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:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The two forms of pituitary adenylate cyclase-activating polypeptide, PACAP27,and PACAP38, are novel members of the vasoactive intestinal peptide (VIP)/secretin/glucagon family of peptides. PACAP receptors that are positively coupled to adenylate cyclase and phospholipase C have been recently identified. We examined the expression of PACAP receptors in the rat cortex, hippocampus, cerebellum and hypothalamus during postnatal development. Functional studies revealed PACAP stimulation of cAMP formation in all the brain areas examined and [3H]inositol monophosphate ([3H]InsP) accumulation only in the cerebellum and hypothalamus. Throughout development, the efficacy or PACAP in stimulating cAMP formation slightly increased in the cortex and hypothalamus and decreased in the hippocampus and cerebellum; PACAP stimulation of [3H]InsP formation decreased in the cerebellum and remained steady in the hypothalamus. The effects of PACAP27 and PACAP38 on cAMP levels and inositol phospholipid hydrolysis were dose-dependent between 1 and 100 nM. In the same brain areas, treatment with VIP increased cAMP formation at doses greater than 100nM and failed to affect [3H]InsP content, thus suggesting the existence of type-1 PACAP receptors. The reverse transcription polymerase chain reaction (RT-PCR) was used to analyse the mRNA expression of type-1 PACAP receptor splice variants. PACAP receptor gene expression in the central nervous system was regulated in a developmental- and tissue-specific manner. The PACAP-R transcript was detected in all the brain areas examined whereas PACAP-R-hop mRNA ocurred only in the cerebellum and hypothalamus. The different expression profiles and functional properties of PACAP receptors in the developing rat brain suggest an involvement of PACAP in histogenesis, maturation and neurotransmission.
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PMID:Tissue-specific and developmental expression of pituitary adenylate cyclase-activating polypeptide (PACAP) receptors in rat brain. 871 2

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) are hypothalamic factors that play roles in the regulation of anterior pituitary cell activity. PACAP exists in 2 forms physiologically, a 38 amino acid form (PACAP38) and a form possessing the N-terminal 27 amino acids of PACAP38 (PACAP27). We have previously shown that PACAP38 stimulates an increase in [Ca2+]i in rat gonadotrophs. In an attempt to identify the PACAP receptor type underlying this effect, we compared the potency of PACAP38, PACAP27 and VIP to stimulate Ca2+ changes in identified single rat gonadotrophs. All 3 peptides at 100 nM were capable of stimulating high amplitude Ca2+ oscillations, which were also observed in the absence of extracellular Ca2+. The order of potency of these peptides was PACAP38 > PACAP27 > VIP, and a potent antagonist of the PACAP/VIP type II binding site ([4-CI-D-Phe6, Leu17]-VIP) failed to block these responses, suggesting that these effects are mediated through a PACAP/VIP type 1 receptor (PVR1). The Ca2+ responses to PACAP38 and VIP were unaffected by overnight treatment of the cells with pertussis toxin (PTX; 250 ng/ml) indicating that these responses are mediated by a PTX-insensitive G-protein. Finally, the Ca2+ responses stimulated by PACAP38 and VIP were blocked by the phospholipase C-beta blocker U73122 (5 microM). In summary, PACAP stimulates Ca2+ oscillations in rat gonadotrophs through the activation of the PVR1 linked to a PTX-insensitive G-protein and the activation of phospholipase C-beta. VIP can stimulate the same pathway in rat gonadotrophs, although it is at least 100 fold less potent than PACAP38.
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PMID:PACAP and VIP stimulate Ca2+ oscillations in rat gonadotrophs through the PACAP/VIP type 1 receptor (PVR1) linked to a pertussis toxin-insensitive G-protein and the activation of phospholipase C-beta. 873 36

Pituitary adenylate cyclase-activating polypeptide (PACAP)i a potent stimulant of catecholamine secretion, increased catecholamine production in cultured porcine adrenal medullary chromaffin cells. PACAP induced dose-and time-dependent increases in mRNAs for the catecholamine synthesizing enzymes, tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH), with maximal 6- and 4-fold increases occurring at 8-16 h, respectively. The half-maximally and maximally effective PACAP concentrations for stimulation of TH and DBH gene expression were 0.5 and 3 nM, respectively. The TH protein level also showed an increase over the unstimulated basal level at 16-24 h in PACAP-stimulate cells. We previously demonstrated that PACAP activates both phospholipase C and adenylate cyclase in adrenal medullary cells. Addition of forskolin alone induced increases in mRNA expression of both TH and DBH. The phosphodiesterase inhibitor 3- isobutyl-1-methylxanthine potentiated the induction of TH and DBH mRNAs by PACAP. Addition of the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) also caused increases in TH and DBH mRNA levels. In protein kinase C-downregulated cells pretreated with PMA for 24 h, the stimulatory effect of PACAP on TH and DBH gene expression was diminished. These results suggest that cAMP and protein kinase C mediate the PACAP-induced TH and DBH gene expression. Removal of extracellular Ca2+ with EGTA enhanced the PACAP-induced increases in both cellular cAMP and mRNA levels of TH and DBH, suggesting that Ca2+ has an inhibitory effect on the induction of TH and DBH mRNAs. In conclusion, the present study indicates that PACAP coordinately upregulates the gene expression of both TH and DBH by activating the cAMP and protein kinase C signaling pathways, leading to simulation of cate-cholamine synthesis, while Ca2+ negatively regulates TH and DBH gene expression in porcine adrenal medullary cells.
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PMID:Pituitary adenylate cyclase-activating polypeptide induces gene expression of the catecholamine synthesizing enzymes, tyrosine hydroxylase and dopamine beta hydroxylase, through 3',5'-cyclic adenosine monophosphate- and protein kinase C-dependent mechanisms in cultured porcine adrenal medullary chromaffin cells. 877 59

Platelet activating factor (PAF) is a potent phospholipid mediator which elicits a diverse array of biological actions by interacting with G protein-coupled PAF receptors (PAFR). Binding of PAF to PAFRs leads to activation of G protein(s) that stimulate phosphoinositide phospholipase C and subsequent intracellular signaling responses. To identify the potential role of intracellular domains of the rat PAFR (rPAFR) in signaling, we examined effects of transfecting minigenes encompassing rPAFR intracellular domains 1 (1i), 2 (2i), and 3 (3i) on inositol phosphate (IP) production mediated by the co-transfected rPAFR cDNA. Although transfection of the rPAFR1i and rPAFR2i minigenes had no effects on PAF-stimulated signaling, transfection of the rPAFR3i minigene inhibited PAF-stimulated IP production by approximately 50% compared to controls. The rPAFR3i domain did not inhibit IP production mediated by the multifunctional rat pituitary adenylate cyclase-activating polypeptide receptor (rPACAPR), demonstrating the specificity of the competition by the rPAFR3i domain. In further experiments, the rPAFR3i domain was engineered onto the homologous domain of a monofunctional transmembrane variant of the rPACAPR (rPACAPR2) that activates only adenylyl cyclase. The rPACAPR2/rPAFR3i chimera responded to PACAP with increases in IP production which were attenuated nearly completely in cells cotransfected with the rPAFR3i domain. In contrast, PACAP had no effects on IP production in a receptor chimera expressing a mutated form of the rPAFR3i domain (rPACAPR2/rPAFR3imut). These results demonstrate the ability of the rPAFR3i domain to confer a phospholipase C-signaling phenotype to a receptor deficient in this activity and show that this activity is specific for the engineered rPAFR3i domain. These results suggest that the third intracellular loop of the rPAFR is a primary determinant in its coupling to phosphoinositide phospholipase C-activating G proteins, providing the first insight into the molecular basis of interaction of PAFRs with signal-transducing G proteins.
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PMID:The third intracellular domain of the platelet-activating factor receptor is a critical determinant in receptor coupling to phosphoinositide phospholipase C-activating G proteins. Studies using intracellular domain minigenes and receptor chimeras. 879 8

Glutamatergic neurotransmission is associated with release of arachidonic acid (AA) from membrane phospholipids of both neurons and astrocytes. Since free AA has been shown to enhance glutamate-mediated synaptic transmission, it can be postulated that glutamate release and AA formation constitute a positive feed-back mechanism for sustained excitatory neurotransmission. In the present study, we examined whether the glutamate-evoked release of AA could be modulated by peptides. Using mouse cortical neurons in primary cultures, we show that the release of AA evoked by glutamate is potentiated by vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide (PACAP). This effect is mediated through the activation of PACAP I receptors. However, several arguments show that this potentiating mechanism does not involve the cAMP/PKA pathway. 1) Increasing intracellular cAMP by either cholera toxin, forskolin, or 8-Br-cAMP treatments does not affect the glutamate-evoked release of AA; 2) potentiation of the glutamate response by PACAP is not prevented by the PKA inhibitor 8-Br-Rp-cAMPS. Also, an involvement of the phospholipase C protein kinase C pathways is unlikely since inhibitors of both phospholipase C (i.e. U-73122) and protein kinase C (i.e. Ro 31-8220) do not affect the potentiation of the glutamate response by PACAP. These observations indicate an effect mediated by PACAP I receptors, which does not involve the second messenger pathways classically associated with activation of this type of receptors. Furthermore, results indicate that this potentiating mechanism mediated by PACAP I receptor acts at a level downstream of the glutamate receptor-mediated calcium influx.
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PMID:Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) potentiate the glutamate-evoked release of arachidonic acid from mouse cortical neurons. Evidence for a cAMP-independent mechanism. 879 93

Pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide belonging to the VIP/secretin/glucagon family, is present in the hypothalamus, anterior pituitary, and adrenal gland where it regulates hormone release, in the GI tract where it modulates motility, and in human tumoral cell lines where it shows a growth-promoting effect. It is now appreciated that alternative splicing of two exons of the rat PACAP-R gene generate four major rPACAP-R splice variants that are differentially expressed in tissues and variably coupled to intracellular second messengers. Because of the potential implications of these findings in human physiology, we cloned the hPACAP-R gene. Similar to the rat, two exons (SV-1 and SV-2) are alternatively spliced to account for four major hPACAP-R receptor splice variants. These splice variants (hPACAP-R-null, hPACAP-R-SV1, hPACAP-R-SV2, hPACAP-R-SV-3) were cloned from a human frontal cortex cDNA library, stably transfected in NIH/ 3T3 cells and each characterized for ligand affinity, stimulation of adenylate cyclase (AC) and phospholipase C (PLC), and ligand-induced expression of the proto-oncogenes, c-fos, and c-myc. Stably transfected NIH/3T3 cells expressing similar numbers of receptors of the four splice variants showed nearly identical responses for ligand affinity and potency for P-38- and P-27-stimulated increases in cAMP and total inositol phosphates. However, each receptor splice variant differed in their ligand-stimulated efficacy for total inositol phosphate stimulation. The hPACAP-R-SV2 showed the greatest efficacy for stimulating phospholipase C that was approximately seven-fold greater than the hPACAP-R-SV1, twofold greater than the hPACAP-R-Null, and 1.5-fold greater than the hPACAP-R-SV-3 splice variants. To determine whether the splice variants also differ in their ability to stimulate immediate early gene expression, c-fos and c-myc transcripts were assayed by Northern blot and quantified by densitometry. PACAP-38 increased c-fos and c-myc expression for all four of the receptor splice variants that paralleled the efficacy for PLC stimulation, with the the SV-2 splice variant showing the greatest stimulation. These results show that the hPACAP-R-SV2 exhibits enhanced efficacy for coupling to both PLC and activation of the protooncogenes, c-fos and c-myc suggesting a novel and potentially important mechanism for differentially activating signal transduction pathways that influence cellular growth and differentiation.
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PMID:Differential signaling and immediate-early gene activation by four splice variants of the human pituitary adenylate cyclase-activating polypeptide receptor (hPACAP-R). 899 93

The current studies have implicated a prominent role for PACAP peptides in modulating the physiological function of cells derived from the sympathoadrenal lineage. Compared to VIP, both PACAP-27 and PACAP-38 demonstrated potent, efficacious, and sustained stimulatory effects on sympathetic neuronal NPY and catecholamine production. The differential effects of PACAP peptides on SCG NPY and catecholamine content and secretion coincided with previous studies that activated directly the sympathetic intracellular cyclic AMP-protein kinase A signaling pathway. These effects appear to be mediated primarily by PACAP1 receptor splice variants coupled to both adenylyl cyclase and phospholipase C in SCG neurons. The actions of PACAP peptides in the SCG shared many parallels with adrenal medullary chromaffin cells, suggesting diverse roles for the PACAP peptidergic system in sympathoadrenal cell development and function. Rather than solutions, these results pose additional questions for the future. What are the endogenous sources of PACAP that regulate sympathetic and adrenal function? Do PACAP peptides, like VIP, have dual roles and also act as sympathetic postganglionic neuromodulators? Are VIP/PACAP receptors expressed during SCG development? What regulates sympathetic PACAP1 receptor isoform expression and how are they differentially coupled to neuronal intracellular signaling cascades? What defines the tissue-specific responses to PACAP-27 and PACAP-38? While many of these questions are not easily approached, future studies of these issues will certainly illuminate the function of PACAP and PACAP receptors in the nervous and endocrine systems.
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PMID:Pituitary adenylate cyclase-activating polypeptides, PACAP-38 and PACAP-27, regulation of sympathetic neuron catecholamine, and neuropeptide Y expression through activation of type I PACAP/VIP receptor isoforms. 899 4

The purpose of this study was to investigate the mechanisms of action of pituitary adenylate cyclase-activating polypeptide (PACAP) in stimulating aldosterone production in two different models: bovine adrenal zona glomerulosa (ZG) cells in primary culture and the human adrenocortical carcinoma cell line H295R. PACAP binds to two major groups of receptors: type I, which prefers PACAP38 and PACAP27 over vasoactive intestinal peptide (VIP); and type II, which has approximately equal affinity for PACAP38, PACAP27, and VIP. The type I subclass comprises multiple splice variants that can be distinguished by their specificity to PACAP38 and PACAP27 in their activation of adenylate cyclase and phospholipase C. Type II PACAP/ VIP receptors couple only to AC. In bovine ZG cells, PACAP38 and PACAP27 stimulated aldosterone production in a dose-dependent manner, whereas VIP was ineffective. In H295R cells, PACAP38, PACAP27, and VIP dose-dependently stimulated aldosterone production with roughly the same ED50. In bovine ZG cells, PACAP38 and PACAP27 stimulated cAMP production with similar efficacy, whereas VIP had no effect. In H295R cells, all three peptides stimulated cAMP accumulation. PACAP38 and PACAP27 also activated PLC in bovine ZG cells as they induced an increase in Ins(1,4,5)Ps production. In H295R cells, neither of these peptides was able to stimulate IP turnover. These results indicate that PACAP stimulation of aldosterone production is mediated by the PVR1s or the PVR1hop splice variants of the type I PACAP-specific receptor subtype in bovine ZG cells, whereas only type II PACAP/VIP receptors seemed to occur in the human H295R cell line. In addition, PACAP-stimulated aldosterone production was inhibited by atrial natriuretic peptide in bovine and human adrenocortical cells, however not by the same mechanism. This further supports species-specific and/or cell type-specific signaling pathways for PACAP in the regulation of aldosterone production.
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PMID:Comparative effect of pituitary adenylate cyclase-activating polypeptide on aldosterone secretion in normal bovine and human tumorous adrenal cells. 900 87

To investigate the regulation of free cytosolic calcium concentration ([Ca2+]i) by the adenosine 3',5'-cyclic monophosphate (cAMP) signaling system in clonal gonadotrophs, microfluorimetric recordings were made in single indo 1-loaded alpha T3-1 cells. Forskolin, 8-bromoadenosine 3',5'-cyclic monophosphate, or a low concentration (100 pM) of the hypothalamic factor pituitary adenylate cyclase-activating polypeptide (PACAP) stimulated Ca2+ step responses or repetitive Ca2+ transients, which were blocked by the removal of extracellular Ca2+ by the dihydropyridine (DHP) (+)PN 200-110 or by preincubation with the protein kinase A (PKA) antagonist H-89 (10 microM). Thus activation of the cAMP/PKA system in alpha T3-1 gonadotrophs stimulates Ca2+ influx through DHP-sensitive (L-type) Ca2+ channels. In contrast, high PACAP concentrations (100 nM) stimulated biphasic Ca2+ spike-plateau responses. The Ca2+ spike was independent of extracellular Ca2+, and similar responses were observed by microperfusion of individual cells with D-myo-inositol 1,4,5-trisphosphate, suggesting the involvement of the phospholipase C (PLC) signaling pathway. The Ca2+ plateau depended on Ca2+ influx, was blocked by (+)PN 200-110, but was only partially blocked by H-89 pretreatment. In conclusion, PACAP stimulates [Ca2+]i increases in alpha T3-1 gonadotrophs through both the PLC and adenylate cyclase signaling pathways. Furthermore, this is the first clear demonstration that the cAMP/PKA system can mediate changes in [Ca2+]i in gonadotroph-like cells.
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PMID:Stimulation of Ca2+ influx in alpha T3-1 gonadotrophs via the cAMP/PKA signaling system. 937 69

Continuous exposure of cells to agonists develops a process that determines the extent to which the cells eventually respond to further stimuli. Here we used CATH.a cells (a catecholaminergic neuron-like cell line), which express pituitary adenylate cyclase-activating polypeptide (PACAP) receptors linked to both adenylyl cyclase and phospholipase C-beta pathways, to investigate the influence of prolonged hormonal treatment on dual signaling and gene transcription. Prolonged incubation of cells with PACAP failed to down-regulate the density and affinity of membrane binding sites and caused opposite changes in messenger systems: PACAP-stimulated cyclic AMP accumulation was attenuated in a time- and dose-dependent fashion (t(1/2) = 6.7 h and IC50 = 0.1 nM), whereas phosphoinositide turnover was overstimulated. Both effects were insensitive to pertussis toxin, whereas the drop in cyclic AMP concentration was also unchanged in the presence of 3-isobutyl-1-methylxanthine, indicating that neither Gi-like proteins nor cyclic nucleotide phosphodiesterases play a critical role in these processes. Blockade of protein synthesis with cycloheximide, as well as inhibition by H89 of cyclic AMP-dependent protein kinase (but not by bisindolylmaleimide of protein kinase C) antagonized the influences exerted by PACAP on adenylyl cyclase activity and inositol phosphate formation. Transcription of the chimeric GAL4-CREB construct, transiently transfected into CATH.a cells, was stimulated by PACAP, and this effect was potentiated as a result of chronic PACAP treatment. The results of the present investigation provide new insight into the possible differential regulation and cross-talks of transduction signals of receptors linked to multiplex signaling. They demonstrate that prolonged exposure of CATH.a cells to PACAP results in the desensitization of the cyclic AMP pathway and superinduction of the inositol phosphate signal, through protein neosynthesis and cyclic AMP-dependent protein kinase activation. At the same time, they show that desensitization of cyclic AMP signaling not only fails to hamper, but actually amplifies PACAP-stimulated CREB-regulated transcription.
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PMID:Continuous activation of pituitary adenylate cyclase-activating polypeptide receptors elicits antipodal effects on cyclic AMP and inositol phospholipid signaling pathways in CATH.a cells: role of protein synthesis and protein kinases. 952 59


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