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B. Trexler; Persistence of Dark Noise in Healthy, Well Dark Adapted Retinas. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3849. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Dark adapted AII amacrine cells display vigorous spontaneous activity that arises primarily from rod bipolars (RBPs). APB application substantially reduces noise, suggesting that RBPs constitutively release glutamate in the dark. However, AII dark noise might be the artifact of damage during tissue preparation or the result of insufficiently dark adapted retinas. Therefore, we examined ganglion cell thresholds and tested for possible damage to rod photoreceptors as a measure of retina sensitivity and health.
Whole cell patch clamping was performed on dark adapted rabbit (New Zealand White) retina slices and flat mount retinas. Retinas were stained with DAPI or FITC-WGA (wheat germ agglutanin) to visualize the retinal pigment epithelium (RPE) layer. Limited intensity and duration 660 nm light during surgery, dissection, and tissue mounting coupled with longer wavelength illumination (860 nm and 940 nm) during slice transfer and microscopic imaging, respectively, ensured that only a minute portion of the 10^8 rhodopsin molecules were bleached before recordings. Light intensities and durations were measured for all procedures. Based on a rod collecting area of 1 micron and assuming perfect absorption of all incident photons, the number of rhodopsin bleached was at worst, 500. More realistic estimates place the number at ~100.
L-APB diminished the dark noise of AII amacrine cells and eliminated the dark adapted light response in both retina slices and flat mount retinas. AII threshold responses are only visible in averages of several flash presentations. Ganglion cell thresholds, however, were quite low (flash intensity ~ 0.001 R* per rod), comparable to thresholds from psychophysical data and other in vitro retina preparations. Both slice and flat mount preparations retain the retinal pigment epithelium, arguing that photoreceptors are intact and releasing normal amounts of glutamate.
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