The diffusion of O2 across the red blood cell (RBC) membrane is critical for life. Although the traditional view had been that all O2 moves through RBC membranes by dissolving in and diffusing through the membrane lipid, our recent work (bioRxiv. doi:10.1101/2020.08.28.265066) indicates that this is not the case. Using stopped-flow absorbance spectroscopy to monitor the hemoglobin (Hb) absorbance spectrum in intact RBCs, we found that, compared with RBCs from wild-type (WT) mice, those from double knockout (dKO) mice genetically deficient in both aquaporin-1 (AQP1) and the rhesus (Rh) protein exhibit a 1/3 decrease in the rate constant for O2 offloading from Hb (kHbO2). Mathematical simulations show that this 1/3 decrease in kHbO2 corresponds to a ~55% decrease in the membrane permeability to O2 (PM,O2). In previous abstracts, we reported that the kHbO2 of RBCs from WT mice are indistinguishable from age 2–3 months through 12 months, but begins to fall at 18 months, and drops by ~13% at 29 months—the lifespan of our mice. However, the relatively low kHbO2 of RBCs from dKO mice remains stable through 29 months. In addition to increasing the number of observations for aging WT and dKO mice, we now report the effect of aging on the kHbO2 of RBCs from mice genetically deficient in just RhAG, studying mice at six ages (2–3 months, 6 months, 12 months, 18 months, 24 months, and 29 months). The kHbO2 of RBCs from RhAG-KO mice begins ~17% lower than that of WT mice at 2–3 months (see above bioRxiv ref.), and remains stable through 29 months (N = 5 to 8 at 6–24 months, 4 at 29 months), similar to that of dKO mice. Changes in hematological parameters or RBC size and shape cannot explain the observed decreases in kHbO2 in WT mice. RBC proteomics data on a single WT mouse at 29 months suggests decreases in AQP1 (11%), RhAG (19%), and mRh (27%), and AE1 (22%). We conclude from our dKO and RhAG-KO data that the non-AQP1/RhAG component of RBC membrane O2 permeability is independent of aging, and that the age-related fall in kHbO2 in WT mice is likely to be due, at least in part, to age-related decreased functional expression (i.e., combination of protein number and activity of individual proteins) of AQP1 and RhAG —which function asO2 channels. We continue to build our number of observations on WT, dKO, and RhAG-KO mice of various ages, and are now extending our work to RBCs from mice genetically deficient in just AQP1.