AAV vector-mediated secretion of chondroitinase provides a sensitive tracer for axonal arborisations

Alves, João Nuno, Muir, Elizabeth M, Andrews, Melissa R, Ward, Anneliese, Michelmore, Nicholas, Dasgupta, Debayan, Verhaagen, Joost, Moloney, Elizabeth B, Keynes, Roger J, Fawcett, James W and Rogers, John H (2014) AAV vector-mediated secretion of chondroitinase provides a sensitive tracer for axonal arborisations. Journal of Neuroscience Methods, 227, 107-20. (doi:10.1016/j.jneumeth.2014.02.010).

Abstract

As part of a project to express chondroitinase ABC (ChABC) in neurons of the central nervous system, we have inserted a modified ChABC gene into an adeno-associated viral (AAV) vector and injected it into the vibrissal motor cortex in adult rats to determine the extent and distribution of expression of the enzyme. A similar vector for expression of green fluorescent protein (GFP) was injected into the same location. For each vector, two versions with minor differences were used, giving similar results. After 4 weeks, the brains were stained to show GFP and products of chondroitinase digestion. Chondroitinase was widely expressed, and the AAV-ChABC and AAV-GFP vectors gave similar expression patterns in many respects, consistent with the known projections from the directly transduced neurons in vibrissal motor cortex and adjacent cingulate cortex. In addition, diffusion of vector to deeper neuronal populations led to labelling of remote projection fields which was much more extensive with AAV-ChABC than with AAV-GFP. The most notable of these populations are inferred to be neurons of cortical layer 6, projecting widely in the thalamus, and neurons of the anterior pole of the hippocampus, projecting through most of the hippocampus. We conclude that, whereas GFP does not label the thinnest axonal branches of some neuronal types, chondroitinase is efficiently secreted from these arborisations and enables their extent to be sensitively visualised. After 12 weeks, chondroitinase expression was undiminished.

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Contributors

Author: Melissa R Andrews

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