Induction of the immediate early gene c- jun in human spinal cord in amyotrophic lateral sclerosis with concomitant loss of NMDA receptor NR-1 and glycine transporter mRNA

Abstract

The aetiology of the sporadic form of amyotrophic lateral sclerosis (ALS) is poorly understood although abnormalities in glutamate and glycine transport have been implicated which both could contribute to a neurodegenerative process mediated through the N-methyl- d-aspartate (NMDA) receptor. In this study we have used in situ hybridization to investigate whether any changes in the expression of NMDA receptors, the glycine transporter or glutamate-mediated injury responses are detectable in ALS. Two immediate early genes were investigated as markers of neuronal injury responses, c- jun and zif-268, both constitutively expressed in the spinal cord. Levels of c- jun mRNA were most abundant in intermediate grey and layer IX of the ventral horn containing motor neurones. This pattern was markedly changed in ALS with large increases (2–3 fold) in c- jun mRNA occurring in dorsal and ventral horn. The marked increase in c- jun mRNA was also substantiated by slot blot analysis of tissue homogenates of spinal cord and a parallel induction of zif-268 mRNA was also seen. NMDA receptor NR-1 mRNA was widely distributed in control spinal cord with the highest concentrations occurring in layers IX, X, intermediate grey and dorsal horn. The ALS cases showed a selective decrease in the level of NR-1 mRNA in the ventral region (50%) whilst no significant decrease was detected in the dorsal region. Quantitation of tissue homogenates with dorsal and ventral regions combined also yielded a significant decrease of 40% which supports the analysis from in situ hybridization densitometry. The distribution of the glycine transporter was characterised with an oligonucleotide probe and showed a specific localisation to motor neurones of the ventral horn, layer II of the dorsal horn and low levels throughout grey matter. In cases of ALS, substantial and selective loss (75%) of the glycine transporter occurred in ventral grey matter ( P < 0.001) with little change occurring in dorsal grey matter. This deficit was also s was also substantiated by slot blot analysis of tissue homogenates where a decrease of 59% was obtained ( P < 0.005). The induction of c- jun and zif-268 detected in ALS spinal cord in this study indicates the presence of a potential cellular response to neuronal ‘stress’ that may play an important role in subsequent neurodegenerative or reparative mechanisms.