Compartmentalized ocular lymphatic system mediates eye-brain immunity.

The eye, an anatomical extension of the central nervous system (CNS), exhibits many molecular and cellular parallels to the brain. Emerging research demonstrates that changes in the brain are often reflected in the eye, particularly in the retina1. Still, the possibility of an immunological nexus between the posterior eye and the rest of the CNS tissues remains unexplored. Here, studying immune responses to herpes simplex virus in the brain, we observed that intravitreal immunization protects mice against intracranial viral challenge. This protection extended to bacteria and even tumours, allowing therapeutic immune responses against glioblastoma through intravitreal immunization. We further show that the anterior and posterior compartments of the eye have distinct lymphatic drainage systems, with the latter draining to the deep cervical lymph nodes through lymphatic vasculature in the optic nerve sheath. This posterior lymphatic drainage, like that of meningeal lymphatics, could be modulated by the lymphatic stimulator VEGFC. Conversely, we show that inhibition of lymphatic signalling on the optic nerve could overcome a major limitation in gene therapy by diminishing the immune response to adeno-associated virus and ensuring continued efficacy after multiple doses. These results reveal a shared lymphatic circuit able to mount a unified immune response between the posterior eye and the brain, highlighting an understudied immunological feature of the eye and opening up the potential for new therapeutic strategies in ocular and CNS diseases.

Intranasal delivery of SARS-CoV-2 spike protein is sufficient to cause olfactory damage, inflammation and olfactory dysfunction in zebrafish.

Anosmia, loss of smell, is a prevalent symptom of SARS-CoV-2 infection. Anosmia may be explained by several mechanisms driven by infection of non-neuronal cells and damage in the nasal epithelium rather than direct infection of olfactory sensory neurons (OSNs). Previously, we showed that viral proteins are sufficient to cause neuroimmune responses in the teleost olfactory organ (OO). We hypothesize that SARS-CoV-2 spike (S) protein is sufficient to cause olfactory damage and olfactory dysfunction. Using an adult zebrafish model, we report that intranasally delivered SARS-CoV-2 S RBD mostly binds to the non-sensory epithelium of the olfactory organ and causes severe olfactory histopathology characterized by loss of cilia, hemorrhages and edema. Electrophysiological recordings reveal impaired olfactory function to both food and bile odorants in animals treated intranasally with SARS-CoV-2 S RBD. However, no loss of behavioral preference for food was detected in SARS-CoV-2 S RBD treated fish. Single cell RNA-Seq of the adult zebrafish olfactory organ indicated widespread loss of olfactory receptor expression and inflammatory responses in sustentacular, endothelial, and myeloid cell clusters along with reduced numbers of Tregs. Combined, our results demonstrate that intranasal SARS-CoV-2 S RBD is sufficient to cause structural and functional damage to the zebrafish olfactory system. These findings may have implications for intranasally delivered vaccines against SARS-CoV-2.

Olfactory sensory neurons mediate ultrarapid antiviral immune responses in a TrkA-dependent manner.

The nervous system regulates host immunity in complex ways. Vertebrate olfactory sensory neurons (OSNs) are located in direct contact with pathogens; however, OSNs' ability to detect danger and initiate immune responses is unclear. We report that nasal delivery of rhabdoviruses induces apoptosis in crypt OSNs via the interaction of the OSN TrkA receptor with the viral glycoprotein in teleost fish. This signal results in electrical activation of neurons and very rapid proinflammatory responses in the olfactory organ (OO), but dampened inflammation in the olfactory bulb (OB). CD8α+ cells infiltrate the OO within minutes of nasal viral delivery, and TrkA blocking, but not caspase-3 blocking, abrogates this response. Infiltrating CD8α+ cells were TCRαß T cells with a nonconventional phenotype that originated from the microvasculature surrounding the OB and not the periphery. Nasal delivery of viral glycoprotein (G protein) recapitulated the immune responses observed with the whole virus, and antibody blocking of viral G protein abrogated these responses. Ablation of crypt neurons in zebrafish resulted in increased susceptibility to rhabdoviruses. These results indicate a function for OSNs as a first layer of pathogen detection in vertebrates and as orchestrators of nasal-CNS antiviral immune responses.

The axillary lymphoid organ - an external, experimentally accessible immune organ in the zebrafish.

Lymph nodes and other secondary lymphoid organs play critical roles in immune surveillance and immune activation in mammals, but the deep internal locations of these organs make it challenging to image and study them in living animals. Here, we describe a previously uncharacterized external immune organ in the zebrafish ideally suited for studying immune cell dynamics in vivo, the axillary lymphoid organ (ALO). This small, translucent organ has an outer cortex teeming with immune cells, an inner medulla with a mesh-like network of fibroblastic reticular cells along which immune cells migrate, and a network of lymphatic vessels draining to a large adjacent lymph sac. Noninvasive high-resolution imaging of transgenically marked immune cells can be carried out in the lobes of living animals, and the ALO is readily accessible to external treatment. This newly discovered tissue provides a superb model for dynamic live imaging of immune cells and their interaction with pathogens and surrounding tissues, including blood and lymphatic vessels.

Olfactory detection of viruses shapes brain immunity and behavior in zebrafish.

Olfactory sensory neurons (OSNs) are constantly exposed to pathogens, including viruses. However, serious brain infection via the olfactory route rarely occurs. When OSNs detect a virus, they coordinate local antiviral immune responses to stop virus progression to the brain. Despite effective immune control in the olfactory periphery, pathogen-triggered neuronal signals reach the CNS via the olfactory bulb (OB). We hypothesized that neuronal detection of a virus by OSNs initiates neuroimmune responses in the OB that prevent pathogen invasion. Using zebrafish ( Danio rerio ) as a model, we demonstrate viral-specific neuronal activation of OSNs projecting into the OB, indicating that OSNs are electrically activated by viruses. Further, behavioral changes are seen in both adult and larval zebrafish after viral exposure. By profiling the transcription of single cells in the OB after OSNs are exposed to virus, we found that both microglia and neurons enter a protective state. Microglia and macrophage populations in the OB respond within minutes of nasal viral delivery followed decreased expression of neuronal differentiation factors and enrichment of genes in the neuropeptide signaling pathway in neuronal clusters. Pituitary adenylate-cyclase-activating polypeptide ( pacap ), a known antimicrobial, was especially enriched in a neuronal cluster. We confirm that PACAP is antiviral in vitro and that PACAP expression increases in the OB 1 day post-viral treatment. Our work reveals how encounters with viruses in the olfactory periphery shape the vertebrate brain by inducing antimicrobial programs in neurons and by altering host behavior.

Sensing the world and its dangers: An evolutionary perspective in neuroimmunology.

Detecting danger is key to the survival and success of all species. Animal nervous and immune systems cooperate to optimize danger detection. Preceding studies have highlighted the benefits of bringing neurons into the defense game, including regulation of immune responses, wound healing, pathogen control, and survival. Here, we summarize the body of knowledge in neuroimmune communication and assert that neuronal participation in the immune response is deeply beneficial in each step of combating infection, from inception to resolution. Despite the documented tight association between the immune and nervous systems in mammals or invertebrate model organisms, interdependence of these two systems is largely unexplored across metazoans. This review brings a phylogenetic perspective of the nervous and immune systems in the context of danger detection and advocates for the use of non-model organisms to diversify the field of neuroimmunology. We identify key taxa that are ripe for investigation due to the emergence of key evolutionary innovations in their immune and nervous systems. This novel perspective will help define the primordial principles that govern neuroimmune communication across taxa.

High-Resolution, 3D Imaging of the Zebrafish Gill-Associated Lymphoid Tissue (GIALT) Reveals a Novel Lymphoid Structure, the Amphibranchial Lymphoid Tissue.

The zebrafish is extensively used as an animal model for human and fish diseases. However, our understanding of the structural organization of its immune system remains incomplete, especially the mucosa-associated lymphoid tissues (MALTs). Teleost MALTs are commonly perceived as diffuse and scattered populations of immune cells throughout the mucosa. Yet, structured MALTs have been recently discovered in Atlantic salmon (Salmo salar L.), including the interbranchial lymphoid tissue (ILT) in the gills. The existence of the ILT was only recently identified in zebrafish and other fish species, highlighting the need for in-depth characterizations of the gill-associated lymphoid tissue (GIALT) in teleosts. Here, using 3-D high-resolution microscopy, we analyze the GIALT of adult zebrafish with an immuno-histology approach that reveals the organization of lymphoid tissues via the labeling of T/NK cells with an antibody directed to a highly conserved epitope on the kinase ZAP70. We show that the GIALT in zebrafish is distributed over at least five distinct sub-regions, an organization found in all pairs of gill arches. The GIALT is diffuse in the pharyngeal part of the gill arch, the interbranchial septum and the filaments/lamellae, and structured in two sub-regions: the ILT, and a newly discovered lymphoid structure located along each side of the gill arch, which we named the Amphibranchial Lymphoid Tissue (ALT). Based on RAG2 expression, neither the ILT nor the ALT constitute additional thymi. The ALT shares several features with the ILT such as presence of abundant lymphoid cells and myeloid cells embedded in a network of reticulated epithelial cells. Further, the ILT and the ALT are also a site for T/NK cell proliferation. Both ILT and ALT show structural changes after infection with Spring Viraemia of Carp Virus (SVCV). Together, these data suggest that ALT and ILT play an active role in immune responses. Comparative studies show that whereas the ILT seems absent in most neoteleosts ("Percomorphs"), the ALT is widely present in cyprinids, salmonids and neoteleosts, suggesting that it constitutes a conserved tissue involved in the protection of teleosts via the gills.

The mitochondrial genome of the planorbid snail Planorbella duryi.

The complete mitochondrial genome of a freshwater planorbid snail, Planorbella duryi (Mollusca, Gastropoda) was recovered from de novo assembly of genomic sequences generated with the Illumina NextSeq500 platform. The P. duryi mitogenome (14,217 base pairs) is AT rich (72.69%) and comprises 13 protein-coding genes, two ribosomal subunit genes, and 22 transfer RNAs. The gene order is identical to that of Biomphalaria glabrata and other snail species in the family Planorbidae. This is the first full characterization of a mitochondrial genome of the genus Planorbella.