Abstract
Purpose :
Recent advances in adaptive optics ophthalmoscopy have enabled exact counts of single red blood cells (RBCs) as they travel within retinal capillaries. Because the approach is non-invasive, it is now possible to reimage the same capillaries from seconds to months providing a unique opportunity to study the functional variables of the capillary network.
Methods :
6 healthy male C57BL6/J mice (6-12 weeks old) were imaged every two weeks with a custom adaptive optics scanning light ophthalmoscope (AOSLO). We imaged retinal capillaries in the deep stratification at the outer plexiform layer (7-22° from the optic disc). RBCs were imaged with phase contrast enabled by lateral offset pinhole imaging (30 Airy disc diameter offset, 796 nm). RBC flux was measured by placing a 15 kHz line scan across each capillary for 20 s and performing manual RBC counts for 1 s of data.
Results :
AOSLO allowed noninvasive imaging of the capillary network at high resolution permitting return to target capillaries at weekly intervals (Fig. 1). We identified 3 key attributes of this network by tracking 55 capillaries in 6 mice:
1) The same capillaries measured 18 s apart show low RBC flux variability. The average standard deviation was 5% (ranging from 0.07% to 35%; mean = 157 cells/s). The dominant sources of this modulation were the cardiac cycle and sporadic events like leukocyte passage or capillary shunting.
2) The same capillaries measured at two-week intervals show greater variability. RBC flux measured across weeks showed an average standard deviation of 21% (ranging from 0.3% to 89%). Despite greater variability in single vessels, the network flux changed by 6% suggestive of a redistribution of flow.
3) RBC flux showed the highest variability when comparing all 55 capillaries across 6 mice. In this case standard deviation was 35% (data compared at 8-week time point). This increase could reveal differences in physiology across mice despite being the same age, gender and strain.
Conclusions :
Here we show the first report of repeat measures of retinal capillary flux in mice across time. At the scale of seconds, flux is not constant in the same capillary; modulation arise from heart rate and stochastic shunting. At the level of weeks, individual capillaries show greater changes though network flux is largely maintained. This study in healthy mice provides a baseline from which future studies can track flux changes in retinal disease.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.