Abstract
Purpose :
Previous work in humans and rodent models of diabetes has identified microvascular change as an early pathology in the retina. The aim of this project was to explore the in vivo capillary response to flicker and correlate this with the recently described role of microglia in vascular control.
Methods :
C57Bl6 mice were rendered diabetic (streptozotocin, 50mg/kg) and monitored out to 12 weeks. Retinal capillaries were imaged (Optical coherence tomography angiography, OCTA) at baseline and during flicker stimulation (10Hz flicker at 100 cd/m2, 10 sec). Data were analyzed to explore capillary reactivity. To explore microglial-mediated vasoregulation, Cx3cr1-GCaMP6 mice were rendered diabetic as above and tamoxifen administered (at 10 weeks) to induce GCaMP6 expression in microglia. At 12 weeks, mice were anesthetized, injected with Rhodamine B to visualize vessels and retinal explants imaged via 2-photon microscopy to explore spontaneous microglial Ca2+ activity and associated capillary constriction.
Results :
Baseline OCTA analysis of retinal capillaries after 12 weeks of diabetes showed no alteration in overall density, or diameter relative to control (1.01±0.05 and 1.03±0.02, respectively). Flicker stimulation resulted in a dilation (3.8±1.2%, p<0.05) in retinal capillaries in control animals compared to baseline, while diabetic animals showed no change in capillary width (-0.2±1.3%). Ex vivo 2 photon analysis showed a similar number of microglial Ca2+ responses were associated with localized constrictions (control 52%, STZ 57%) and there was no change in microglial Ca2+ frequency, or constriction frequency and magnitude. Diabetes did result in an increased microglial Ca2+ amplitude (control, 0.29±0.02; diabetes, 0.34±0.01, ΔF/F p<0.05), while the duration of the associated capillary constriction also increased (control, 56.3±2.6; diabetes, 71.1±4.2, secs p<0.05) compared to control explants.
Conclusions :
This work shows that in diabetes, capillaries are unable to respond to normal vasoregulatory cues. This dysfunction correlates with altered microglial vasoregulation kinetics, namely increased Ca2+ amplitude and prolonged capillary constriction. These data further support the role of immune cells in vessel regulation within the retina and highlight early diabetes-induced changes.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.