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C O'Neill

Publications and source records attributed to C O'Neill.

At least 19 recordsLinked to original sources
PubMed1992

Preservation of Gi-protein inhibited adenylyl cyclase activity in the brains of patients with Alzheimer's disease.

The coupling of inhibitory guanine nucleotide binding (Gi) proteins to the adenylyl cyclase signal transduction complex was compared in 4 brain regions from a series of Alzheimer's disease and matched control subjects by measuring the inhibition of membrane enzyme activities in response to guanosine 5'-[beta gamma-imido]diphosphate (Gpp[NH]p) and aluminium fluoride (AlF4-). Basal adenylyl cyclase activities were significantly lower in preparations of angular gyrus and frontal and temporal cortices, but not cerebellum, from the Alzheimer's disease cases compared to controls. Gpp[NH]p and AlF4- gave significant inhibitions of adenylyl cyclase activity in all brain regions. The magnitude of these inhibitions, when corrected for altered basal activities, were similar for the Alzheimer's disease and control cases. These results indicate that there is no impairment of Gi-protein mediated inhibition of adenylyl cyclase activity in Alzheimer's disease brain.

Adenylyl Cyclases↗
PubMed1992

Embryo-derived platelet activating factor.

Platelet-activating factor (PAF) is secreted by the preimplantation embryo of a number of species. The role of this secretion is yet to be fully elucidated. Evidence to date indicates that it has an important function as an autocrine stimulant of embryonic metabolism, growth and viability. Production of PAF by embryos appears to be severely compromised in vitro. This may be a major cause of reduced embryo viability following embryo culture and it may also help to explain the controversy in the literature regarding the production of PAF by embryos. PAF also alters several aspects of maternal physiology during early pregnancy including endometrial prostaglandin secretion. The significance of these changes remains to be defined.

Animals↗
PubMed1992

Metabolic and developmental responses of preimplantation embryos to platelet activating factor (PAF).

Platelet activating factor (PAF) is an ether phospholipid produced by preimplantation embryos of a number of species. Production of PAF by embryos has been measured by detecting thrombocytopenia in a splenectomized mouse bioassay, platelet aggregation bioassays in vitro and a specific radioimmunoassay. Production is highly variable and is adversely affected by culture in vitro. It has, however, been correlated with morphology, development rates in vitro and the pregnancy potential of embryos following transfer. Investigations using PAF-antagonists have established an essential role for PAF in early pregnancy. Together with studies that have shown PAF to have direct effects on embryonic metabolism during culture in vitro, these observations suggest that PAF acts as an embryonic autocoid. Hence, a major site of action for embryo-derived PAF in vivo is the embryo itself. Supplementation of embryo culture media with PAF had no effect on the rate of development in vitro of 2-cell mouse embryos through to the blastocyst stage. However, PAF increased cell numbers of blastocysts cultured from the 2-cell stage and the mitotic index of embryos at both the 8-cell and blastocyst stages. Supplementation of culture media with PAF has also been shown to increase the implantation potential of both mouse and human embryos cultured in vitro. In the mouse, the effect of PAF in enhancing implantation rates was most evident when the developmental potential of untreated embryos was suboptimal. These observations suggest that the production of embryo-derived PAF is one limiting factor in maintaining the viability of embryos cultured in vitro.

Animals↗
PubMed1992

Actions of platelet activating factor (PAF) on gametes and embryos: clinical aspects.

Platelet activating factor (PAF) is a phospholipid widespread in body tissues. Previous reviews have discussed its production by preimplantation embryos and the evidence implicating it as an autocrine mediator in aspects of gamete and embryo physiology. Human spermatozoa contain variable amounts of PAF, the amount contained depending on the source and method of preparation of the sperm. Incubation of human sperm with PAF tends to increase their forward velocity, especially in samples with slow motility. PAF treatment causes an increase in the proportion of acrosome-reacted sperm and in their ability to penetrate both zona-free hamster ova and cervical mucus. PAF has been found in human follicular fluid at ovulation. A role for PAF in ovulation has been suggested, because PAF antagonists reduce the rate of ovulation in rats. In some studies, modest improvements to mouse in vitro fertilization (IVF) rates have been achieved with PAF supplementation of media under specific conditions. Furthermore, in the rabbit and mouse, PAF antagonists have been reported to inhibit fertilization in vivo and in vitro respectively. However, addition of PAF to human IVF medium, but only at the time of insemination and fertilization, had no effect on either fertilization or pregnancy rates. Sensitive bio- and immuno-assays have shown that PAF is secreted by human embryos into their fluid milieu. PAF secretion by these zygotes during culture, although markedly variable, has been correlated with the achievement of pregnancy and pregnancy outcome. Although the secretion of PAF by the mouse embryo decreases during culture in vitro, exogenous PAF enhances embryo viability during culture. Similarly, culture of human zygotes in PAF-supplemented medium prior to embryo transfer significantly increases the chance of achieving pregnancy. Both the implantation and live-birth rates are increased in human IVF by addition of PAF to the medium.

Embryo, Mammalian↗
PubMed1992

Structural heterogeneity of platelet-activating factor produced by murine preimplantation embryos.

Platelet-activating factor (PAF) is 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine with the alkyl moiety predominantly a mixture of saturated hexadecyl and octadecyl chains (C16:0 and C18:0 PAF, respectively). Previously, a PAF bioassay was compared with a radioimmunoassay for PAF (NEN Du Pont). Both assays were sensitive and quantitative, but the correlation between PAF measured by the bioassay compared to the radioimmunoassay was poor for murine embryo-derived PAF (r = 0.773, n = 88), while being completely adequate (r = 0.961) for a PAF standard which was an equimolar mixture of C16:0 and C18:0 PAF (C16:0/C18:0 PAF). This study compared a larger sample size of murine embryo-derived PAF (n = 154) and found that the poor correlation between the two assays persisted (r = 0.791). When dose-response curves were generated with C16:0, C16:0/C18:0 and C18:0 PAF (over a concentration range of 0.3-30 ng/ml), the concentrations which gave a 50% response were equivalent in the bioassay (i.e. 6 ng/ml), but differed in the radioimmunoassay (i.e. 1.5, 3 and 6 ng/ml, respectively). Following separation of murine embryo-derived PAF (from medium in which 30 two-cell embryos had been cultured for 24 h) into C16:0 and C18:0 PAF by reverse phase high performance liquid chromatography, 9/20 cultures produced 100% C16:0 PAF, 2/20 cultures produced 100% C18:0 PAF and the remaining 9/20 cultures produced varying proportions of both.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Evidence that platelet-activating factor suppresses uterine oxytocin-induced 13,14-dihydro-15-keto-prostaglandin F2 alpha release and phosphatidylinositol hydrolysis in the ewe.

This study was conducted to determine whether platelet-activating factor (PAF) (1) attenuated oxytocin-induced secretion of the prostaglandin (PG) F2 alpha metabolite, PGFM, by the ovine uterus in situ and (2) inhibited the generation of the inositol phosphate secondary messengers by endometrial tissue in response to oxytocin challenge in vitro. Ovariectomized ewes received steroid replacement to mimic the luteal phase. Six ewes received intrauterine injections of 200 micrograms PAF/uterine horn/day on Days 11-15, and 6 ewes were treated with vehicle. All ewes received 1 microgram oxytocin i.v. on Days 13-16. Pretreatment of ewes with PAF significantly suppressed PGFM release in response to oxytocin on Days 14 and 15 (p less than 0.005) compared to vehicle-treated ewes. PAF was not administered on Day 16, and the PGFM response to oxytocin was not different between groups. In a second experiment, ewes were given intrauterine injections of 200 micrograms PAF/uterine horn/day (n = 8) or vehicle (n = 7) on Days 11-15, and all ewes received 1 microgram oxytocin i.v. on Days 13 and 14. On Day 15 the uterus was removed, and the incorporation of 3H-inositol into inositol phosphates was determined in caruncular endometrium. Treatment of ewes with PAF in vivo reduced inositol monophosphate (IP1) generated by oxytocin (10(-6) M) by 56.4%, compared to that in endometrium from vehicle-treated controls, and also inhibited the incorporation of 3H-inositol into glycerophosphoinositol (GPI). If PAF was added to the endometrium during the incubation in vitro, the attenuation of inositol phosphate generation did not occur.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Adenylyl cyclase activity in postmortem human brain: evidence of altered G protein mediation in Alzheimer's disease.

The effects of agonal status, postmortem delay, and age on human brain adenylyl cyclase activity were determined in membrane preparations of frontal cortex from a series of 18 nondemented subjects who had died with no history of neurological or psychiatric disease. Basal and guanosine 5'-O-(3-thiotriphosphate)-, aluminum fluoride-, and forskolin-stimulated enzyme activities were not significantly reduced over an interval from death to postmortem of between 3 and 37 h and were also not significantly different between individuals dying with a long terminal phase of an illness and those dying suddenly. Basal and aluminum fluoride-stimulated enzyme activities showed a negative correlation with increasing age of the individual. In subsequent experiments, basal and guanosine 5'-O-(3-thiotriphosphate)-, aluminum fluoride-, and forskolin-stimulated enzyme activities were compared in five brain regions from a series of eight Alzheimer's disease and seven matched nondemented control subjects. No significant differences were observed between the groups for either basal activity or activities in response to forskolin stimulation of the catalytic subunit of the enzyme. In contrast, enzyme activities in response to stimulation with guanosine 5'-O-(3-thiotriphosphate) and aluminum fluoride were significantly reduced in preparations of neocortex and cerebellum from the Alzheimer's disease cases compared with the nondemented controls. Lower guanosine 5'-O-(3-thiotriphosphate)-, but not aluminum fluoride-, stimulated activity was also observed in preparations of frontal cortex from a group of four disease controls compared with nondemented control values. The disease control group, which contained Parkinson's disease and progressive supranuclear palsy patients, showed increased forskolin-stimulated activity compared with both the nondemented control and the Alzheimer's disease groups. These findings indicate a widespread impairment of G protein-stimulated adenylyl cyclase activity in Alzheimer's disease brain, which occurs in the absence of altered enzyme catalytic activity and which is unlikely to be the result of non-disease-related factors associated with the nature of terminal illness of individuals.

Adenylyl Cyclases↗
PubMed1992

Biosynthesis of platelet-activating factor by two-cell mouse embryos.

Incubation of two-cell mouse embryos with a range of radiolabelled compounds resulted in the incorporation of label into platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) in the culture media. The demonstration that known precursors ([1-14C]hexadecanol, [1-3H]hexadecanol, 1-O-[alkyl-1'2'-3H]lyso-PAF, 1-O-[alkyl-1'2'-3H]acetyl-glycerol and [methyl-3H]choline chloride) were incorporated into PAF showed that embryo-derived PAF biosynthesis occurred via pathways present in other PAF-producing cells. The enzyme responsible for the formation of the ether linkage of the PAF molecule, alkyl-dihydroxyacetone-phosphate synthase, was present in the preimplantation embryo as [1-3H]hexadecanol was incorporated into PAF. Incorporation of label from alkylacetyl-glycerol and choline chloride into lyso-PAF was also observed, suggesting a role for lyso-PAF in the metabolism of embryo-derived PAF. Incubation of embryos with each of three [14C]carbohydrate energy substrates resulted in the incorporation of label into PAF in culture media, indicating that the composition of embryo culture media is important in the synthesis of PAF precursors. Incorporation of label from [2-14C]pyruvate was greatest and is consistent with the suggestion that pyruvate is the major energy source at the two-cell stage of development. L-[U-14C]Lactate was also incorporated into embryo-derived PAF, but the mean amount incorporated relative to the concentration of labelled substrate in the medium was 40 times less. The incorporation of D-[U-14C]glucose into PAF was 2405 times less than that from pyruvate, relative to the concentration in the medium.

Alkyl and Aryl Transferases↗
PubMed1991

Preservation of 5-hydroxytryptamine1A receptor-G protein interactions in the cerebral cortex of patients with Alzheimer's disease.

The coupling of 5-hydroxytryptamine1A (5-HT1A) receptors to guanine nucleotide binding (G) proteins was investigated in membranes prepared from frontal and parietal cortices of control and Alzheimer's disease brains by characterising the effect of guanosine 5'-[beta gamma-imido]diphosphate (Gpp[NH]p) on [3H]8-hydroxy-2-(di-n-propylamino)-tetralin ([3H]8-OH-DPAT) binding parameters. In the absence of guanine nucleotides, [3H]8-OH-DPAT bound to a single high affinity binding site in all membrane types. The number of [3H]8-OH-DPAT binding sites was significantly decreased in the parietal cortex of Alzheimer's disease samples compared with controls, whereas in the frontal cortex the number of binding sites remained unchanged. Gpp[NH]p reduced the [3H]8-OH-DPAT binding affinity and the number of binding sites to the same degree in both regions in control and Alzheimer's disease cases. [3H]8-OH-DPAT binding was inhibited in a concentration dependent manner with an IC50 value of approximately 1 microM in all cases. These results suggest that the 5-HT1A receptor-G protein complex is functionally intact in these regions in Alzheimer's disease brain.

8-Hydroxy-2-(di-n-propylamino)tetralin↗
PubMed1991

Coupling of human brain cerebral cortical alpha 2-adrenoceptors to GTP-binding proteins in Alzheimer's disease.

The coupling of alpha 2-adrenoceptors to guanine nucleotide binding G-proteins was investigated in cerebral cortical membranes from control and Alzheimer's disease brain by characterizing the effects of MnCl2 and Gpp[NH]p on [3H]clonidine binding. The manganese induced elevation of [3H]clonidine binding was apparent in both control and Alzheimer's disease samples and the effect showed no significant difference between the two groups in the frontal cortex. However, the MnCl2 concentration dependence curves for control and Alzheimer's disease samples were significantly different from one another in the temporal cortex, whereas the pattern of stimulation by MnCl2 remained the same. The guanine nucleotide analogue, Gpp[NH]p inhibited [3H]clonidine binding in a concentration-dependent manner, the profiles of inhibition showing no significant differences between control and Alzheimer's disease samples. Analysis of the effect of Gpp[NH]p on [3H]clonidine saturation binding curves showed no significant differences between control and Alzheimer's disease samples in either frontal (Kd = 9.68 +/- 1.38, 9.1 +/- 2.6 nM; Bmax = 40.23 +/- 4.33, 44.3 +/- 9.4 fmol/mg, control and Alzheimer's disease values, respectively), or temporal (Kd = 11.61 +/- 4.04, 5.38 +/- 2.5 nM; Bmax = 52.0 +/- 14.0, 31.07 +/- 8.00 fmol/mg control and Alzheimer's disease values, respectively) cortices.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1991

Assay of a phosphatidylinositol bisphosphate phospholipase C activity in postmortem human brain.

The activity of a phospholipase C which hydrolyses exogenous phosphatidylinositol-4,5-bisphosphate [( 3H]PtdIns(4,5)P2) in membranes prepared from frozen postmortem human brain and rat brain was investigated. Enzyme characteristics were essentially similar in membranes prepared from frozen postmortem brain and fresh or frozen rat brain. The [3H]PtdIns(4,5)P2 solubilization and assay procedure employed resulted in an efficient availability of the substrate for the enzyme. The non-hydrolysable guanosine triphosphate analogue guanosine 5'-[beta gamma-imido]diphosphate (Gpp[NH]p) stimulated hydrolysis rapidly with a half maximum activity of approximately 25 microM. This stimulation was not specific for guanine nucleotides as ATP, imidodiphosphate and pyrophosphate also caused enzyme activation. However these activation effects could be distinguished by the polyanion spermine. The non-hydrolysable guanine dinucleotide analogue guanosine 5'-[beta-thio]diphosphate acted as a partial agonist thereby inhibiting the stimulatory effect of Gpp[NH]p. Gpp[NH]p-stimulated enzyme activity showed a maximum response in the presence of 1 mM deoxycholate and displayed a pH optima in the range 7.0-7.5. PtdIns(4,5)P2 hydrolysis was observed in the absence of added calcium, but hydrolytic cleavage was inhibited in the presence of divalent ion chelators. Magnesium inhibited PtdIns(4,5)P2 hydrolysis in a concentration-dependent manner. Elucidation of these aspects of the phosphatidylinositol cycle in normal human postmortem brain will permit comparative studies in CNS disease states.

Aged↗
PubMed1991

The effect of inhibitors of platelet aggregation on the metabolism of platelet-activating factor (PAF) in washed rabbit platelets.

The rabbit platelet metabolizes platelet-activating factor (PAF) intracellularly. PAF is deacetylated to produce lysoPAF which, in turn, can be acylated to produce 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine (alkylacyl GPC). Some PAF receptor antagonists have been shown to inhibit this metabolic conversion. In the present study we examined whether the PAF receptor antagonists SRI 63-441 and WEB 2086 would inhibit the metabolism of PAF by intact rabbit platelets. In addition, we examined whether iloprost, a stable analogue of prostaglandin I2 (PGI2), and a potent inhibitor of platelet activation induced by a range of agonists, would also inhibit PAF metabolism. We found that SRI 63-441 and WEB 2086 caused an almost complete inhibition of the conversion of PAF to alkylacyl GPC. Iloprost caused up to a 50% inhibition of PAF metabolism compared to antagonist-free controls. Iloprost (and PGI2) is thought to inhibit platelet response by elevation of cAMP, while receptor antagonists act by blocking PAF binding to its receptor. Since iloprost caused partial inhibition of PAF metabolism, the results of this study suggest that inhibition of PAF metabolism does not occur solely due to competitive inhibition of PAF binding to its receptor.

Animals↗
PubMed1991

Platelet-activating factor (PAF) receptor antagonists inhibit arachidonic acid induced platelet aggregation in rabbit whole blood.

The potency of several platelet-activating factor (PAF) receptor antagonists was measured by observing their inhibitory effects against PAF induced platelet aggregation. Their selectivity was assessed by monitoring their effect on platelet aggregation induced by arachidonic acid (AA) and adenosine diphosphate (ADP). The antagonists inhibited platelet aggregation induced at the PAF EC50 (0.023 microM) with the following rank order of potency: WEB 2086 greater than WEB 2170 greater than SRI 64-412 greater than SRI 63-675 greater than BN 52021 greater than kadsurenone greater than SRI 63-441 greater than alprazolam. While the antagonists had no inhibitory effect at the EC50 for ADP (10 microM), they did inhibit platelet aggregation induced at the EC50 for AA (55 microM). However, there was considerable variability in the slope of the inhibitory response and the relative potency of each antagonist against PAF induced platelet aggregation as compared to AA induced platelet aggregation. The antagonist IC50 (microM) against PAF and AA were as follows, with those that showed significantly different (p less than 0.01) slopes indicated by an asterisk: SRI 63-441* (3.8, 15.1); SRI 63-675 (1.4, 36.2); SRI 64-412 (0.5, 10.5); BN 52021* (2.4, 58.9); kadsurenone* (2.8, 28.3); alprazolam* (10, 25); WEB 2086 (0.055, 0.220), and WEB 2170 (0.107, 0.534). Therefore, in rabbit whole blood the antagonists were potent, although not completely selective, inhibitors of PAF induced platelet aggregation. These results suggest that the mode of action of PAF and AA induced platelet aggregation may share some common features. However, since the slope of the inhibitory response against PAF and AA for some antagonists differed, mechanistic differences in their action appear to exist.

Adenosine Diphosphate↗