Master projekt ved Neurobiologisk Forskningsenhed, Rigshospitalet

Project title: Expression and function of a unique human-specific duplicated alpha7 nicotinic receptor gene in human brain-derived primary microglia cells

Background: Deficits in cholinergic neurotransmission is associated with impaired cognitive function in schizophrenia and Alzheimer’s disease, As a consequence, nicotinic acetylcholine receptors (nAChRs) are a major focus for human genetic, molecular and pharmacological studies. Genetic variations in the CHRNA7 gene are linked to schizophrenia, with the alpha7 nAChR subtype (encoded by the CHRNA7 gene) presenting a promising target for ameliorating cognitive deficits in schizophrenia. Genetic studies indicate that both the classical CHRNA7 gene as well as a partially duplicated alpha7 nAChR gene (CHRFAM7A) is linked to schizophrenia. The CHRFAM7A gene encodes a truncated duplicated variant of the alpha7 nAChR subunit, with the corresponding protein termed dup-alpha7. Dup-alpha7 may act as a dominant-negative modulator of alpha7 nAChR function, which could potentially influence the pharmacodynamics of alpha7 nAChR activators and thus the therapeutic effect in cognitive disease states. It is major importance that this duplicated gene is unique to humans, thus not found in e.g. rodents and non-human primates, which makes it conceivable that CHRFAM7A/dup-alpha7 plays an important role in both normal and pathological function in the human brain, in particular in relation to schizophrenia. Interestingly, the CHRFAM7A gene is found abundant in peripheral immune cells, where it may potentially interfere with immunomodulatory properties of alpha-7 nAChR activators. Because central anti-inflammatory properties of alpha-7 nAChR activators are proposed to contribute to their therapeutic effects in cognitive diseases, it is important to determine the relevance of CHRFAM7A expression and function in the context of inflammatory activity in principal immune cells of the human brain, i.e. microglia.

Project aim: The master project aims to assess the expression and functional implications of CHRNA7/CHRFAM7A genes in the human brain cells. To this end, human microglia isolated from brain cortical lobectomy samples obtained from human donors undergoing resection surgery will be employed. (1) Cultured primary microglia will be assessed for expression dynamics of CHRNA7/CHRFAM7A, as well as other nAChR subunits, in the context of different proinflammatory and nAChR agonist stimuli, respectively; (2) To assess the contribution of microgliaderived CHRNA7/CHRFAM7A expression, the mRNA expression profile will be compared to astrocyte and mixed neuronal/glia cell preparations derived from the same human brain sample; (3) Cellular phenotyping of CHRNA7/CHRFAM7A expression will also be applied to human brain
cells isolated from the brain specimens; (4) Finally, alpha-7 nAChR activators will be tested for functional effects (anti-inflammatory properties, intracellular signaling) in cultured human primary microglia cells.

Research disciplines: Cell cultures, pharmacology, molecular biology, neuroinflammation.

Methods: Quantitative PCR (qPCR), Western blotting, ELISA, immunocytochemistry.

Project start: Autumn/Winter 2015, duration: 1 year.

Research lab: Neurobiology Research Unit (NRU), Juliane Maries Vej 28, 3rd floor, Copenhagen University Hospital Rigshospitalet, Building 6931, DK-2100 Copenhagen, Denmark.

Contact: Henrik B. Hansen, PhD, Senior Scientist (henrik.hansen@nru.dk; tlf. 35456711).