Time course Analysis of Gene expression patterns in ZebrafIsh Eye during Optic Nerve Regeneration

Abstract

It is well-established that neurons in the adult mammalian central nervous system (CNS) are terminally differentiated and, if injured, will be unable to regenerate their connections. In contrast to mammals, zebrafish and other teleosts display a robust neuroregenerative response. Following optic nerve crush (ONX), retinal ganglion cells (RGC) regrow their axons to synapse with topographically correct targets in the optic tectum, such that vision is restored in ∼21 days. What accounts for these differences between teleostean and mammalian responses to neural injury is not fully understood. A time course analysis of global gene expression patterns in the zebrafish eye after ONX can help to elucidate cellular and molecular mechanisms that contribute to a successful neuroregeneration. To define different phases of regeneration after ONX, alpha tubulin 1 (tuba1) and growth-associated protein 43 (gap43), markers previously shown to correspond to morphophological events, were measured by real time quantitative PCR (qPCR). Microarray analysis was then performed at defined intervals (6 hours, 1, 4, 12, and 21 days) post-ONX and compared to SHAM. Results show that optic nerve damage induces multiple, phase-related transcriptional programs, with the maximum number of genes changed and highest fold-change occurring at 4 days. Several functional groups affected by optic nerve regeneration, including cell adhesion, apoptosis, cell cycle, energy metabolism, ion channel activity, and calcium signaling, were identified. Utilizing the whole eye allowed us to identify signaling contributions from the vitreous, immune and glial cells as well as the neural cells of the retina. Comparisons between our dataset and transcriptional profiles from other models of regeneration in zebrafish retina, heart and fin revealed a subset of commonly regulated transcripts, indicating shared mechanisms in different regenerating tissues. Knowledge of gene expression patterns in all components of the eye in a model of successful regeneration provides an entry point for functional analyses, and will help in devising hypotheses for testing normal and toxic regulatory factors.

The scales and tales of myelination: using zebrafish and mouse to study myelinating glia

“…heart and fin), represent an excellent vertebrate model organism for studying regeneration (Shi et al 2015). Recently, several groups have taken advantage of the elevated regenerative capacity of zebrafish to understand repair after injury in the nervous system (McCurley et al, 2010; Ma et al 2012; Moore et al, 2012; Münzel et al, 2012; Chung et al 2013). In the PNS of zebrafish and mouse, axons are readily remyelinated by SCs after injury (Wang and Jin, 2011; Moore et al, 2012; Jessen et al, 2015).…”

A transgenic zebrafish model of targeted oocyte ablation and de novo oogenesis

“…The robust capacity of the zebrafish (Danio rerio) to regenerate fins, neural tissues (retinal cells, optic nerves, spinal cord), and cardiac muscle has led to the widespread use of this model species for tissue and organ regeneration studies (Chablais and Jazwinska, 2010;Jopling et al, 2010;McCurley and Callard, 2010;Cameron, 2000;Poss et al, 2002Poss et al, , 2003Tawk et al, 2002;Lien et al, 2006;Sherpa et al, 2008). The innate regenerative capability of this species, combined with its relative ease of genetic manipulation, has consequently led to many novel insights into the molecular and cellular regulation of tissue regeneration (Curado et al, 2007(Curado et al, , 2008Davison et al, 2007;Pisharath, 2007;Pisharath et al, 2007;Moss et al, 2009;Pisharath and Parsons, 2009;Zhao et al, 2009).…”

Complementary research models and methods to study axonal regeneration in the vertebrate retinofugal system

“…In the first model, the nerve is cut with scissors, thereby creating a proximal and distal nerve stump that are physically separated from each other. In contrast, in the ONC model, the nerve is crushed with forceps to separate the axons, but leaving the connective tissue around them intact, and thus holding the proximal and distal ends together (Becker et al 2000;McCurley and Callard 2010;Fleisch et al 2011;Lemmens et al 2015;Van Houcke et al 2017). As expected, the ONT model is somewhat more drastic and regeneration progresses slower as compared to ONC (Zou et al 2013).…”

“…Correct tectal reinnervation includes restoration of the retinotopic map, which is achieved around 6 weeks after the optic nerve lesion Becker 2007, 2014). Complete visual repair, including complex behaviors with visual aspects such as chasing and shoaling, is restored at 60-100 dpi (Bormann et al 1998;Hieber et al 1998;Becker 2007, 2014;McCurley and Callard 2010;Kato et al 2013).…”

“…Indeed, both tuba1a and gap43 have long been recognized as a hallmark for axonal growth during CNS development in vertebrates, while their expression declines drastically upon CNS maturation. However, in neurons with regenerative potential, such as zebrafish RGCs after optic nerve injury, they are re-induced throughout the regeneration process, which could thus be followed by keeping track of gap43 or tuba1a expression, as was discussed in section "Imaging modalities to evaluate optic nerve regeneration" (Hieber et al 1998;Goldman et al 2001;Kaneda et al 2008;Udvadia 2008;Kusik et al 2010;McCurley and Callard 2010;Diekmann et al 2015b).…”