR independently of increasing cAMP. Forskolin and CPT-cAMP blocked the effect of insulin on increasing mTOR phosphorylation, which was assessed using mTAb1, an antibody whose binding is inhibited by phosphorylation of mTOR. Although the mTAb1 epitope contains a consensus site for protein kinase B, neither agent inhibited the activation of protein kinase B produced by insulin. These findings support the interpretation that increasing cAMP attenuates the effects of insulin on PHAS-I, p70(S6K), and other downstream targets of the mTOR signaling pathway by inhibiting the phosphorylation and activation of mTOR.
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PMID:Attenuation of mammalian target of rapamycin activity by increased cAMP in 3T3-L1 adipocytes. 985 18

The ability of Toxoplasma gondii tachyzoites to differentiate into latent bradyzoite forms is essential for pathogenesis of clinical disease. We examined the effects of cyclic nucleotides on T. gondii bradyzoite differentiation in vitro. Differentiation of tachyzoites to bradyzoites was measured in an immunofluorescence assay using ME49 or its clonal derivative PLK, two well-characterized T. gondii strains. Treatment of human fibroblast cultures infected with T. gondii with 8-(4-chlorophenylthio)-cyclic GMP (CPT-cGMP), a membrane-permeable, nonhydrolyzable analogue of cGMP, resulted in an increased percentage of bradyzoite-positive vacuoles. Cyclic AMP (cAMP) also induced in vitro conversion of PLK, but the method of cAMP elevation was critical. Forskolin raises cAMP levels transiently and induced bradyzoites, whereas agents predicted to cause sustained elevation of cAMP were inhibitory to parasite conversion. Levels of cAMP were measured in host cells and extracellular tachyzoites. Forskolin, CPT-cGMP, and agents known to induce bradyzoite formation elevated cAMP in host cells and PLK parasites. These data suggest cyclic nucleotide signaling pathways are important in the stress-induced conversion of T. gondii tachyzoites to bradyzoites. Furthermore, because cAMP elevation was seen in PLK but not RH, a T. gondii strain that did not differentiate well in our assay, cAMP signaling within the parasite is likely to be critical.
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PMID:Cyclic nucleotide signaling in Toxoplasma gondii bradyzoite differentiation. 1111

Pigmented (PE) and nonpigmented (NPE) ciliary epithelial cells comprise the ciliary epithelium, the site of aqueous humor formation in the eye. In man, catecholamines increase the rate of aqueous humor formation, but the mechanism underlying these effects is not understood. Recent evidence suggests that Na-K-Cl cotransport plays a central role in blood-to-aqueous chloride transport across ciliary epithelium in cow and rabbit. We therefore investigated whether catecholamines stimulate Na-K-Cl cotransport in human PE cells. Na-K-Cl cotransporter protein was detected as a 170 kDa protein band on immunoblots. Immunofluorescence microscopy detected cotransporter on the basolateral membranes of the PE layer of ciliary epithelium from a human donor. Cotransporter immunofluorescence was also detected in cultured PE cells. Na-K-Cl cotransport activity measured as ouabain-insensitive bumetanide-sensitive(86)Rb uptake was stimulated by isoproterenol 1.6-fold, with an EC(50) = 28 n M and maximal stimulation at 1 microM. Other transport mechanisms involved in(86)Rb uptake were not affected. Stimulation by 1 microM isoproterenol was blocked by 10 n M ICI 118,551, a beta(2)-specific receptor antagonist, whereas the receptor subtype-specific antagonists yohimbine (alpha(2)), prazosin (alpha(1)) and atenolol (beta(1)) were ineffective. Norepinephrine stimulation (EC(50) = 280 n M) was also blocked by ICI 118,551. Dopamine stimulated Na-K-Cl cotransport 1.6-fold with an EC(50) = 14 microM. The dopamine effect could not be blocked by 10 microM SCH 23390, a D1-antagonist, but was abolished by ICI 118,551. Forskolin and CPT-cAMP stimulated Na-K-Cl cotransport 1.79- and 1.71-fold, respectively, whereas the inactive forskolin analogue 1,9-dideoxyforskolin had no effect. However, high concentrations of the PKA inhibitors PKI amide 14-22 and KT 5720 were needed to inhibit both PKA activity in cell lysates and isoproterenol stimulation of cotransport. This finding may indicate the presence of a novel PKA isoform in PE cells. Inhibitors of other protein kinases, including myosin light chain kinase, protein kinase G, calmodulin-dependent kinase and tyrosine kinase, were without effect on stimulated Na-K-Cl cotransport. When EC(50)s for catecholaminergic stimulations of Na-K-Cl cotransport in PE were compared to those in NPE, values within five-fold of one another were seen for isoproterenol and norepinephrine. In contrast, dopamine was 28-fold more potent in NPE than in PE. The data suggest that both PE and NPE possess beta(2)adrenergic receptors, but only NPE cells possess dopamine D1 receptors linked to Na-K-Cl cotransport.
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PMID:Catecholaminergic regulation of Na-K-Cl cotransport in pigmented ciliary epithelium: differences between PE and NPE. 1113 77

1. The mechanisms and receptors involved in the vasoactive intestinal peptide (VIP)- and pituitary adenylate cyclase-activating polypeptide (PACAP)-induced relaxations of the pig intravesical ureter were investigated. 2. VIP, PACAP 38 and PACAP 27 concentration-dependently relaxed U46619-contracted ureteral strips with a similar potency. [Ala(11,22,28)]-VIP, a VPAC(1) agonist, showed inconsistent relaxations. 3. The neuronal voltage-gated Ca(2+) channel inhibitor, omega-conotoxin GVIA (omega-CgTX, 1 microm), reduced the VIP relaxations. Urothelium removal or blockade of capsaicin-sensitive primary afferents, nitric oxide (NO) synthase and guanylate cyclase with capsaicin (10 microm), N(G)-nitro-l-arginine (l-NOARG, 100 microm) and 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 5 microm), respectively, did not change the VIP relaxations. However, the PACAP 38 relaxations were reduced by omega-CgTX, capsaicin, l-NOARG and ODQ. 4. The VIP and VIP/PACAP receptor antagonists, [Lys(1), Pro(2,5), Arg(3,4), Tyr(6)]-VIP (1 microm) and PACAP (6-38) (0.4 microm), inhibited VIP and VIP and PACAP 38, respectively, relaxations. 5. The nonselective and large-conductance Ca(2)-activated K(+) channel blockers, tetraethylammonium (3 mm) and charybdotoxin (0.1 microm), respectively, and neuropeptide Y (0.1 microm) did not modify the VIP relaxations. The small-conductance Ca(2)-activated K(+) channel blocker apamin (1 microm) did not change the PACAP 27 relaxations. 6. The cAMP-dependent protein kinase A (PKA) blocker, 8-(4-chlorophenylthio)adenosine-3',5'-cyclic monophosphorothioate (Rp-8-CPT-cAMPS, 100 microm), reduced VIP relaxations. The phosphodiesterase 4 inhibitor rolipram and the adenylate cyclase activator forskolin relaxed ureteral preparations. The rolipram relaxations were reduced by Rp-8-CPT-cAMPS. Forskolin (30 nm) evoked a potentiation of VIP relaxations. 7. These results suggest that VIP and PACAP relax the pig ureter through smooth muscle receptors, probably of the VPAC(2) subtype, linked to a cAMP-PKA pathway. Neuronal VPAC receptors localized at motor nerves and PAC(1) receptors placed at sensory nerves and coupled to NO release, seem also to be involved in the VIP and PACAP 38 relaxations.
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PMID:Heterogeneity of neuronal and smooth muscle receptors involved in the VIP- and PACAP-induced relaxations of the pig intravesical ureter. 1466 37

The removal by phagocytosis of degenerated myelin is central for repair in Wallerian degeneration that follows traumatic injury to axons and in autoimmune demyelinating diseases (e.g., multiple sclerosis). We tested for roles played by the cAMP cascade in the regulation of myelin phagocytosis mediated by complement receptor-3 (CR3/MAC-1) and scavenger receptor-AI/II (SRAI/II) separately and combined in mouse microglia and macrophages. Components of the cAMP cascade tested are cAMP, adenylyl cyclase (AC), Gi, protein kinase A (PKA), exchange protein directly activated by cAMP (Epac), and phosphodiesterases (PDE). PKA inhibitors H-89 and PKI(14-22) amide inhibited phagocytosis at normal operating cAMP levels (i.e., those occurring in the absence of reagents that alter cAMP levels), suggesting activation of phagocytosis through PKA at normal cAMP levels. Phagocytosis was inhibited by reagents that elevate endogenous cAMP levels to above normal: Gi-inhibitor Pertussis toxin (PTX), AC activator Forskolin, and PDE inhibitors IBMX and Rolipram. Phagocytosis was inhibited also by cAMP analogues whose addition mimics abnormal elevations in endogenous cAMP levels: nonselective 8-bromo-cAMP, PKA-specific 6-Benz-cAMP, and Epac-specific 8-CPT-2'-O-Me-cAMP, suggesting that abnormal high cAMP levels inhibit phagocytosis through PKA and Epac. Altogether, observations suggest a dual role for cAMP and PKA in phagocytosis: activation at normal cAMP levels and inhibition at higher. Furthermore, a balance between Gi-controlled cAMP production by AC and cAMP degradation by PDE maintains normal operating cAMP levels that enable efficient phagocytosis.
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PMID:cAMP cascade (PKA, Epac, adenylyl cyclase, Gi, and phosphodiesterases) regulates myelin phagocytosis mediated by complement receptor-3 and scavenger receptor-AI/II in microglia and macrophages. 1634 30


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