Amyotrophic lateral sclerosis (ALS) is a lethal neurodegenerative disease in which motoneurons (MN) in the nervous system die. The study of ALS, namely the main MN disease in the adulthood, has been hampered by the absence, for a long time, of available animal models. The discovery of mutation in the gene for the cytosolic Cu/Zn super-oxide dismutase in 20% of familiar ALS patients led to an animal model in which the human mutant SOD1 is over expressed in mice (G93A). These transgenic mice develop limb paralysis associated with loss of spinal motor neurons (Gurney et al 1994). The development of new in vitro models of SOD1 mutations is instrumental to investigate the mechanisms of MN death associated with this gene defect. We investigated the interactions between mutations associated to ALS and the spinal micro-environment, focusing on early functional markers of disease. In particular we monitored functional aspects, such as spinal network synaptic activity in cultured models. We employed the long term spinal cord organotypic culture developed from G93A embryonic mice and their wild type (WT) littermates. We previously reported in such cultures (Avossa et al., 2006) an increase susceptibility of MN to excitotoxic stimuli and the presence of a discrete synaptic rearrangement in G93A ventral cord during the second week in vitro (Avossa et al., 2006). Based on these results, we decided to characterize the spinal network activity, via interneuron patch clamp recordings (voltage clamp) at several time points of in vitro growth, of both WT and G93A. During the first week in vitro, both WT and G93A recorded interneurons, displayed spontaneous bursting. At later stages of in vitro growth, spontaneous synaptic activity was characterised by a large increase in PSCs frequency. In the majority of ventral interneurons we did not detect spontaneous population bursts similar to those recorded at one week, although PSCs temporal summation often led to the appearance of clusters of synaptic currents. In the presence of CNQX (10 mM) inhibitory post synaptic currents (IPSC) and mIPSC (during co-application of CNQX and TTX, 1 mM) displayed a decrease in frequency in the G93A samples when compared to WT ones. Within the temporal window of 2 and 3 weeks in culture, glycinergic PSC underwent a different development profile in G93A interneurons, displaying a mean decay time constant (t) which was significantly faster when compared to that detected in control WT neurons at similar age in vitro.

Gurney et al., 1994 Science 264:1772-1775.

Avossa et al., 2006 Neuroscience, 138: 1179-1194.