Diaphragm neuromuscular transmission failure in a mouse model of an early-onset neuromotor disorder

The spa transgenic mouse displays spasticity and hypertonia that develops during the early postnatal period, with motor impairments that are remarkably similar to symptoms of human cerebral palsy. Previously, we observed that spa mice have fewer phrenic motor neurons innervating the diaphragm muscle (DIAm). We hypothesize that spa mice exhibit increased susceptibility to neuromuscular transmission failure (NMTF) due to an expanded innervation ratio. We retrogradely labeled phrenic motor neurons with rhodamine and imaged them in horizontal sections (70 µm) using confocal microscopy. Phrenic nerve-DIAm strip preparations from wild type and spa mice were stretched to optimal length, and force was evoked by phrenic nerve stimulation at 10, 40, or 75 Hz in 330-ms duration trains repeated each second (33% duty cycle) across a 120-s period. To assess NMTF, force evoked by phrenic nerve stimulation was compared to force evoked by direct DIAm stimulation superimposed every 15 s. Total DIAm fiber number was estimated in hematoxylin and eosin-stained strips. Compared to wild type, spa mice had over twofold greater NMTF during the first stimulus train that persisted throughout the 120 s period of repetitive activation. In both wild type and spa mice, NMTF was stimulation-frequency dependent. There was no difference in neuromuscular junction morphology or the total number of DIAm fibers between wild type and spa mice, however, there was an increase innervation ratio (39%) in spa mice. We conclude that early-onset developmental neuromotor disorders impair the efficacy of DIAm neuromuscular transmission, likely to contribute to respiratory complications.NEW & NOTEWORTHY Individuals with motor control deficits, including cerebral palsy (CP) often have respiratory impairments. Glycine-receptor mutant spa mice have early-onset hypertonia, and limb motor impairments, similar to individuals with CP. We hypothesized that in the diaphragm of spa mice, disruption of glycinergic inputs to MNs would result in increased phrenic-DIAm neuromuscular transmission failure. Pathophysiologic abnormalities in neuromuscular transmission may contribute to respiratory dysfunction in conditions where early developmental MN loss or motor control deficits are apparent.