Tauopathy in zebrafish using life imaging transgenic zebrafish

Abstract

Axonal transport is a pivotal process for neurons. Transport defects can lead to a deficiency of essential proteins and accumulation of cellular debris at areas distant from the soma, leading to starvation and malfunction of synapses and finally neurodegeneration. Axonal transport deficits have been described in neurodegenerative diseases, including AD and FTD. The Tau protein, which plays a central role in both diseases as hyperphosphorylated constituent of neurofibrillary tangles, is important for regulating microtubules, the intracellular transport tracks. Transport alterations appear to occur early in the cascade of neurological damage, long before neurons die. Therefore, axonal transport represents a promising drug target for neurodegenerative diseases. Mechanisms of axonal transport have been mainly studied in cultured neurons. While this approach has greatly expanded our knowledge of transport phenomena, it is also limited, as in vitro systems neither fully reflect the geometrical complexity of neurons in vivo and their interplay with other cells, nor easily mimic processes related to human disease. More intact preparations derived from invertebrates or mice circumvented some of these problems, but more versatile systems that allow studying axonal transport in vivo, together with screening for modulators, are still needed. We addressed this by developing an in vivo ‘tool box’ to study axonal transport in optically transparent and genetically accessible zebrafish. We have established the characteristics of mitochondrial transport in individually labelled neurons. Furthermore, we generated transgenic ‘mito-fish’ with fluorescently labelled neuronal mitochondria and performed quantitative measurements of mitochondrial density and transport in normal and compound-treated animals. As ‘proof-of-principle’, we crossed the ‘mito-fish’ to a previously developed zebrafish tauopathy-model, which recapitulates key pathological features of FTD, including hyperphosphorylation, tangle formation and cell death (Paquet et al. 2009). Tau-transgenic fish show a prominent reduction in mitochondrial density and movements in peripheral neurons, which can be rescued by over-expression of MARK, a kinase that regulates Tau binding to microtubules. These approaches allow imaging transport of mitochondria in intact animals with high efficiency, and relating transport to the physiology of complete neurons in their natural habitat. Furthermore, ‘mito-fish’ are useful tools to assess mechanisms of transport disruptions and screen for modulating compounds.