Endogenous Hydrogen Peroxide Regulates the Excitability of Midbrain Dopamine Neurons via ATP-Sensitive Potassium Channels

Abstract

ATP-sensitive K⁺ (K_ATP) channels link metabolic state to cell excitability. Here, we examined regulation of K_ATP channels in substantia nigra dopamine neurons by hydrogen peroxide (H₂O₂), which is produced in all cells during aerobic metabolism. Blockade of K_ATP channels by glibenclamide (100 nm) or depletion of intracellular H₂O₂ by including catalase, a peroxidase enzyme, in the patch pipette increased the spontaneous firing rate of all dopamine neurons tested in guinea pig midbrain slices. Using fluorescence imaging with dichlorofluorescein to visualize intracellular H₂O₂, we found that moderate increases in H₂O₂ during partial inhibition of glutathione (GSH) peroxidase by mercaptosuccinate (0.1-0.3 mm) had no effect on dopamine neuron firing rate. However, with greater GSH inhibition (1 mm mercaptosuccinate) or application of exogenous H₂O₂, 50% of recorded cells showed K_ATP channel-dependent hyperpolarization. Responsive cells also hyperpolarized with diazoxide, a selective opener for K_ATP channels containing sulfonylurea receptor SUR1 subunits, but not with cromakalim, a selective opener for SUR2-based channels, indicating that SUR1-based K_ATP channels conveyed enhanced sensitivity to elevated H₂O₂. In contrast, when endogenous H₂O₂ levels were increased after inhibition of catalase, the predominant peroxidase in the substantia nigra, with 3-amino-1,2,4-triazole (1 mm), all dopamine neurons responded with glibenclamide-reversible hyperpolarization. Fluorescence imaging of H₂O₂ indicated that catalase inhibition rapidly amplified intracellular H₂O₂, whereas inhibition of GSH peroxidase, a predominantly glial enzyme, caused a slower, smaller increase, especially in nonresponsive cells. Thus, endogenous H₂O₂ modulates neuronal activity via K_ATP channel opening, thereby enhancing the reciprocal relationship between metabolism and excitability.