Figure 6.

(A) Fluorescence signal increased over time in the nuclei of cells in kidney slices loaded with HEt, as a result of the production of ROS, and the basal rate of ROS production was higher in the PTs than in the DTs. After 20 min, 10 μM rotenone was added to the perfusate. This caused a relatively greater increase in the rate of ROS production in the PTs than in the DTs. Data are means ± SE fluorescence signal per nuclei (expressed as a percentage of the starting signal) from a total of 54 PT and 26 DT cells from four separate slices. (B) Experiments were repeated in the presence of 500 μM apocynin, an inhibitor of NADPH oxidase. As expected, the rates of ROS production were lower, so different microscope settings were used. Under these conditions, the basal rate of ROS production was found to be similar in both tubule types, but, again, a relatively greater rise in the rate of ROS production was observed in PTs in response to rotenone; mean values given are from a total of 37 PT and 31 DT cells from three separate slices. (C) A representative image is depicted of the HEt signal 10 min after rotenone (in the absence of apocynin). (D) Levels of the antioxidant GSH were higher in the PTs than in the DTs, as measured using the fluorescence indicator monochlorobimane. Steady state was reached after 50 min. Data are means ± SE signal per tubule from a total of 40 PTs and 26 DTs from four separate slices. (E) A representative image after 50 min is depicted. Arrowhead, PT; arrow, DT. Bar = 20 μm.