Two‐photon AMPK and ATP imaging reveals the bias between rods and cones in glycolysis utility

Abstract

In vertebrates, retinal rod and cone photoreceptor cells rely significantly on glycolysis. Lactate released from photoreceptor cells fuels neighboring retinal pigment epithelium cells and Müller glial cells through oxidative phosphorylation. To understand this highly heterogeneous metabolic environment around photoreceptor cells, single-cell analysis is needed. Here, we visualized cellular AMP-activated protein kinase (AMPK) activity and ATP levels in the retina by two-photon microscopy. Transgenic mice expressing a hyBRET-AMPK biosensor were used for measuring the AMPK activity. GO-ATeam2 transgenic mice were used for measuring the ATP level. Temporal metabolic responses were successfully detected in the live retinal explants upon drug perfusion. A glycolysis inhibitor, 2-deoxy-d-glucose (2-DG), activated AMPK and reduced ATP. These effects were clearly stronger in rods than in cones. Notably, rod AMPK and ATP started to recover at 30 min from the onset of 2-DG perfusion. Consistent with these findings, ex vivo electroretinogram recordings showed a transient slowdown in rod dim flash responses during a 60-min 2-DG perfusion, whereas cone responses were not affected. Based on these results, we propose that cones surrounded by highly glycolytic rods become less dependent on glycolysis, and rods also become less dependent on glycolysis within 60 min upon the glycolysis inhibition.

Catalytic isoforms of AMP-activated protein kinase differentially regulate IMPDH activity and photoreceptor neuron function

“…We measured IMPDH activity from retinal lysates of wild-type mice treated with metformin and observed a substantial abrogation of IMPDH activity in metformin-treated lysates. He et al also demonstrated that the effect of AMPK activation by metformin in the retina is notably slower than other that of other AMPK activators (40), which corroborates the delay in IMPDH activity inhibition seen in our assay. These results suggest that a potent effect of metformin in the retina is inhibition of IMPDH activity, which could be through activation of AMPK.…”

Photoreceptor Manifestations of Primary Mitochondrial Optic Nerve Disorders

“…41 , 42 In particular, a recent study using reporter mice showed that rods are more dependent on glycolysis than cones, which may imply that cones are more reliant on mitochondria as their energy source and therefore more susceptible to mitochondrial dysfunction than rods. 43 Taken together, we would expect more cone than rod dysfunction in patients with LHON or OPA1.…”

Glucose uptake by GLUT1 in photoreceptors is essential for outer segment renewal and rod photoreceptor survival

“…Previous studies have shown that cones are less dependent on glycolysis than rods in non‐degenerating retinas 6,14,43 . However, in models of RP, cones maintain their viability by upregulating aerobic glycolysis through stabilization of GLUT1 expression at the plasma membrane of cone photoreceptors 2,6 .…”

“…In a non-diseased rod dominant retina, it was proposed that cones may rely more on oxidative phosphorylation than on glycolysis. 6,43 A recent study using the rd10 mouse showed exogenous lactate was sufficient to promote cone survival when TCA cycle activity was boosted by overexpression of thioredoxin interacting protein (Txnip) allele. The rescue was also dependent on LDHB, which catalyzes the conversion of lactate to pyruvate.…”

“…Previous studies have shown that cones are less dependent on glycolysis than rods in non-degenerating retinas. 6,14,43 However, in models of RP, cones maintain their viability by upregulating aerobic glycolysis through stabilization of GLUT1 expression at the plasma membrane of cone photoreceptors. 2,6 Given that cones represent only 3% of photoreceptors in the mouse retina, it is difficult to assess impacts on cone viability with SD-OCT. To examine if loss of cone photoreceptors contributed to the observed thinning of the Ret∆Glut1 ONL, 4-month-old retinal flat mounts were immunolabeled for cone arrestin (ARR3) to quantify cone density (Figure 4A).…”