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A quantitative phospho-proteomic study on rat hippocampal synaptoneurosomes following a single 19-nortestosterone administration.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences, Biological Research on Drug Dependence.
GE Healthcare, Bio-Sciences AB, Uppsala, Sweden.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences, Biological Research on Drug Dependence.
Department of Medical Biochemistry and Biophysics, Division of Molecular Biometry, Karolinska Institute, Stockholm, Sweden.
(English)Manuscript (preprint) (Other academic)
Abstract [en]

In the past, several studies have been conducted investigating the impact of androgens on the activation of known signaling pathways in cultured primary hippocampal cells and C6 cells. We have recently reported two studies showing altered in vivo phosphorylation of key components in cellular signaling in rat hippocampal synaptoneurosomes after administration of anabolic androgenic steroids. In this paper we report a quantitative liquid chromatography mass spectrometry-based screening for androgen-induced alterations of phosphoproteins in rat hippocampal synaptoneurosomes following a single 19-nortestosterone administration. We identified over 80 phosphorylated peptides with functions associated with the cytoskeleton and transmembrane signaling. DeCyder™ MS based quantitative comparison of the relative phosphopeptide abundances between the two groups of treatment was computed for the majority of the phosphopeptides.

National Category
Neurosciences
Identifiers
URN: urn:nbn:se:uu:diva-120302OAI: oai:DiVA.org:uu-120302DiVA: diva2:303093
Available from: 2010-03-11 Created: 2010-03-11 Last updated: 2010-03-11
In thesis
1. The Impact of Drugs of Abuse on the Neuroproteome: Application of Immunoblotting and LC-MS-based Proteomics
Open this publication in new window or tab >>The Impact of Drugs of Abuse on the Neuroproteome: Application of Immunoblotting and LC-MS-based Proteomics
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Qualitative and quantitative analysis of proteins can be based either on immunological or on mass spectrometry-utilizing methods. In this thesis, different approaches of protein analysis were applied to elucidate the impact of drugs of abuse on the rat brain.

Androgens have been reported to affect mood, anxiety and memory function. The hippocampus plays an important role in memory formation, and studies have reported androgen-induced alteration in hippocampal dendritic spine density. The process of memory and learning requires synaptic plasticity, which depends on key components of signaling pathways, e.g. the ERK and the NMDA receptor. Often, misuse of anabolic androgenic steroids (AAS) leads to administration of supratherapeutical doses with partly unknown impact on the brain.

Rapid and subchronic effects of high AAS doses on proteins of a synapse-enriched rat hippocampal preparation (synaptoneurosomes) were investigated using the immunoblot technique and mass spectrometry. A single high dose of the AAS nandrolone decanoate was found to increase the in vivo phosphorylation of both NMDA receptor subunits and ERKs 24 h after administration. Subchronic treatment with multiple doses showed no effect. A subsequent study focused on rapid effects of the free nandrolone. The phosphorylation of the NMDA receptor subunit GluN2B and ERK1 was found diminished after 2 h and restored after 6 h. This suggests a biphasic impact of nandrolone on signaling components. Moreover, this study aimed to elucidate whether the effects were mediated via the androgen receptor (AR). The AR antagonist flutamide abolished AAS-induced effects. Determination of the CaMKIIα and ephrinB2 proteins, being a potential link of the analogous phosphorylation of NMDA receptor and ERK, could be excluded. Proteins involved in synaptogenesis were not found to be altered. A mass spectrometry-based study was conducted to screen for phosphoproteins and their potential alteration 2 h after a single dose of nandrolone. This analysis identified several proteins with known and not yet described phosphorylation sites. A differential analysis of the identified phosphoproteins was performed.

In a fourth study, quantitative MS-based peptidomics were applied to investigate endogenous neuropeptides in the mesolimbic system during morphine withdrawal. Moreover, strain-dependence was investigated. In accordance to earlier reports, comparable regulation was observed of proenkephalin-, prodynorphin- and preprotachykinin-derived neuropeptides levels.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2010. 67 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 121
Identifiers
urn:nbn:se:uu:diva-120303 (URN)978-91-554-7749-3 (ISBN)
Public defence
2010-04-28, B41, BMC, Husargatan 3, Uppsala, Sweden, 09:15 (English)
Opponent
Supervisors
Available from: 2010-04-06 Created: 2010-03-11 Last updated: 2011-05-05Bibliographically approved

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