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  • Central Cholinergic Synapse Formation in Optimized Primary Septal-Hippocampal Co-cultures.

Central Cholinergic Synapse Formation in Optimized Primary Septal-Hippocampal Co-cultures.

Cellular and molecular neurobiology (2020-08-30)
Sarra Djemil, Claire R Ressel, Mai Abdel-Ghani, Amanda K Schneeweis, Daniel T S Pak
ABSTRACT

Septal innervation of basal forebrain cholinergic neurons to the hippocampus is critical for normal learning and memory and is severely degenerated in Alzheimer's disease. To understand the molecular events underlying physiological cholinergic synaptogenesis and remodeling, as well as pathological loss, we developed an optimized primary septal-hippocampal co-culture system. Hippocampal and septal tissue were harvested from embryonic Sprague-Dawley rat brain and cultured together at varying densities, cell ratios, and in the presence of different growth factors. We identified conditions that produced robust septal-hippocampal synapse formation. We used confocal microscopy with primary antibodies and fluorescent ligands to validate that this system was capable of generating developmentally mature cholinergic synapses. Such synapses were comprised of physiological synaptic partners and mimicked the molecular composition of in vivo counterparts. This co-culture system will facilitate the study of the formation, plasticity, and dysfunction of central mammalian cholinergic synapses.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-MAP2 Antibody, Chemicon®, from chicken
Sigma-Aldrich
Trypsin Inhibitor, Defined (1X) Solution, Animal component free, BioReagent, suitable for cell culture
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Anti-Choline Acetyltransferase Antibody, Chemicon®, from goat