Abstract
Peripheral glia are known to have a critical role in the initial response to axon damage and degeneration. However, little is known about the cellular responses of non-myelinating glia to nerve injury. In this study, we analyzed the transcriptomes of wild-type and mutant (lacking peripheral glia) zebrafish larvae that were treated with metronidazole. This treatment allowed us to conditionally and selectively ablate cranial sensory neurons whose axons are ensheathed only by non-myelinating glia. While transcripts representing over 27,000 genes were detected by RNAseq, only a small fraction (~1% of genes) were found to be differentially expressed in response to neuronal degeneration in either line at either 2 hrs or 5 hrs of metronidazole treatment. Analysis revealed that most expression changes (332 out of the total of 458 differentially expressed genes) occurred over a continuous period (from 2 to 5 hrs of metronidazole exposure), with a small number of genes showing changes limited to only the 2 hr (55 genes) or 5 hr (71 genes) time points. For genes with continuous alterations in expression, some of the most meaningful sets of enriched categories in the wild-type line were those involving the inflammatory TNF-alpha and IL6 signaling pathways, oxidoreductase activities and response to stress. Intriguingly, these changes were not observed in the mutant line. Indeed, cluster analysis indicated that the effects of metronidazole treatment on gene expression was heavily influenced by the presence or absence of glia, indicating that the peripheral non-myelinating glia play a significant role in the transcriptional response to sensory neuron degeneration. This is the first transcriptome study of metronidazole-induced neuronal death in zebrafish and the response of non-myelinating glia to sensory neuron degeneration. We believe this study provides important insight into the mechanisms by which non-myelinating glia react to neuronal death and degeneration in sensory circuits.
Highlights
In the peripheral nervous system, damage to and/or loss of sensory neurons can result in debilitating neuropathies [1,2,3,4,5,6,7] that often have a dramatic impact on quality of life
Investigations into the pathophysiology of neuronal damage/death typically have utilized experimental paradigms involving non-selective damage to peripheral nerves; e.g., constricting, crushing or cutting nerves [8,9,10] or using non-targeted pharmacological agents [11, 12]. Such studies have shown that peripheral glia play critical roles in both the degenerative and regenerative processes that are involved in the responses to peripheral nerve damage [13,14,15]
Our working hypothesis is that alterations in gene expression underlie the morphological responses that are seen with metronidazole (MET)-induced cranial neuron ablation and axon degeneration in this transgenic line
Summary
In the peripheral nervous system, damage to and/or loss of sensory neurons can result in debilitating neuropathies [1,2,3,4,5,6,7] that often have a dramatic impact on quality of life. GO analysis of the late response DEGs (a total of 71 genes) revealed that as the larvae were exposed to MET for longer times (5 hrs), new sets of DEGs were identified that revealed enrichment in new categories such as those involved in metabolism (cers2, mgst1, gclm, sgpl1, gls, papss2, sqrdl, cyp46a1, g6pd, ptgs2, ggt1 and slc5a1), transcriptional regulation (cebpb, cebpd, bach1, epas1, cers2, junb, maff and stat3), glutathione metabolism (mgst1, gclm, g6pd and ggt1) and IL-6/TNF signaling pathways (sele, cebpb, cebpd, stat3, junb, creb3l3, tnfrsf1a and il6st) (Fig 5 and S7 Table).
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