Energy metabolism in cochlear outer hair cells in vitro

ATP levels in outer hair cells in vitro were measured using the luciferin/luciferase method. Hair cells were isolated from the guinea pig cochlea and maintained for 2 h in a balanced salt solution with 5.5 mM glucose. Ten to 20 cells sufficed for a robust and reproducible luminescence signal, indicating an ATP content of 6.2 +/- 0.4 fmol/cell. This ATP concentration is similar to that found in cultures of other cell types and agrees well with the classical measurements in freeze-dried preparations. The ATP levels were reduced by the following treatments: (1) the omission of glucose in the culture medium lowered ATP levels by 28%; (2) the inhibition of glycolysis by 2-deoxyglucose lowered ATP levels by 66%; (3) the inhibition of oxidative phosphorylation by carbonyl cyanide m-chlorophenylhydrazine (CCCP) lowered ATP levels by 75%, and (4) the inhibition of both pathways reduced the ATP content to non-detectable levels. Acetoacetate was able to restore ATP levels partially when glycolysis was inhibited. These results suggest that (1) the major pathway of ATP synthesis in outer hair cells is the aerobic metabolism of glucose; (2) endogenous energy stores (e.g. glycogen) can maintain ATP levels in the absence of glucose; and (3) ketone bodies may be alternative energy sources.