Acute intermittent hypoxia induced phrenic long-term facilitation despite increased SOD1 expression in a rat model of ALS

Abstract

Amyotrophic lateral sclerosis (ALS) is a progressive and fatal neurodegenerative disease characterized by motor neuron death. Since most ALS patients succumb to ventilatory failure from loss of respiratory motor neurons, any effective ALS treatment must preserve and/or restore breathing capacity. In rats over-expressing mutated superoxide dismutase-1 (SOD1G93A), the capacity to increase phrenic motor output is decreased at disease end-stage, suggesting imminent ventilatory failure. Acute intermittent hypoxia (AIH) induces phrenic long-term facilitation (pLTF), a form of spinal respiratory motor plasticity with potential to restore phrenic motor output in clinical disorders that compromise breathing. Since pLTF requires NADPH oxidase activity and reactive oxygen species (ROS) formation, it is blocked by NADPH oxidase inhibition and SOD mimetics in normal rats. Thus, we hypothesized that SOD1G93A (mutant; MT) rats do not express AIH-induced pLTF due to over-expression of active mutant superoxide dismutase-1. AIH-induced pLTF and hypoglossal (XII) LTF were assessed in young, pre-symptomatic and end-stage anesthetized MT rats and age-matched wild-type littermates. Contrary to predictions, pLTF and XII LTF were observed in MT rats at all ages; at end-stage, pLTF was actually enhanced. SOD1 levels were elevated in young and pre-symptomatic MT rats, yet superoxide accumulation in putative phrenic motor neurons (assessed with dihydroethidium) was unchanged; however, superoxide accumulation significantly decreased at end-stage. Thus, compensatory mechanisms appear to maintain ROS homoeostasis until late in disease progression, preserving AIH-induced respiratory plasticity. Following intrathecal injections of an NADPH oxidase inhibitor (apocynin; 600µM; 12µL), pLTF was abolished in pre-symptomatic, but not end-stage MT rats, demonstrating that pLTF is NADPH oxidase dependent in pre-symptomatic, but NADPH oxidase independent in end-stage MT rats. Mechanisms preserving/enhancing the capacity for pLTF in MT rats are not known.

Mechanisms of Enhanced Phrenic Long-Term Facilitation in SOD1G93A Rats

“…Indeed, phrenic motor neuron death/survival was comparable across all treatment groups ( Fig. 1; Nichols et al, 2013Nichols et al, , 2015. Thus, enhanced moderate AIH-induced G93A and wild-type rats.…”

Intermittent Hypoxia Enhances Functional Connectivity of Midcervical Spinal Interneurons

“…To evaluate functional connectivity, cross-correlation histograms were constructed for all possible pairs of simultaneously recorded neurons using a bin width of 0.2 ms (Moore et al, 1970). The detectability index (DI; Aertsen and Gerstein, 1985) was calculated for each correlogram as the peak (or trough) relative to average background activity (calculated over 12 ms before the trigger), divided by the SD.…”

“…The response of C-INs to repeated bouts of hypoxia [acute intermittent hypoxia (AIH)] is of interest because AIH can trigger spinal neuroplasticity and sustained increases in respiratory, autonomic, and/or somatic motor output (Dick et al, 2007;Lovett-Barr et al, 2012;. For these reasons, AIH is being actively explored as a therapeutic modality to promote motor recovery following neurologic disorders including spinal cord injury (Trumbower et al, 2012;Hayes et al, 2014;Tester et al, 2014;Gonzalez-Rothi et al, 2015) and amyotrophic lateral sclerosis (ALS; Nichols et al, 2013Nichols et al, , 2015.…”

Quantitative assessment of integrated phrenic nerve activity

“…Heterozygous SOD1 G93A progeny were identified with polymerase chain reaction (PCR) of tail DNA with primers specific for hSOD1. SOD1 G93A rats were considered end-stage when they had lost 20% of their peak body mass, as in previous reports (Nichols et al, 2013a; Nichols et al, 2014; Nichols et al, 2015a). …”