Abstract
Purpose :
In darkness, there is a current that circulates through the rod; Na+along with some Ca2+enters the outer segment through cyclic-nucleotide-gated (CNG) ion channels and K+exits the inner segment through voltage gated ion channels. Photon stimulation hydrolyzes cGMP, closes CNG channels and suppresses the circulating current. By stimulating rod outer segment guanylate cyclase (ROS-GC) to synthesize cGMP at a faster rate in a Ca2+-dependent fashion. bicarbonate has been shown to affect the circulating current in darkness as well as the photon response amplitude and kinetics in amphibian rods. Here, we wanted to see how bicarbonate impacts the physiology of murine rods, which are considerably smaller in size, operate at higher temperature and generate faster photon responses.
Methods :
The ex vivo electroretinogram (ERG) was recorded from retinas of mice deficient in cone transducin, in order to isolate the rod response to photic stimulation. Retina were perfused with Ames solution containing either 20μM bicarbonate or equimolar Cl-in place of bicarbonate. 50μM DL-AP4 and 100μM BaCl2 were added to block post-photoreceptoral and glial responses, respectively. Solutions were maintained at 35°C and bubbled with 95% O2/5% CO2. Retinas were stimulated with 20ms flashes at 500nm and with 10s steps at 540nm through various neutral density filters.
Results :
Murine rods responded reversibly to bicarbonate with an increased in circulating current by 1.4-fold(n=7). The integration time of dim flash response decreased by 1.3-fold(n=6) indicating faster flash response recovery. Sensitivity to flashes were reduced by 1.7-folds(n=3).
The time in saturation, measured from midflash to 20% recovery of circulating current, were lower for bicarbonate experiments. Flash almost 2-fold brighter was required to hold rods in saturation.
Bicarbonate decreased the step sensitivity of rods by 1.4-fold (for 540nm light), this is attributed to 1.3-fold lower integration time of dim flash response.
Conclusions :
In contrast to 20μM bicarbonate used here, previously observed in amphibian rods, bicarbonate at 50mM increased circulating current by 17± 3% with similar dim flash response as in murine rods. Amphibian’s sensitivity to flashes was reduced by 52± 10%, while that of murine rods reduced by 7± 0.5% (n=3).
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.