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Characterization of three‐dimensional rat central nervous system culture maturation, with applications to monitor cholinergic integrity
Author(s) -
Andersen Parker L.,
Vermette Patrick,
Khalil Abdelouahed,
Witkowski Jacek M.,
Fülöp Tamas
Publication year - 2020
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1002/btpr.2976
Subject(s) - acetylcholine , cholinergic , biology , central nervous system , neurite , glutamate receptor , synapse , neuroscience , microbiology and biotechnology , tissue culture , in vivo , in vitro , biochemistry , endocrinology , genetics , receptor
Studying age‐related neuropathologies in vitro requires a three‐dimensional (3D) culture system presenting mature phenotypes. In this study, we aimed to determine whether aged reaggregate cultures physiologically represent mature brain tissue. Results support that embryo‐derived rat central nervous system (CNS) reaggregate cultures develop into mature‐like tissues, comparable to in vivo maturation, including the following characteristics: (a) progressive reduction in cell proliferation (reduced anti‐Ki‐67 immunoreactivity), (b) progressive restriction of long neurite growth potential (as explant cultures), and (c) increased and sustained synaptic enzyme (acetylcholine esterase, AChE) activity. The acquisition of mature‐like reaggregate cultures has allowed us to pursue the hypothesis that the physiological integrity of 3D CNS cultures may be monitored by synaptic enzyme activity. To assess this hypothesis, mature‐like reaggregates were exposed to H 2 O 2 , glutamate, or amyloid β(1–42); each resulted in diminished AChE activity. H 2 O 2 exposure resulted in nuclear fragmentation. Glutamate and amyloid β(1–42) exposure resulted in acetylcholine content reduction. Simultaneous reduction of AChE activity and acetylcholine content verified diminished cholinergic integrity. This scheme exploiting synapse enzyme activity of mature‐like 3D CNS tissue is therefore applicable to age‐related neuropathology research including in vitro screening of conditions potentially affecting synapse integrity, including the promotion of dementia.