Abstract
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the selective loss of defined motoneuron populations in the brainstem and spinal cord. Although low cytosolic calcium ([Ca2+]i) buffering and a strong interaction between metabolic mechanisms and [Ca2+]i have been associated with selective motoneuron vulnerability, the underlying cellular mechanisms are barely understood. To elucidate the underlying molecular events, we used rapid charge-cooled device imaging to evaluate Ca2+ signaling and metabolic signatures in the brainstem slices of SOD1G93A mice, the mouse model of human ALS, at 8 to 9 and 14 to 15 weeks of age, corresponding to the presymptomatic and symptomatic stages of motor dysfunction, respectively, and compared the results with corresponding age-matched wild-type littermates. We also monitored the mitochondrial membrane potential (ΔΨm) of brainstem motoneurons, a valuable tool for characterizing the metabolic signature of intrinsic energy profiles and considered to be a good experimental measure for monitoring energy metabolism in cells. We found that different pharmacological interventions substantially disrupt ΔΨm in SOD1G93A motoneurons during the symptomatic stage. Furthermore, we investigated the impact of impaired mitochondrial mechanisms on [Ca2+]i regulation by using the membrane-permeable indicator fura-acetoxy methyl ester. Taken together, the results indicate that mitochondrial disruptions are critical elements of SOD1G93A-mediated motoneuron degeneration in which selective motoneuron vulnerability results from a synergistic accumulation of risk factors, including the disruption of electrochemical potential, low Ca2+ buffering, and strong mitochondrial control of [Ca2+]i. The stabilization of mitochondria-related signal cascades may represent a useful strategy for clinical neuroprotection in ALS.
Footnotes
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This study was supported by the Bundesministerium für Bildung und Forschung (BioRegioN/ERANET) [Grant 031 3610A]; the Bernstein Center for Computational Neuroscience Göttingen [Grant 136 2870]; and the Göttingen Center for Molecular Physiology of the Brain [Grant 135 6834].
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ABBREVIATIONS: ALS, amyotrophic lateral sclerosis; Δψm, mitochondrial membrane potential; [Ca2+]i, cytosolic calcium; [Ca2+]mito, mitochondrial calcium; aCSF, artificial cerebrospinal fluid; AM, acetoxymethyl ester; bp, base pair(s); CCD, charge-coupled device; CPA, cyclopiazonic acid; DMSO, dimethyl sulfoxide; ER, endoplasmic reticulum; F, fluorescence; fALS, familial amyotrophic lateral sclerosis; FCCP, carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone; hALS, human amyotrophic lateral sclerosis; HMN, hypoglossal motoneuron; PCR, polymerase chain reaction; rhod123, rhodamine 123; ROS, reactive oxygen species; RT, room temperature; SERCA, sarco-endoplasmic reticulum Ca2+ ATPase; SOD1, Cu/Zn superoxide dismutase 1; WT, wild-type.
- Received August 9, 2008.
- Accepted December 4, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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