Purchase this article with an account.
Yabana Takeshi, Michele Hooper, Spring Farrell, Balwantray C Chauhan; Characteristics of GCaMP expressing cells in retinal ganglion cell layer of Thy1-GCaMP transgenic mice before and after optic nerve injury.. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3122. doi: https://doi.org/.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
The genetically encoded green fluorescent protein-based calcium sensor, GCaMP, has been used to detect calcium transients and report neuronal activity. We evaluated the specificity of GCaMP expression in retinal ganglion cells (RGCs) of the transgenic Thy1-GCaMP3 mouse line after optic nerve transection (ONT).
A total of 14 Thy1-GCaMP3 mice were used for this study. Control retinas (n = 4) were isolated and stained for RNA-binding protein with multiple splicing (RBPMS) and choline acetyltransferase (ChAT), specific markers for RGCs and amacrine cells, respectively. GCaMP3 expression was detected by GFP immunoreactivity. In a subset of animals, ONT was performed 3, 7, or 14 days before sacrifice (n = 3, 3, 4, respectively). Cells positive for GCaMP, RBPMS, ChAT, as well as the population of co-labeled cells, were quantified in each quadrant at 1.0, 1.5 and 2.0 mm from the optic nerve head.
The mean (SD) densities of GCaMP, RBPMS, and ChAT expressing cells in control retinas were 2663 (110), 3401 (175), and 1041 (47) cells/mm2, respectively. Of the GCaMP positive cells, 92.1 (0.8)% were co-labeled with RBPMS, while 72.2 (1.3)% of RBPMS-labeled cells expressed GCaMP. ChAT expressing cells were neither labeled with GCaMP nor RBPMS (Fig. 1). The number of GCaMP+ and RBPMS+ cells decreased dramatically after ONT; 74.0%, 41.5%, and 18.0% of GCaMP+ and 78.3%, 42.5%, and 15.4% of RBPMS+ cells survived 3, 7, and 14 days after ONT. Furthermore, 71.3%, 36.2%, and 11.5% of GCaMP+/RBPMS+ and 96.3%, 59.2%, and 25.5% of GCaMP-/RBPMS+ cells survived at each time point. However, the number of ChAT+ or GCaMP+/RBPMS- cells did not change (Fig. 2).
In the RGC layer of Thy1-GCaMP mice, GCaMP is expressed by the majority of RGCs. There was an expected progressive and specific loss of GCaMP expression after ONT. Thy1-GCaMP3 transgenic mice show potential for longitudinal assessment of RGCs in injury models.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
This PDF is available to Subscribers Only