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In addition to a fast activating and immediately inactivating inward sodium current, many types of excitable cells possess a noninactivating or slowly inactivating component: the persistent sodium current (INaP). The INaP is found in normal primary sensory neurons where it is mediated by tetrodotoxin-sensitive sodium channels. The dorsal root ganglion (DRG) is the gateway for ectopic impulses that originate in pathological pain signals from the periphery. However, the role of INaP in DRG neurons remains unclear, particularly in neuropathic pain states. Using in vivo recordings from single medium- and large-diameter fibers isolated from the compressed DRG in Sprague-Dawley rats, we show that local application of riluzole, which blocks the INaP, also inhibits the spontaneous activity of A-type DRG neurons in a dose-dependent manner. Significantly, riluzole also abolished subthreshold membrane potential oscillations (SMPOs), although DRG neurons still responded to intracellular current injection with a single full-sized spike. In addition, the INaP was enhanced in medium- and large-sized neurons of the compressed DRG, while bath-applied riluzole significantly inhibited the INaP without affecting the transient sodium current (INaT). Taken together, these results demonstrate for the first time that the INaP blocker riluzole selectively inhibits INaP and thereby blocks SMPOs and the ectopic spontaneous activity of injured A-type DRG neurons. This suggests that the INaP of DRG neurons is a potential target for treating neuropathic pain at the peripheral level.

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Cathelicidin antimicrobial peptides (CAMPs) represent a crucial component of the innate immune system in vertebrates. Although widely studied in mammals, little is known about the structure and function of fish CAMPs. Further to the previous findings, two more cathelicidin genes and multiple transcripts from rainbow trout were identified in the present study. Interestingly, we found that trout have evolved energy-saving forms of cathelicidins with the total deletion of the characteristic cathelin-like domain. Sequence analysis revealed that salmonid CAMPs have formed a special class of antimicrobial peptides in vertebrates with three distinctive hallmarks: the N terminus is intensified by positive charges, the central region consists of repetitive motifs based on RPGGGS, and the C terminus is lowly charged. Immunofluorescence localization of trout CAMPs demonstrated that these peptides expressed mainly at the mucosal layer of gut. Meanwhile, signals around sinusoids were also detected in head kidney. Moreover, the biological activities of trout CAMPs were proved to be mediated by the N terminus. Additionally, the repetitive motifs characteristically existing in Salmonidae increased the structural flexibilities of peptides and further increased the antibacterial and IL-8–stimulating activities. Unlike most α helical and cytotoxic mammalian CAMPs, trout CAMPs, mainly consisting of β-sheet and random coil, exhibited no cytotoxic activities. The distinctive structural features of trout CAMPs provide new insights into the understanding of the evolution of CAMPs in vertebrates. Moreover, the high bacterial membrane selectivity of trout CAMPs will help to design excellent peptide antibiotics.

The gut-associated lymphoid tissue, connected with liver via bile and blood, constructs a local immune environment of both defense and tolerance. The gut-liver immunity has been well-studied in mammals, yet in fish remains largely unknown, even though enteritis as well as liver and gallbladder syndrome emerged as a limitation in aquaculture. In this study, we performed integrative bioinformatic analysis for both transcriptomic (gut and liver) and proteomic (intestinal mucus and bile) data, in both healthy and infected tilapias. We found more categories of immune transcripts in gut than liver, as well as more adaptive immune in gut meanwhile more innate in liver. Interestingly reduced differential immune transcripts between gut and liver upon inflammation were also revealed. In addition, more immune proteins in bile than intestinal mucus were identified. And bile probably providing immune effectors to intestinal mucus upon inflammation was deduced. Specifically, many key immune transcripts in gut or liver as well as key immune proteins in mucus or bile were demonstrated. Accordingly, we proposed a hypothesized profile of fish gut-liver immunity, during either homeostasis or inflammation. Current data suggested that fish gut and liver may collaborate immunologically while keep homeostasis using own strategies, including potential unique mechanisms.

For a virus to replicate efficiently, it must try and inhibit host IFN expression because IFN is an important host defense at early stages after viral infection. For aquatic viruses, the mechanisms used to escape the hosts IFN system are still unclear. In this study, we show that the N protein of spring viremia of carp virus (SVCV) inhibits zebrafish IFNφ1 production by degrading the mitochondrial antiviral signaling protein (MAVS). First, the upregulation of IFNφ1 promoter activity stimulated by polyinosinic:polycytidylic acid, retinoic acid–inducible gene I (RIG-I) or MAVS was suppressed by the SVCV infection. However, the upregulation by the downstream factor of the RIG-I–like receptor signaling pathway, TANK-binding kinase 1, was not affected. Notably, at the protein level, MAVS decreased remarkably when cells were infected with SVCV. Second, consistent with the result of the SVCV infection, overexpression of the N protein of SVCV blocked the IFNφ1 transcription activated by MAVS and downregulated MAVS expression at the protein level but not at the mRNA level. Further analysis demonstrated that the N protein targeted MAVS for K48-linked ubiquitination, which promoted the degradation of MAVS. These data indicated that fish MAVS could be degraded by the N protein of SVCV through the ubiquitin-proteasome pathway. To our knowledge, this is the first article of a fish RIG-I–like receptor pathway interfered by an aquatic virus in an ubiquitin-proteasome manner, suggesting that immune evasion of a virus also exists in lower vertebrates.

As the teleost specific immunoglobulin, IgT plays important roles in systemic and mucosal immunity. In the current study, in rainbow trout, we have cloned the heavy chain (Igτ) genes of a secretory form of IgT2 as well as the membrane and secretory forms of a third IgT subclass, termed IgT3. Conserved cysteine and tryptophan residues that are crucial for the folding of the immunoglobulin domain as well as hydrophobic and hydrophilic residues within CART motif were identified in all IgT subclasses. Through analysis of the rainbow trout genome assembly, Igτ3 gene was found localized upstream of Igτ1 gene, while Igτ2 gene situated on another scaffold. At the transcriptional level, Igτ1 was mainly expressed in both systemic and mucosal lymphoid tissues, while Igτ2 was largely expressed in systemic lymphoid organs. After LPS and poly (I:C) treatment, Igτ1 and Igτ2 genes exhibited different expression profiles. Interestingly the transcriptional level of Igτ3 was negligible, although its protein product could be identified in trout serum. Importantly, a previously reported monoclonal antibody directed against trout IgT1 was able to recognize IgT2 and IgT3. These data demonstrate that there exist three subclasses of IgT in rainbow trout, and that their heavy chain genes display different expression patterns during stimulation. Overall, our data reflect the diversity and complexity of immunoglobulin in trout, thus provide a better understanding of the IgT system in the immune response of teleost fish.

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