Regional and Ultrastructural Immunolocalization of Copper-Zinc Superoxide Dismutase in Rat Central Nervous System

Abstract

SUMMARYWe examined the distribution of copper-zinc superoxide dismutase (CuZn-SOD) in adult rat central nervous system by light and electron microscopic immunocytochemistry, using an affinity-purified polyclonal antibody. The enzyme appeared to be exclusively localized in neurons. No immunoreactivity was seen in non-neuronal cells. The staining intensity was variable, depending on the brain region and, within the same region, on the neuron type. Highly immunoreactive elements included cortical neurons evenly distributed in the different layers, hippocampal interneurons, neurons of the reticular thalamic nucleus, and Golgi, stellate, and basket cells of the cerebellar cortex. Other neurons, i.e., pyramidal cells of the neocortex and hippocampus, Purkinje and granule cells of the cerebellar cortex, and the majority of thalamic neurons, showed much weaker staining. In the spinal cord, intense CuZnSOD immunoreactivity was present in many neurons, including motor neurons. Pre-embedding immunoelectron microscopy of the neocortex, hippocampus, reticular thalamic nucleus, and cerebellar cortex showed cytosolic and nucleoplasmic labeling. Moreover, single membrane-limited immunoreactive organelles identified as peroxisomes were often found, even in neurons that appeared weakly stained at the light microscopic level. In double immunogold labeling experiments, particulate CuZn-SOD immunoreactivity co-localized with catalase, a marker enzyme for peroxisomes, thus demonstrating that in neural tissue CuZnSOD is also present in peroxisomes.

Fewer active motors per vesicle may explain slowed vesicle transport in chick motoneurons after three days in vitro

“…MEDIC is described in more detail elsewhere (Hill et al 2004;Hill, Macosko and Holzwarth, in preparation). Using the MEDIC technique 0.1 µm diameter polystyrene beads moving 1 µm/s are at the limit of detectability, thus likely candidates for the vesicular organelles that we observe in this paper are mitochondria (Friberg et al 1998), lysosomes (Bednarski et al 1997), peroxisomes (Moreno et al 1997), and phagosomes (Martin et al 1999)-all of which are larger than 0.1 µm in neuronal cells.…”

The Copper Chaperone CCS Directly Interacts with Copper/Zinc Superoxide Dismutase

“…Sequence analysis of CCS revealed a potential peroxisomal localization sequence in the carboxyl terminus (Fig. 1), and although recent studies have suggested that in neurons SOD1 is localized in part to peroxisomes (32), no overlap was observed for either SOD1 or CCS when double-immunofluorescent studies were performed in these same cells utilizing a sheep anti-human catalase antibody that identified peroxisomes (data not shown).…”

Quantitative Histochemical Assay for Superoxide Dismutase in Rat Brain

“…At present it is possible to localize and quantify the amount of SOD enzyme immunohistochemically by use of specific antibodies, but not its activity (Moreno et al 1997).…”