Activation of phospholipase C (PLC) is a central component of the signal transduction process in numerous cells, including platelets. U73122 has been widely used as a selective PLC inhibitor. In the present study, the effects of U73122 on platelet function have been further examined. Platelets were stimulated with collagen (via PLC-gamma), the stable thromboxane mimetic U46619 (via PLC-beta), or phorbol myristate acetate (PMA) via protein kinase C (PKC). Consistent with inhibition of PLC, U73122 inhibited platelet aggregation and [3H]-serotonin release in response to collagen and U46619 in a concentration-dependent manner. Similarly, U73122 blocked collagen-induced release of thromboxane A2. U73122 also inhibited U46619-induced [32P]phosphatidic acid production and phosphorylation of the major PKC substrate, pleckstrin. U73122 had no effect on PMA-induced pleckstrin phosphorylation, [3H]-serotonin release, or intracellular vacuole formation. However, U73122 did inhibit PMA-induced platelet aggregation and fibrinogen binding. Overall, these results suggest that U73122, in addition to its inhibition of PLC, also affects PKC-independent events that interfere with platelet aggregation.
"Next, it was demonstrated that administration of SR141716A before the injection of ANA either icv or into the PPTg blocked the sleep-inducing effects of ANA . Furthermore, if activity of PLC, which is coupled to the CB 1 cannabinoid receptors [113, 114] was prevented using U73122 (a PLC inhibitor)   the sleep-promoting properties of ANA were also blocked . "
[Show abstract][Hide abstract] ABSTRACT: The endocannabinoid system comprises amides, esters and ethers of long chain polyunsaturated fatty acids. Narachidonoylethanolamide (anandamide; ANA) and 2-arachidonoylglycerol (2-AG) are endogenous cannabinoids (endocannabinoids) ligands for the cannabinoid family of G-protein-coupled receptors named CB1 and CB2. Endocannabinoids are released upon demand from lipid precursors in a receptor-dependent manner and behave as retrograde signaling messengers, as well as modulators of postsynaptic transmission, interacting with other neurotransmitters systems. The two principal enzymes that are responsible for the metabolism of ANA and 2-AG are fatty acid amide hydrolase and monoacylglycerol lipase, respectively. Pharmacological experiments have shown that the administration of endocannabinoids induce cannabimimetic effects, including sleep promotion. This review will focus on some of the current evidence of the pharmacological potential of the endocannabinoid system on sleep modulation.
Current Medicinal Chemistry - Central Nervous System Agents 09/2011; 11(3):189-96. DOI:10.2174/187152411798047780
"We next turned our attention to the signaling events occurring after the Frizzled receptor and especially the possible role of phospolipase C (PLC) signaling, which is known to be activated by Gβγ (Ahumada et al., 2002). Addition of a specific PLC inhibitor, u73122 (Lockhart and McNicol, 1999) to cells expressing β 2 AR/RFz1 or β 2 AR/RFz2 and induced with Iso prevented the loss of Dishevelled (Figure 5D). Because PLC involvement occurs after Gβγ and before Ca +2 /PKC the pathway involved in the downregulation of Dishevelled is expected to follow this order (Wnt/Frizzled→Gβγ→ PLC→Ca +2 /PKC signaling) (Figure 4C-E and 5D). "
[Show abstract][Hide abstract] ABSTRACT: Wnt signaling is known to be important for diverse embryonic and post-natal cellular events and be regulated by the proteins Dishevelled and Axin. Although Dishevelled is activated by Wnt and involved in signal transduction, it is not clear how Dishevelled-mediated signaling is turned off. We report that guanine nucleotide binding protein beta 2 (Gnb2; Gbeta2) bound to Axin and Gbeta2 inhibited Wnt mediated reporter activity. The inhibition involved reduction of the level of Dishevelled, and the Gbeta2gamma2 mediated reduction of Dishevelled was countered by increased expression of Axin. Consistent with these effects in HEK293T cells, injection of Gbeta2gamma2 into Xenopus embryos inhibited the formation of secondary axes induced either by XWnt8 or Dishevelled, but not by beta-catenin. The DEP domain of Dishevelled is necessary for both interaction with Gbeta2gamma2 and subsequent degradation of Dishevelled via the lysosomal pathway. Signaling induced by Gbeta2gamma2 is required because a mutant of Gbeta2, Gbeta2 (W332A) with lower signaling activity, had reduced ability to downregulate the level of Dishevelled. Activation of Wnt signaling by either of two methods, increased Frizzled signaling or transient transfection of Wnt, also led to increased degradation of Dishevelled and the induced Dishevelled loss is dependent on Gbeta1 and Gbeta2. Other studies with agents that interfere with PLC action and calcium signaling suggested that loss of Dishevelled is mediated through the following pathway: Wnt/Frizzled-->Gbetagamma-->PLC-->Ca(+2)/PKC signaling. Together the evidence suggests a novel negative feedback mechanism in which Gbeta2gamma2 inhibits Wnt signaling by degradation of Dishevelled.
Experimental and Molecular Medicine 07/2009; 41(10):695-706. DOI:10.3858/emm.2009.41.10.076 · 3.45 Impact Factor
"Phospholipase D (PLD) is also involved in ABA-induced stomatal closure (Sang et al., 2001), but the action of this enzyme is believed to be either independent or downstream of elevations in [Ca 2 ] cyt (Jacob et al., 1999). Previous works (MacRobbie, 2000; Staxen et al., 1999) on the role of PI-PLC in guard cell ABA signalling have largely relied on indirect approaches, such as the use of the PLC inhibitor U73122 which in animals can have non-speci®c effects (Cenni and Picard, 1999; Lockhart and McNicol, 1999; Pulcinelli et al., 1998; Walker et al., 1998). Therefore, we generated transgenic tobacco plants with reduced levels of PI-PLC in their guard cells. "
[Show abstract][Hide abstract] ABSTRACT: The calcium-releasing second messenger inositol 1,4,5-trisphosphate is involved in the regulation of stomatal aperture by ABA. In other signalling pathways, inositol 1,4,5-trisphosphate is generated by the action of phospholipase C. We have studied the importance of phospholipase C in guard cell ABA-signalling pathways. Immunolocalisation of a calcium-activated phospholipase C confirmed the presence of phospholipase C in tobacco guard cells. Transgenic tobacco plants with considerably reduced levels of phospholipase C in their guard cells were only partially able to regulate their stomatal apertures in response to ABA. These results suggest that phospholipase C is involved in the amplification of the calcium signal responsible for reductions in stomatal aperture in response to ABA. As full ABA-induced inhibition of stomatal opening was not observed, our results support a role for the action of other calcium-releasing second messengers in the guard cell ABA-signalling pathway. It is not known whether these different calcium-releasing second messengers act in the same or parallel ABA-signalling pathways.
The Plant Journal 05/2003; 34(1):47-55. DOI:10.1046/j.1365-313X.2003.01698.x · 5.97 Impact Factor
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