Decrease in a Leak Conductance Associated with Mitochondrial Complex V and Improved Bioenergetic Efficiency may Underlie Cytoprotection of at-Risk Neurons by Dexpramipexole

Abstract

Chronic neurodegenerative disorders have been linked to mitochondrial dysfunction. Pharmaceutical agents which reduce mitochondrial dysfunction may provide therapeutic benefit by protecting neurons at risk. In the current study we have investigated the effects of dexpramipexole ((6R)-4,5,6,7-tetrahydro-N6-propyl-2,6-benzothiazole-diamine), a drug currently under clinical study in ALS patients and a putative mitochondria-targeted neuroprotectant. Mitochondria isolated from proteasome-inhibitor-injected rat brain had large or intermediate-level multiconductance channel activity not present in controls; dexpramipexole inhibited this activity. Dexpramipexole was cytoprotective over a concentration range that represented the EC80-100 for the effects on the conductances, suggesting that full inhibition may be required for significant cytoprotection. To focus on the relationship of dexpramipexole to cell bioenergetics, recordings were made from submitochondrial vesicles enriched in complex V (SMVs). Control SMVs were leaky in response to stimulus voltage, and currents were attenuated in a concentration-dependent manner by application of ATP or dexpramipexole. Dexpramipexole enhanced the rate of ATP hydrolysis and synthesis by complex V in a concentration-dependent manner and maintained ATP production in cultured hippocampal neurons and SHSY5Y cells. Using two methods, the Seahorse respirometry system and an oxygen sensor that detects oxygen flux in single neurons, we determined that dexpramipexole decreases cellular oxygen use, consistent with the effect on inhibiting inner membrane proton leak. Removal of F1 from SMVs eliminated the effects of dexpramipexole (but not ATP) on the inhibition of conductance, and significantly attenuated binding of 14C-dexpramipexole. Dexpramipexole also eliminated the switch from mitochondrial to glycolytic respiratory parameters elicited by treatment of cultured cells with PSI. These data suggest that an increase in efficiency of oxidative phosphorylation produced by dexpramipexole may result from inhibition of complex V leak conductance, enhancing the survivability of neurons.