Abstract
Abstract: :
Purpose: In a recent screen for Enhanced Green Fluorescent Protein (EGFP)-expressing cells in transgenic mice generated in our lab, we have found a line in which EGFP is expressed in a subset of amacrine cells in the inner nuclear layer (INL). The purpose of this study was to characterize the amacrine cell based upon general morphology and thier pattern of stratification, using known immunological markers in the retina. Methods: Transgenic mice were generated by injecting a construct consisting of a 1.2 kb CD44 promoter linked to the EGFP gene. Mice were initially screened by PCR and transgenic animals were examined by confocal microscopy. Retinal whole mounts were used to estimate the density of EGFP-positive cells. Double label immunohistochemistry was done on 10µm frozen sections to enable the stratification pattern of the EGFP-labeled terminals to be examined. Results: A population of amacrine cells was found to express EGFP in the retinas from a line of CD44 - EGFP transgenic mice. The labeled cells had a density of 1978± 19.8 cells per mm2. The cells extended their axon arbors into stratum 3 of the retina, which lies between the two bands of ChAT positive amacrine cell processes. The EGFP expressing cells were positive for GAD immunoreactivity suggesting that they were GABAergic. Double labeling with an antibody against the glutamate transporter GLT-1, showed that the cells co-stratified with the GLT-1 positive cone bipolar cell terminals, indicating that these cells may be post synaptic to the GLT-1 cone bipolar cell. Based upon the stratification pattern of the cell, it is likely that the EGFP-labeled cells in our transgenic mouse line are homologs of the On-Off amacrine cell that has previously been described in rabbit retina, and recently named the WF3-1 cell. Conclusion: The present study describes a GABAergic amacrine cell in mouse retina that stratifies in the center of the IPL at the same level as the GLT-1 positive bipolar cell. The EGFP-labeling of the wide-field amacrine cell makes it possible to target this cell type for future electrophysiological and molecular studies.
Keywords: 312 amacrine cells • 434 immunohistochemistry • 557 retina: proximal(bipolar, amacrine, and ganglion cells)