Abstract
Basal forebrain cholinergic neurons (BFCNs) are believed to be one of the first cell types to be affected in all forms of AD, and their dysfunction is clinically correlated with impaired short-term memory formation and retrieval. We present an optimized in vitro protocol to generate human BFCNs from iPSCs, using cell lines from presenilin 2 (PSEN2) mutation carriers and controls. As expected, cell lines harboring the PSEN2N141I mutation displayed an increase in the Aβ42/40 in iPSC-derived BFCNs. Neurons derived from PSEN2N141I lines generated fewer maximum number of spikes in response to a square depolarizing current injection. The height of the first action potential at rheobase current injection was also significantly decreased in PSEN2N141I BFCNs. CRISPR/Cas9 correction of the PSEN2 point mutation abolished the electrophysiological deficit, restoring both the maximal number of spikes and spike height to the levels recorded in controls. Increased Aβ42/40 was also normalized following CRISPR/Cas-mediated correction of the PSEN2N141I mutation. The genome editing data confirms the robust consistency of mutation-related changes in Aβ42/40 ratio while also showing a PSEN2-mutation-related alteration in electrophysiology.
Highlights
The “amyloid hypothesis” is one of the most popular formulations for the pathogenesis of Alzheimer’s disease (AD)
Optimization of protocol for basal forebrain cholinergic neurons (BFCNs) differentiation The scheme of BFCN differentiation is described in Fig. 1a. iPSCs from control subjects or AD patients were plated in feeder-free conditions and allowed to reach 100% confluency prior to differentiation using mTeSR1 basal media
Lines and controls With the aim of determining the impact of PSEN2N141I mutation on the differentiation, gene expression, function, and communication of BFCNs, we characterized cells at later time points for appropriate expression markers; our goal was to explore whether PSEN2N141I iPSC were able to complete BFCN maturation process and if so, if any abnormalities along later stages of BFCN differentiation may account for the pathophysiology of early onset familial Alzheimer’s disease (EOFAD) (Fig. 3)
Summary
The “amyloid hypothesis” is one of the most popular formulations for the pathogenesis of Alzheimer’s disease (AD). Ortiz-Virumbrales et al Acta Neuropathologica Communications (2017) 5:77 presence of a presynaptic cholinergic deficit in the basal forebrain of AD patients [1, 71]. Post-mortem brain biochemical and volumetric studies at different stages of the disease identified several other regions of the brain that were affected early in the course of AD [63]. These studies have traditionally focused on the hippocampus and cortex, but more recently, attention has begun shifting back to the basal forebrain and adding other areas, such as the striatum [27, 62]. The latest analyses suggest that cholinergic basal forebrain volume measurement may be a better predictor of the transition from MCI to AD than the previous standard, hippocampal volume [10]
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call