Bachmann, Gesine ; Frohns, Florian ; Thangaraj, Gopenath ; Bausch, Alexander ; Layer, Paul G. (2019)
IPL Sublamination in Chicken Retinal Spheroids Is Initiated via Müller Cells and Cholinergic Differentiation, and Is Disrupted by NMDA Signaling.
In: Investigative ophthalmology & visual science, 60 (14)
doi: 10.1167/iovs.18-24952
Article, Bibliographie
Abstract
Purpose
Reaggregates from E6 embryonic chicken retina exhibit areas corresponding to an inner plexiform layer (IPL), which presents an ideal in vitro model to test conditions and constraints of cholinergic and glutamatergic network formation, providing a basis for retinal tissue engineering. Here, we show that ipl formation is regulated by cholinergic starburst amacrine cells (SACs), a glial scaffold and by L-glutamate.
Methods
Rosetted spheroids were cultured in absence or presence of 0.2 to 0.4 mM L-glutamate and analyzed by immuno- and enzyme histochemistry, proliferation, and apoptosis assays.
Results
After 2 days in vitro (div), ipl formation was announced by acetylcholinesterase+ (AChE) and choline acetyltransferase+ (ChAT) cells. Individual vimentin+ or transitin+ Müller glial cell precursors (MCPs) in ipl centers coexpressed ChAT. Comparable to in vivo, pairwise arranged ChAT+ SACs formed two laminar subbands. Projections of calretinin+ amacrine cells (ACs) into ipl associated with MCP processes. In L-glutamate-, or NMDA-treated spheroids ipls were disrupted, including loss of SACs and MCs; coincubation with NMDA receptor inhibitor MK-801 prevented these effects. Also, many Pax6+ cells, comprising most ACs, were lost, while rho4D2+ rod photoreceptors were increased. Cell proliferation was slightly increased, while apoptosis remained unaffected.
Conclusions
This demonstrated: (1) a far-advanced differentiation of an IPL in retinal spheroids, as never described before; (2) ipl sublamination was initiated by cholinergic precursor cells, which-functioning as "ipl founder cells"-(3) gave rise to neurons and glial cells; (4) these SACs and MCPs together organized ipl formation; and (5) this process was counteracted by NMDA-dependent glutamate actions.
| Item Type: | Article |
|---|---|
| Erschienen: | 2019 |
| Creators: | Bachmann, Gesine ; Frohns, Florian ; Thangaraj, Gopenath ; Bausch, Alexander ; Layer, Paul G. |
| Type of entry: | Bibliographie |
| Title: | IPL Sublamination in Chicken Retinal Spheroids Is Initiated via Müller Cells and Cholinergic Differentiation, and Is Disrupted by NMDA Signaling. |
| Language: | English |
| Date: | 1 November 2019 |
| Journal or Publication Title: | Investigative ophthalmology & visual science |
| Volume of the journal: | 60 |
| Issue Number: | 14 |
| DOI: | 10.1167/iovs.18-24952 |
| Abstract: | Purpose Reaggregates from E6 embryonic chicken retina exhibit areas corresponding to an inner plexiform layer (IPL), which presents an ideal in vitro model to test conditions and constraints of cholinergic and glutamatergic network formation, providing a basis for retinal tissue engineering. Here, we show that ipl formation is regulated by cholinergic starburst amacrine cells (SACs), a glial scaffold and by L-glutamate. Methods Rosetted spheroids were cultured in absence or presence of 0.2 to 0.4 mM L-glutamate and analyzed by immuno- and enzyme histochemistry, proliferation, and apoptosis assays. Results After 2 days in vitro (div), ipl formation was announced by acetylcholinesterase+ (AChE) and choline acetyltransferase+ (ChAT) cells. Individual vimentin+ or transitin+ Müller glial cell precursors (MCPs) in ipl centers coexpressed ChAT. Comparable to in vivo, pairwise arranged ChAT+ SACs formed two laminar subbands. Projections of calretinin+ amacrine cells (ACs) into ipl associated with MCP processes. In L-glutamate-, or NMDA-treated spheroids ipls were disrupted, including loss of SACs and MCs; coincubation with NMDA receptor inhibitor MK-801 prevented these effects. Also, many Pax6+ cells, comprising most ACs, were lost, while rho4D2+ rod photoreceptors were increased. Cell proliferation was slightly increased, while apoptosis remained unaffected. Conclusions This demonstrated: (1) a far-advanced differentiation of an IPL in retinal spheroids, as never described before; (2) ipl sublamination was initiated by cholinergic precursor cells, which-functioning as "ipl founder cells"-(3) gave rise to neurons and glial cells; (4) these SACs and MCPs together organized ipl formation; and (5) this process was counteracted by NMDA-dependent glutamate actions. |
| Identification Number: | pmid:31738824 |
| Divisions: | 10 Department of Biology 10 Department of Biology > Developmental Biology and Neurogenetics |
| Date Deposited: | 25 Nov 2019 13:42 |
| Last Modified: | 25 Nov 2019 13:42 |
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