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Appearance of cholinergic myenteric neurons during enteric nervous system development: comparison of different Ch AT fluorescent mouse reporter lines
Author(s) -
Erickson C. S.,
Lee S. J.,
BarlowAnacker A. J.,
Druckenbrod N. R.,
Epstein M. L.,
Gosain A.
Publication year - 2014
Publication title -
neurogastroenterology and motility
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.489
H-Index - 105
eISSN - 1365-2982
pISSN - 1350-1925
DOI - 10.1111/nmo.12343
Subject(s) - enteric nervous system , choline acetyltransferase , myenteric plexus , cholinergic , cholinergic neuron , green fluorescent protein , biology , neurotransmitter , microbiology and biotechnology , neurogenesis , acetylcholine , neuroscience , vesicular acetylcholine transporter , central nervous system , endocrinology , immunohistochemistry , biochemistry , immunology , gene
Background Cholinergic neurons have been identified with the acetylcholine synthetic enzyme choline acetyltransferase (Ch AT ). However, Ch AT is difficult to localize in newly differentiated peripheral neurons making the study of cholinergic neuronal development problematic. Consequently, researchers have used mouse reporter lines to indicate the presence of Ch AT . Methods Our objective was to determine which Ch AT reporter line was the most sensitive indicator of Ch AT expression. We utilized two different fluorescent Ch AT reporter lines ( Ch AT ‐ GFP and Ch AT ‐Cre;R26R:flox STOP :tdTomato ) together with immunolocalization of Ch AT protein (Ch AT ‐ IR ) to characterize the spatial and temporal expression of Ch AT in myenteric neurons throughout enteric nervous system ( ENS ) development. Key Results Ch AT ‐ IR cells were first seen in the intestine at E10.5, even within the migration wavefront of neural precursors. Myenteric neurons within the distal small intestine (d SI ) and proximal colon were first labeled by Ch AT ‐ IR , then Ch AT ‐ GFP , and finally Ch AT ‐Cre tdTomato. The percentage of Ch AT ‐ IR neurons is equivalent to adult levels in the d SI by E13.5 and proximal colon by P0. After these stages, the percentages remained relatively constant throughout development despite dramatic changes in neuronal density. Conclusions & Inferences These observations indicate that neurotransmitter expression occurs early and there is only a brief gap between neurogenesis and neurotransmitter expression. Our finding that the proportion of Ch AT myenteric neurons reached adult levels during embryonic development suggests that the fate of cholinergic neurons is tightly regulated and that their differentiation might influence further neuronal development. Ch AT ‐ GFP is a more accurate indicator of early ENS cholinergic neuronal differentiation than the Ch AT ‐Cre;R26R:flox STOP :tdTomato reporter mouse.