Mitochondria in motor nerve terminals: function in health and in mutant superoxide dismutase 1 mouse models of familial ALS

Abstract

Mitochondria contribute to neuronal function not only via their ability to generate ATP, but also via their ability to buffer large Ca(2+) loads. This review summarizes evidence that mitochondrial Ca(2+) sequestration is especially important for sustaining the function of vertebrate motor nerve terminals during repetitive stimulation. Motor terminal mitochondria can sequester large amounts of Ca(2+) because they have mechanisms for limiting both the mitochondrial depolarization and the increase in matrix free [Ca(2+)] associated with Ca(2+) influx. In mice expressing mutations of human superoxide dismutase -1 (SOD1) that cause some cases of familial amyotrophic lateral sclerosis (fALS), motor terminals degenerate well before the death of motor neuron cell bodies. This review presents evidence for early and progressive mitochondrial dysfunction in motor terminals of mutant SOD1 mice (G93A, G85R). This dysfunction would impair mitochondrial ability to sequester stimulation-associated Ca(2+) loads, and thus likely contributes to the early degeneration of motor terminals.

Fig. 1

Mitochondrial regulation of cytosolic [Ca²⁺] is important for maintaining neuromuscular transmission at mouse motor nerve terminals. a. fluorescence micrograph of motor terminal ionophoretically injected with a Ca²⁺ indicator. The online version of the small pictures at right shows the response to 50 Hz stimulation of the motor axon; the intensity of the fluorescence of the Ca-indicator (normalized to pre-stimulation values) is encoded on a pseudo-color scale, where blue, green, yellow and red represent increasing [Ca²⁺] elevations. b. elevation of cytosolic [Ca²⁺] induced by stimulation (horizontal bar) under control conditions and following inhibition of mitochondrial Ca²⁺ uptake by depolarizating Ψ_m with antimycin (inhibits complex III of the ETC; Oligomycin blocks Complex V, the ATP synthase. c. changes in EPP quantal content (m/m_prestim) during and following stimulation under control conditions, in oligomycin, and following Ψ_m depolarization (depol). Ψ_m depolarization enhances depression and eliminates post-tetanic potentiation. d. Changes in cytosolic [Ca²⁺], mitochondrial matrix [Ca²⁺] and Ψ_m evoked by 3 bouts of stimulation at 100 Hz, monitored (respectively) by changes in the fluorescence of ionophoretically injected Oregon Green (OG)-5 N, X-rhod-1 loaded into mitochondria, and rhodamine (Rh)-123. Each trace comes from a different preparation of the levator auris longus muscle, whose contractions were blocked with d-tubocurarine or μ-conotoxin GIIIA. Records are normalized to pre-stimulation values. Parts B and C adapted from David and Barrett (2003); part D from Nguyen et al. (2009)

Fig. 2

Stimulation-induced Ψ_m depolarizations increase in motor terminals of mutant SOD1 mice, with transient opening of the mPTP in symptomatic fALS mice. a. Representative traces comparing Ψ_m depolarizations (monitored as increase in Rh-123 fluorescence) in motor terminals of wild-type (WT) mice, mice lacking SOD1 (SOD1-KO), mice that express normal human SOD1 (in addition to mouse SOD1, SOD1-OX), and presymptomatic mice expressing the fALS-causing G85R and G93A mutations of human SOD1. Dots below the traces indicate stimulus trains (each 100 Hz, 5 s; same pattern as in Fig. 1d). Bar graph plots averaged results for the first stimulus train, indicating that fALS motor terminals show a ~5-fold greater fluorescence increase than WT terminals. The inset micrograph illustrates regions of an Rh-123-loaded presymptomatic G85R mouse motor terminal whose fluorescence increased during stimulation (difference between resting and stimulated images). b. Superimposed traces illustrating the larger, summating, stimulation-induced Ψ_m depolarizations recorded in motor terminals of older, symptomatic G85R and G93A mice. On this scale the smaller Ψ_m depolarizations in a motor terminal expressing normal human SOD1 (hSOD1wt) are nearly undetectable. Arrows indicate additional, asynchronous Ψ_m depolarizations. c. The large Ψ_m depolarizations recorded in a symptomatic G93A motor terminal are reduced by cyclo-sporin A (CsA), which inhibits mPTP opening. Part A adapted from Nguyen et al. (2009); parts B and C from Nguyen et al. (2011)