The central theme of this Program is to develop animal models both for the understanding of the biology of Alzheimer~s Disease (AD) and for the development of gene therapy of AD. Project 4 focuses on the roles of neurotrophins and acetylcholine within central cholinergic systems in learning and memory. One approach central to this project is to either over-express or abolish expression of the genes encoding nerve growth factor (NGF) or acetylcholine transferase (ChAT), the key enzyme in acetylcholine synthesis, only in specific cholinergic pathways of the adult basal forebrain of the mouse, while expression of these genes is unaffected in the periphery and the remainder of the brain. Over- expression will be achieved by using a tetracycline inducible system. Abolishment of expression will be achieved by employed a conditional mutation system, which is derived from the Cre-loxP recombination system of P1 phage. Recombinase Cre is a 38 kd protein that is able to recognize two recombination sites (termed loxP sites) and catalyze the excision of DNA sequences between two loxP sites. Our preliminary results show that when introduced into hippocampus in an adenoviral vector via stereotaxic injection, the Cre recombinase is indeed able to mediate recombination of a beta-galactosidase target gene construct containing two 1oxP sites. Hence, the strategy is to introduce two loxP sites into the NGF or ChAT gene followed by introduction of the Cre recombinase in specific cholinergic nuclei or target areas of cholinergic neurons. Consequences of these genetic manipulations will be investigated by both neuroanatomical analysis and behavioral tests.
The specific aims are: 1. To generate and characterize mice carrying a lost-or-gains-of-function of the NGF or ChAT gene in adult basal forebrain cholinergic pathways. 2. To determine the specificity of the functional (behavioral) consequences of a loss-or gain-of-function of NGF or ChAT in different basal forebrain cholinergic pathways in young and aged mice. 3. To determine the effects of gene therapy in these models by either ex vivo or in vivo gene delivery methods.
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