For evaluation of ultrastructural changes in MNs in the L3–L4 segments of the spinal cord at day 30 of SOD1G93A mice (N = 5, total of 38 MNs) were compared with findings from age-matched WT (N = 5, total of 48 MNs) animals. On the basis of the number of MNs labeled by retrograde transport and motor unit number

estimation (MUNE) results in the literature, we estimate a total of 100 MNs that together compose the TA and soleus motor pools. We analyzed 5–10% of this population at the ultrastructural level. These same MNs were used to evaluate afferent Inhibitors,research,lifescience,medical synaptic input described below and mitochondria number and area. Mitochondria in the MNs identified in the high-resolution images were counted and areas measured using Image J software. Significant differences between WT and SOD1 groups was determined using unpaired t-tests. Glial Inhibitors,research,lifescience,medical cells also identified based on their ultrastructural characteristics (Peters et al. 1991) in the same sections used for analysis of MNs were used for characterization of glial cells. Evaluation of afferent synapses on lumbar MNs For examination

of synapses on MN soma or distal dendrites, using the same segment levels as used for the MNs described above, 25–30 dendrites with synapses in the ventro-lateral white matter were identified and photographed at 16,000×. Inhibitors,research,lifescience,medical Only those synapses with a clear synaptic density and presynaptic vesicles were scored. Synapses were evaluated using thereby classical criteria for identification of Type 1 or R (round), Type 2 or P (pleomorphic), or selleck kinase inhibitor C-type synapses (Bodian 1964, 1966, 1970, 1972; Uchizono 1965, 1966; Hellström et al. 2003). Synapses were defined as having an identifiable synaptic density Inhibitors,research,lifescience,medical and/or fused vesicles at the presynaptic terminal (see Fig. 19 in accompanying paper, doi: 10.1002/brb3.142). Type 1 synapses

(R) have ±50 nmol/L spherical vesicles, light ground substance, and were asymmetrical, with the postsynaptic density and Inhibitors,research,lifescience,medical synaptic cleft being larger than that of other synapse types (see Fig. 19 C and F in accompanying paper). Type 1 synapses are considered excitatory. Type 2 synapses (P) are considered inhibitory and have pleiomorphic or flattened vesicles, GSK-3 lighter symmetrical synaptic densities, and a thin synaptic cleft (see Fig. 19B, E, and F in accompanying paper). Type 3 synapses or C-terminals (C) are only found on αMNs and are cholinergic. These synapses overlie subsynaptic cisterns and organelles (rER), have a distinctive postsynaptic density that appears connected to ER, and have a high packing density of round or slightly flattened vesicles that were not as uniform as those found in Type 1 synapses (Fig. 19A, D, H, and G in accompanying paper). There are at least two other types of synapses (R bulbs or M terminals, and T terminals that exhibit electron dense bodies immediately adjacent to the postsynaptic density called taxi bodies); these synapse types are quite rare (<1%) and were not included in our analysis.