Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies have demonstrated that stimulation of phospholipase C-linked G-protein-coupled receptors, including muscarinic M1 and M3 receptors, increases the release of the soluble form of amyloid precursor protein (sAPPalpha) by alpha-secretase cleavage. In this study, we examined the involvement of capacitative Ca2+ entry (CCE) in the regulation of muscarinic acetylcholine receptor (mAChR)-dependent sAPPalpha release in neuroblastoma SH-SY5Y cells expressing abundant M3 mAChRs. The sAPPalpha release stimulated by mAChR activation was abolished by EGTA, an extracellular Ca2+ chelator, which abolished mAChR-mediated Ca2+ influx without affecting Ca2+ mobilization from intracellular stores. However, mAChR-mediated sAPPalpha release was not inhibited by thapsigargin, which increases basal [Ca2+]i by depletion of Ca2+ from intracellular stores. While these results indicate that the mAChR-mediated increase in sAPPalpha release is regulated largely by Ca2+ influx rather than by Ca2+ mobilization from intracellular stores, we further investigated the Ca2+ entry mechanisms regulating this phenomenon. CCE inhibitors such as Gd3+, SKF96365, and 2-aminoethoxydiphenyl borane (2-APB), dose dependently reduced both Ca2+ influx and sAPPalpha release stimulated by mAChR activation, whereas inhibition of voltage-dependent Ca2+ channels, Na+/Ca2+ exchangers, or Na+-pumps was without effect. These results indicate that CCE plays an important role in the mAChR-mediated release of sAPPalpha.
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PMID:Capacitative Ca2+ entry is involved in regulating soluble amyloid precursor protein (sAPPalpha) release mediated by muscarinic acetylcholine receptor activation in neuroblastoma SH-SY5Y cells. 1652 74

The activation of the Ras-related GTPase R-Ras, which has been implicated in the regulation of various cellular functions, by G protein-coupled receptors (GPCRs) was studied in HEK-293 cells stably expressing the M3 muscarinic acetylcholine receptor (mAChR), which can couple to several types of heterotrimeric G proteins. Activation of the receptor induced a very rapid and transient activation of R-Ras. Studies with inhibitors and activators of various signaling pathways indicated that R-Ras activation by the M3 mAChR is dependent on cyclic AMP formation but is independent of protein kinase A. Similar to the rather promiscuous M3 mAChR, two typical G(s)-coupled receptors also induced R-Ras activation. The receptor actions were mimicked by an Epac-specific cyclic AMP analog and suppressed by depletion of endogenous Epac1 by small interfering RNAs, as well as expression of a cyclic AMP binding-deficient Epac1 mutant, but not by expression of dominant negative Rap GTPases. In vitro studies demonstrated that Epac1 directly interacts with R-Ras and catalyzes GDP/GTP exchange at this GTPase. Finally, it is shown that the cyclic AMP- and Epac-activated R-Ras plays a major role in the M3 mAChR-mediated stimulation of phospholipase D but not phospholipase C. Collectively, our data indicate that GPCRs rapidly activate R-Ras, that R-Ras activation by the GPCRs is apparently directly induced by cyclic AMP-regulated Epac proteins, and that activated R-Ras specifically controls GPCR-mediated phospholipase D stimulation.
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PMID:Cyclic AMP-dependent and Epac-mediated activation of R-Ras by G protein-coupled receptors leads to phospholipase D stimulation. 1675 64

Neurotrophins are important modulators of synaptic function at both developing and mature synapses in the CNS and PNS. At the neuromuscular junction (NMJ), neurotrophins, as well as perisynaptic Schwann cells (PSCs) are critical for the long-term maintenance and stability of the synapse. Considering this correlation and the acute interactions that occur at the synapse between PSCs and the nerve terminal, we wondered if neurotrophins could also be involved in neuron-glia signalling. To test if neurotrophins were able to signal to PSCs we used brief applications of neurotrophin-3 (NT-3), brain-derived neurotophic factor (BDNF) or nerve growth factor (NGF; 100 ng/mL). Soleus muscles of mice were incubated with the Ca(2+) indicator Fluo-4AM and Ca(2+) responses in PSCs were elicited through nerve stimulation (50 Hz, 30 s). Our results indicate that acute application of both NT-3 and BDNF, but not NGF, increased PSC Ca(2+) responses. Investigation of the mechanisms involved in these increases revealed distinct pathways for BDNF and NT-3. BDNF increased PSC responsiveness through potentiation of ATP responses while NT-3 modulated muscarinic acetylcholine receptor signalling. Using local applications of the neurotrophins, we found that both neurotrophins were able to elicit Ca(2+) responses in PSCs where BDNF used a phospholipase C-inositol 1,4,5-triphosphate (PLC-IP(3)) mechanism, while NT-3 required extracellular Ca(2+). Our results demonstrate a neurotrophin-dependent modulation of neuron-glia signalling through differential mechanisms employed by NT-3 and BDNF. Hence, neurotrophins precisely and differentially regulate PSC functions through modulation of either purinergic or cholinergic signalling pathways.
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PMID:Neurotrophins modulate neuron-glia interactions at a vertebrate synapse. 1735 53

M-channels are voltage-gated K+ channels that regulate the excitability of many neurons. They are composed of Kv7 (KCNQ) family subunits, usually Kv7.2 + Kv7.3. Native M-channels and expressed Kv7.2 + 7.3 channels are inhibited by stimulating G(q/11)-coupled receptors - prototypically the M1 muscarinic acetylcholine receptor (M1-mAChR). The channels require membrane phosphatidylinositol-4,5-bisphosphate (PIP(2)) to open and the effects of mAChR stimulation result primarily from the reduction in membrane PIP(2) levels following G(q)/phospholipase C-catalysed PIP(2) hydrolysis. However, in sympathetic neurons, M-current inhibition by bradykinin appears to be mediated through the release and action of intracellular Ca(2)+ by inositol-1,4,5-trisphosphate (IP(3)), a product of PIP(2) hydrolysis, rather than by PIP(2) depletion. We have therefore compared the effects of bradykinin and oxotremorine-M (a muscarinic agonist) on membrane PIP(2) in sympathetic neurons using a fluorescently tagged mutated C-domain of the PIP(2) binding probe, 'tubby'. In concentrations producing equal M-current inhibition, bradykinin produced about one-quarter of the reduction in PIP(2) produced by oxotremorine-M, but equal reduction when PIP(2) synthesis was blocked with wortmannin. Likewise, wortmannin restored bradykinin-induced M-current inhibition when Ca(2)+ release was prevented with thapsigargin. Thus, inhibition by bradykinin can use product (IP(3)/Ca(2)+)-dependent or substrate (PIP(2)) dependent mechanisms, depending on Ca(2)+ availability and PIP(2) synthesis rates.
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PMID:Regulation of M(Kv7.2/7.3) channels in neurons by PIP(2) and products of PIP(2) hydrolysis: significance for receptor-mediated inhibition. 1739 26

Invertebrate visual iG(q)alpha is homologous to mammalian mG(q)alpha in two of the three domains important for G protein interaction with receptors; the C-terminus and the linker regions that connect the helical and ras-like domains of the alpha subunit. The third receptor-interacting domain, the N-terminus, contains a six amino acid extension MTLESI in mG(q)alpha that is not present in iG(q)alpha. In co-expression studies we assessed the promiscuity and efficacy of receptor coupling to phospholipase C (PLC) by iG(q)alpha, a non-palmitoylated mutant iG(q)alpha(C3,4A), mG(q)alpha and G(15)alpha. The invertebrate G proteins and mG(q)alpha only coupled to G(q)-coupled receptors (m1 muscarinic acetylcholine receptor (mChR1), alpha(1A)-adrenergic receptor (alpha1-AR)) and not to the G(i/s)-coupled receptors (CCR1 receptor, beta2-adrenergic receptor or dopamine D1 receptor) while G(15)alpha coupled to all receptors. iG(q)alpha and iG(q)alpha(C3,4A) both had double the efficacy for PLC activation compared to the mammalian G proteins when co-expressed with mChR1 and alpha1-AR. The increased efficacy of iG(q)alpha compared to mG(q)alpha was also seen downstream of PLC with carbachol stimulation of the mitogen-activated protein kinase, ERK1/2. Addition of the MTLESI extension onto the N-terminus of iG(q)alpha decreased its efficacy by 35% whereas deletion of this sequence from mG(q)alpha increased its efficacy by 60% in the PLC and ERK1/2 assays. iG(q)alpha, iG(q)alpha(C3,4A) and mG(q)alpha all displayed similar receptor-independent AlF(4)(-)activation of PLC and guanosine triphosphate hydrolysis (GTPase) activity. iG(q)alpha, and iG(q)alpha(C3,4A) both had increased receptor-activated guanosine 5'-[gamma-[(35)S]thio]triphosphate ([(35)S]GTPgammaS) binding when compared to mG(q)alpha when co-expressed with the mChR1. These results demonstrate that G(q) protein efficacy is at least partially determined by the presence of the amino-terminal MTLESI extension. Comparison of [(35)S]GTPgammaS binding rates helps explain the increased efficacy of the invertebrate G proteins.
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PMID:Receptor-coupling properties of the invertebrate visual guanine nucleotide binding protein iGqalpha. 1756 78

The production and further metabolism of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] require several calcium-dependent enzymes, but little is known about subsequent calcium-dependent changes in cellular Ins(1,4,5)P3. To study the calcium dependence of muscarinic acetylcholine receptor-induced Ins(1,4,5)P3 increases in PC12h cells, we utilized an Ins(1,4,5)P3 imaging system based on fluorescence resonance energy transfer and using green fluorescent protein variants fused with the pleckstrin homology domain of phospholipase C-delta1. The intracellular calcium concentration, monitored by calcium imaging, was adjusted by thapsigargin pretreatment or alterations in extracellular calcium concentration, enabling rapid receptor-independent changes in calcium concentration via store-operated calcium influx. We found that Ins(1,4,5)P3 production was increased by a combination of receptor- and calcium-dependent components, rather than by calcium alone. The level of Ins(1,4,5)P3 induced by the receptor was found to be half that induced by the combined receptor and calcium components. Increases in calcium levels prior to receptor activation did not affect the subsequent receptor-induced Ins(1,4,5)P3 increase, indicating that calcium does not influence Ins(1,4,5)P3 production without receptor activation. Removal of both the receptor agonists and calcium rapidly restored calcium and Ins(1,4,5)P3 levels, whereas removal of calcium alone restored calcium to its basal concentration. Similar calcium-dependent increases in Ins(1,4,5)P3 were also observed in Chinese hamster ovary cells expressing m1 muscarinic acetylcholine receptor, indicating that the observed calcium dependence is common to Ins(1,4,5)P3 production. To our knowledge, our results are the first showing receptor- and calcium-dependent components within cellular Ins(1,4,5)P3.
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PMID:Receptor- and calcium-dependent induced inositol 1,4,5-trisphosphate increases in PC12h cells as shown by fluorescence resonance energy transfer imaging. 1785 Mar 33

Activity of the cholinergic muscarinic system is associated with modulation of locomotor activity, although the precise mechanism remains unclear. The phospholipase C-beta1 knockout mouse displays both M1 muscarinic receptor dysfunction and a hyperactive locomotor phenotype. This mouse serves as an ideal model for the analysis of muscarinic modulation of locomotor activity. The clozapine metabolite N-desmethylclozapine (NDMC) has shown some promise as an alternative or adjunct treatment for psychotic disorders. NDMC shows strong muscarinic acetylcholine receptor affinities, which may contribute to the clinical efficacy of clozapine and account for the correlation between NDMC/clozapine ratio and treatment response. Administration of NMDC reversed a striking hyperactive phenotype in the phospholipase C-beta1 knockout mouse, whereas no significant effects were observed in wild-type animals. This highlights the potential role of muscarinic activity in the behavioural response to NDMC. The M1 muscarinic antagonist pirenzepine, however, also reduced the hyperactive phenotype of these mice, emphasizing the importance of muscarinic function in the control of locomotor behaviour, but also calling into question the specific mechanism of action of NMDC at muscarinic receptors.
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PMID:Role of muscarinic receptors in the activity of N-desmethylclozapine: reversal of hyperactivity in the phospholipase C knockout mouse. 1869 Jan 8

While it is generally accepted that phospholipase C (PLC) and protein kinase C (PKC) are down-stream proteins involved in metabotropic glutamate receptor 5 (mGluR5)-related signal transduction, we still do not know which subtype of PLC or PKC is specifically regulated after mGluR5 activation. In the present study in mGluR5 wild-type (mGluR5+/+) mice, we showed induced PKCbeta2 or PKCgamma expression at the border between the stratum oriens and alveus (O/A border) at 2h during pilocarpine induced status epilepticus (SE), and in the stratum pyramidale in CA1 area at 1 day after pilocarpine induced SE; at 1 day, induced expression of PLCbeta4 in the stratum pyramidale of CA1 area was observed. Furthermore, double labeling revealed the co-localization of induced PKCbeta2 or PKCgamma with mGluR5 or with induced PLCbeta4 in the stratum pyramidale of CA1 area. These induced expression, however, were not found in mGluR5 mutant (mGluR5-/-) mice. It suggests that induced PLCbeta4-PKCbeta2/PKCgamma at 1 day after pilocarpine induced SE in pyramidal neurons or PKCbeta2 or PKCgamma in interneurons at O/A border at 2h during pilocarpine induced SE may be specifically linked to the activation of mGluR5. When compared to mGluR5+/+ mice, significant shorter latency (from pilocarpine injection to the occurrence of status epilepticus) and maintenance period (from beginning to the end of status epilepticus) for status epilepticus in mGluR5-/- mice were also demonstrated. It is possible that mGluR5 may play a negative role in initiation of status epilepticus by interacting with muscarinic acetylcholine receptor in mGluR5+/+ mice.
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PMID:mGluR5-PLCbeta4-PKCbeta2/PKCgamma pathways in hippocampal CA1 pyramidal neurons in pilocarpine model of status epilepticus in mGluR5+/+ mice. 1877 62

Upon activation of receptors coupled to the Gq subclass of G proteins, phospholipase C (PLC)beta hydrolyses membrane phospholipid to yield a pair of second messengers, inositol 1,4,5-trisphosphate and 1,2-diacylglycerol. Of four PLCbeta isoforms, PLCbeta1 is transcribed predominantly in the telencephalon and its gene inactivation in mice impairs metabotropic glutamate receptor- and muscarinic acetylcholine receptor-dependent hippocampal oscillations, endocannabinoid production in the hippocampus and barrel formation in the somatosensory cortex. Here we examined cellular and subcellular distributions of PLCbeta1 in adult mouse brains. In the telencephalon, high levels of PLCbeta1 were observed in principal neurons, including pyramidal cells in the cortex and hippocampus, granule cells and mossy cells in the dentate gyrus, and medium spiny neurons in the caudate-putamen, whereas most interneurons had low levels of or were negative for PLCbeta1 and, instead, expressed PLCbeta4. By immunofluorescence, tiny clusters of PLCbeta1 were distributed in somatodendritic compartments of principal neurons and positioned close to those of metabotropic glutamate receptor 5, muscarinic acetylcholine receptor M1 and diacylglycerol lipase-alpha, respectively. Immunoelectron microscopy revealed that PLCbeta1 was often associated with the smooth endoplasmic reticulum, cell membrane or postsynaptic density. In particular, it was highly accumulated at the perisynapse of dendritic spines forming asymmetrical synapses. In the cerebellum, PLCbeta1 was generally low but was enriched in axons and dendrites of basket cells. These results suggest that PLCbeta1 is the key effector in telencephalic principal neurons and cerebellar interneurons. Furthermore, the well-orchestrated molecular arrangement appears to be the anatomical basis for the specificity, efficiency and convergence of the neuronal phosphoinositide signaling system.
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PMID:Predominant expression of phospholipase Cbeta1 in telencephalic principal neurons and cerebellar interneurons, and its close association with related signaling molecules in somatodendritic neuronal elements. 1897 90

Exposure to sodium fluoride (NaF) resulted in an increased accumulation (up to 10-fold) of total [(3)H]inositol phosphates (T-InsP) in rat PC 12 cells. The magnitude of the NaF effect was comparable to that for muscarinic acetylcholine receptor-mediated stimulation of T-InsP accumulation in the presence of saturating concentrations of carbachol, but effects of NaF and muscarinic agonists were additive at subsaturating concentrations. The NaF effect was atropine insensitive; was not mimicked by effects of NaCl (10 mM), aluminum fluoride (1 to 100 muM), forskolin (up to 100 muM), or dibutyryl cyclic AMP (1 mM); and was not altered by treatment with pertussis or cholera toxins (1 mug/ml for 24 h). By contrast, the carbachol response was fully sensitive to atropine and partly sensitive to pertussis toxin. Chelation of extracellular calcium ion following 10 min of pretreatment with EDTA or EGTA (3 mM) inhibited carbachol-stimulated T-InsP accumulation by 50%, but resulted in an enhancement of NaF-stimulated T-InsP accumulation. By contrast, inhibition of the mobilization of intracellular calcium ion with 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate inhibited NaF stimulation of T-InsP accumulation by more than 50% but inhibited carbachol-stimulated TInsP accumulation to a much lower extent. Enhanced calcium influx and cell depolarization stimulated by high extracellular concentrations of KCl markedly potentiated carbachol, but not NaF, stimulation of T-InsP accumulation. This differential sensitivity to muscarinic antagonists, cell depolarization, and manipulation of intra- and extracellular calcium ion indicates that different mechanisms underly NaF and carbachol stimulation of T-InsP accumulation. However, stimulation of T-InsP accumulation in the presence of carbachol alone, NaF alone, or carbachol plus NaF was inhibited to a similar extent in the presence of the phorbol ester, phorbol 12-myristate13-acetate. Taken together, these observations suggest that NaF and carbachol effects are mediated through distinct mechanisms but share a common target, perhaps a GTP-binding protein and/or phospholipase C, whose activity is known to be influenced by protein kinase C.
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PMID:Differential sensitivity of phosphoinositide metabolism to sodium fluoride and carbachol treatments in PC12 cells. 1991 22


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