Article Figures & Data
Figures
- Figure 1.
The lack of CD73 prevents the metabolism of AMP and ATP to adenosine. A, Superimposed adenosine biosensor and null traces illustrating production of adenosine from 30 μm AMP by cerebellar slices from CD73+/+ and CD73−/− mice. B, As in A but for 30 μm ATP. C, Addition of 5′nucleotidase (e5′n; 5 kU/ml) increased adenosine production from ATP (30 μm) in a slice from a CD73−/− mouse. D, Application of 5′nucleotidase (5 kU/ml) had little effect on adenosine production in a slice from a CD73+/+ mouse. E, Graph summarizing conversion of AMP, ATP, and cAMP into adenosine by slices from CD73+/+ and CD73−/− mice (e5′n: breakdown of ATP into adenosine by slices from CD73−/− mice in the presence of 5 kU/ml of 5′-nucleotidase; n = 4–5). **p < 0.01, Student's t test. F, Graph plotting the amplitude of individual normalized parallel fiber fEPSPs against time recorded from a cerebellar slice from a CD73+/+ mouse. Inset, superimposed fEPSP averages in control and 100 μm ATP. G, As in F but for a slice from a CD73−/− mouse. Inset, superimposed fEPSP averages in control and 100 μm ATP.
- Figure 2.
Properties of adenosine release in CD73 mice. A, Adenosine biosensor trace showing adenosine release evoked twice in control and then reduced after EHNA (20 μm) application. B, TTX (1 μm) abolished adenosine release (arrows show where stimulations occurred but no adenosine was detected). C, Adenosine release was abolished in ACSF containing no Ca2+ and 1 mm EDTA. The biosensor traces in A and B were recorded from slices from CD73+/+ mice, whereas in C the slice was from a CD73−/− mouse.
- Figure 3.
Loss of CD73 expression reveals direct adenosine release. A, Graph summarizing adenosine release measured from 21 and 18 slices from CD73−/− and CD73+/+ mice, respectively. Each point represents the concentration of adenosine evoked by stimulation from an individual slice. The lines represent the mean concentrations of adenosine detected in slices from CD73+/+ and CD73−/− mice. B, Adenosine release evoked in a slice from a CD73+/+ mouse is reduced by ∼ 50% after the block of ionotropic glutamate receptors with 20 μm CNQX. C, CNQX (20 μm) has no effect on the adenosine release evoked in a slice from a CD73−/− mouse. D, Graph summarizing the adenosine release data (n = 5–6). ** p < 0.01, Student's t test. E, Evoked adenosine release in a slice from a CD73−/− mouse was reversibly reduced by the mGlu4 receptor agonist l-AP4 (50 μm).
- Figure 4.
Direct adenosine release is blocked by bafilomycin. A, Adenosine release evoked in a slice from a CD73−/− mouse was reduced and almost abolished after incubation with bafilomycin (3 μm). B, Graph summarizing the effect of bafilomycin incubation on adenosine release evoked in slices from CD73−/− mice (n = 7) compared with interleaved control slices (n = 6). Where bafilomycin (3 μm) or DMSO (0.1%) was applied is indicated by the arrow. The adenosine release has been normalized by dividing by the mean concentration of adenosine released by the control stimulations. The rundown of adenosine release in control was described by a linear fit (R, 0.93; slope, 0.01 min−1). Inset, same as B but in slices from rats. C, Graph plotting the mean percentage of adenosine release remaining after 40–60 min in control and bafilomycin. ***p < 0.001, Student's t test.
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