Measures of bulbar and spinal motor function, muscle innervation, and mitochondrial function in ALS rats

Abstract

Symptom onset in amyotrophic lateral sclerosis (ALS) may occur in the muscles of the limbs (spinal onset) or those of the head and neck (bulbar onset). Most preclinical studies have focused on spinal symptoms, despite the prevalence of and increased morbidity and mortality associated with bulbar disease. We measured lick rhythm and tongue force to evaluate bulbar disease in the SOD1-G93A rat model of familial ALS. Body weight and grip strength were measured concomitantly. Testing spanned the early (maturation), middle (pre-symptomatic), and late (symptomatic and end-stage) phases of the disease. We measured a persistent tongue motility deficit that became apparent in the early phase of the disease, providing behavioral evidence of bulbar pathology. At end-stage, however, cytochrome oxidase (CO) activity was normal in the hypoglossal nucleus, and in the tongue, neuromuscular innervation, citrate synthase (CS) protein levels and activity, and uncoupling protein 3 (UCP3) protein levels remained unchanged. Interestingly, significant denervation and atrophy were evident in the end-stage sternomastoid muscle, providing peripheral anatomical evidence of bulbar pathology. Changes in body weight and grip strength occurred in the late phase of the disease. Extensive atrophy and denervation were observed in the end-stage gastrocnemius muscle. In contrast to our findings in the tongue, CS protein levels were decreased in the extensor digitorum longus (EDL) and soleus, although CS activity was maintained or increased. UCP3 protein was decreased also in the EDL. These data provide evidence of differential effects in muscles that were more or less affected by disease.

Histological Bulbar Manifestations in the ALS Rat

“…While this conflicts with the study by Smittkamp et al [14] that did not show denervation in tongue muscle of the SOD1-G93A rat, they did report denervation in the sternomastoid muscle, which we did not evaluate in the current study. Moreover, it should be noted that our current study utilized a relatively small sample size (n = 3-6).…”

“…This latter finding may be due to the rapid demise of the animals used in that study, such that there was not enough time for MNs in cranial nerve nuclei to undergo degeneration; the authors did, however, observe vacuolation in hypoglossal and facial nuclei [16]. Similarly, in another study the hypoglossal nucleus had no decrease in activity [14]. Furthermore, the reported heterogeneity in MN counts across studies can likely be explained by the susceptibility of SOD1-G93A rats to genetic drift [9].…”

“…Alternatively, there may be inherent differences between disease symptomology in SOD1-G93A rats relative to human ALS subjects. Of note, however, tongue motility and functioning, which exhibited deficits in a separate study [14], were not tested here. Lastly, our SOD1-G93A rats exhibited motor deficits earlier than what is described in the literature and died later than expected; therefore, the longer, slower progression suggests that these rats might have had a less aggressive form of disease that mitigated the development of overt facial motor function deficits.…”

Pre- and postsynaptic mechanisms underlying inhibition of hypoglossal motor neuron excitability by riluzole

“…Progressive bulbar impairment of speech and swallowing are also frequently co-occurrences in limb-onset ALS patients (Fujimura-Kiyono et al 2011;Küh-nlein et al 2008;Weikamp et al 2012), and symptoms due to bulbar deficits are among the most distressing for ALS patients (Hillel and Miller 1989). Loss of HMs (Dal Canto and Gurney 1995;Ferrucci et al 2010) and lingual movement deficits (Smittkamp et al 2008(Smittkamp et al , 2010 are also seen in transgenic animal models of ALS; HMs from transgenic models of ALS show an early onset of hyperexcitability due to an increased persistent Na ϩ current density (van Zundert et al 2008). HMs are therefore an important motor neuron group known to be susceptible to early changes in excitability and adult demise in ALS and its transgenic models.…”

Differential effects of amphetamine and GBR-12909 on orolingual motor function in young vs aged F344/BN rats

“…We used the experimental chamber that was designed and first used by Fowler et al (Fowler and Das, 1994; Fowler and Mortell, 1992) and in our recent studies (Smittkamp et al, 2008, 2010; Stanford et al, 2003). Data were recorded in a modified rodent operant chamber with a front panel containing a 6-cm square hole at floor level.…”