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[3H]PtdIns hydrolysis in postmortem human brain membranes is mediated by the G-proteins Gq/11 and phospholipase C-beta.

ABSTRACT: A method utilizing exogenously added [3H]PtdIns incubated with membranes prepared from postmoretem human brain has been shown to provide a means of measuring agonist-induced, guanosine 5'-O-(thiotriphosphate) (GTP[S])-dependent hydrolysis of [3H]PtdIns, thus allowing investigations of the activity of the phosphoinositide second-messenger system in accessible human brain tissue. Agonists inducing [3H]PtdIns hydrolysis include carbachol, trans-1-aminocyclopentyl-1,3-dicarboxylate (ACPD; a glutamatergic metabotropic receptor agonist), serotonin and ATP, with the latter two agonists producing the largest responses. In addition to ATP, [3H]PtdIns hydrolysis was induced by ADP and by 2-methylthio-ATP, indicating that P2-purinergic receptors mediate this process. Subtype-selective antibodies we used to identify Gq/11 and phospholipase C-beta as the G-protein and phospholipase C subtypes that mediated GTP[S]-induced and agonist-induced [3H]PtdIns hydrolysis. These results demonstrate that this method reveals that agonist-induced, GTP[S]-dependent [3H]PtdIns hydrolysis is retained in postmortem human brain membranes with properties similar to rat brain. This method should allow studies of the modulation of phosphoinositide hydrolysis in human brain and investigations of potential alterations in postmortem brain from subjects with neurological and psychiatric diseases.

SUBMITTER: Jope RS

PROVIDER: S-EPMC1137541 | biostudies-other | 1994 Dec

REPOSITORIES: biostudies-other

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Project description:Lipid dysregulations have been critically implicated in Alzheimer's disease (AD) pathology. Chemical analysis of amyloid-β (Aβ) plaque pathology in transgenic AD mouse models has demonstrated alterations in the microenvironment in the direct proximity of Aβ plaque pathology. In mouse studies, differences in lipid patterns linked to structural polymorphism among Aβ pathology, such as diffuse, immature, and mature fibrillary aggregates, have also been reported. To date, no comprehensive analysis of neuronal lipid microenvironment changes in human AD tissue has been performed. Here, for the first time, we leverage matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) through a high-speed and spatial resolution commercial time-of-light instrument, as well as a high-mass-resolution in-house-developed orbitrap system to characterize the lipid microenvironment in postmortem human brain tissue from AD patients carrying Presenilin 1 mutations (PSEN1) that lead to familial forms of AD (fAD). Interrogation of the spatially resolved MSI data on a single Aβ plaque allowed us to verify nearly 40 sphingolipid and phospholipid species from diverse subclasses being enriched and depleted, in relation to the Aβ deposits. This included monosialo-gangliosides (GM), ceramide monohexosides (HexCer), ceramide-1-phosphates (CerP), ceramide phosphoethanolamine conjugates (PE-Cer), sulfatides (ST), as well as phosphatidylinositols (PI), phosphatidylethanolamines (PE), and phosphatidic acid (PA) species (including Lyso-forms). Indeed, many of the sphingolipid species overlap with the species previously seen in transgenic AD mouse models. Interestingly, in comparison to the animal studies, we observed an increased level of localization of PE and PI species containing arachidonic acid (AA). These findings are highly relevant, demonstrating for the first time Aβ plaque pathology-related alteration in the lipid microenvironment in humans. They provide a basis for the development of potential lipid biomarkers for AD characterization and insight into human-specific molecular pathway alterations.

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[3H]PtdIns hydrolysis in postmortem human brain membranes is mediated by the G-proteins Gq/11 and phospholipase C-beta.

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