Fast whole-brain imaging of seizures in zebrafish larvae by two-photon light-sheet microscopy

Light-sheet fluorescence microscopy (LSFM) enables real-time whole-brain functional imaging in zebrafish larvae. Conventional one-photon LSFM can however induce undesirable visual stimulation due to the use of visible excitation light. The use of two-photon (2P) excitation, employing near-infrared invisible light, provides unbiased investigation of neuronal circuit dynamics. However, due to the low efficiency of the 2P absorption process, the imaging speed of this technique is typically limited by the signal-to-noise-ratio. Here, we describe a 2P LSFM setup designed for non-invasive imaging that enables quintuplicating state-of-the-art volumetric acquisition rate of the larval zebrafish brain (5 Hz) while keeping low the laser intensity on the specimen. We applied our system to the study of pharmacologically-induced acute seizures, characterizing the spatial-temporal dynamics of pathological activity and describing for the first time the appearance of caudo-rostral ictal waves (CRIWs).     

Emerging Respiratory Viruses of Cats

In recent years, advances in diagnostics and deep sequencing technologies have led to the identification and characterization of novel viruses in cats as protoparviruses and chaphamaparvoviruses, unveiling the diversity of the feline virome in the respiratory tract. Observational, epidemiological and experimental data are necessary to demonstrate firmly if some viruses are able to cause disease, as this information may be confounded by virus- or host-related factors. Also, in recent years, researchers were able to monitor multiple examples of transmission to felids of viruses with high pathogenic potential, such as the influenza virus strains H5N1, H1N1, H7N2, H5N6 and H3N2, and in the late 2019, the human hypervirulent coronavirus SARS-CoV-2. These findings suggest that the study of viral infections always requires a multi-disciplinary approach inspired by the One Health vision. By reviewing the literature, we provide herewith an update on the emerging viruses identified in cats and their potential association with respiratory disease.     

VGLUT‐VGAT expression delineates functionally specialised populations of vasopressin‐containing neurones including a glutamatergic perforant path‐projecting cell group to the hippocampus in rat and mouse brain

The origin and functional significance of vasopressin (AVP)‐containing fibres in limbic regions has been an ongoing subject of investigation for several years. We have previously identified AVP‐magnocellular neurones of rat hypothalamus that provide glutamatergic projections to the hippocampus, amygdala, lateral habenula and locus coeruleus. However, we also reported AVP‐immunopositive fibres in those regions that are thin and make Gray type II synapses, which are unlikely to be of magnocellular origin. Therefore, in the present study, we characterised AVP mRNA co‐expression with expression of mRNAs marking glutamatergic (vesicular glutamate transporter [VGLUT]) and GABAergic (vesicular GABA transporter [VGAT]) neuronal traits in rat and mouse brain, using high‐resolution in situ hybridisation methods, including a radio‐ribonucleotide and RNAscope 2.5 HD duplex assay, with Slc17a7, Slc17a6, Slc32a1 and Avp probes corresponding to mRNAs of VGLUT1, VGLUT2, VGAT and AVP, respectively. We located 18 cell groups expressing Avp and identified their molecular signatures for VGLUT and VGAT mRNA expression. Avp cell groups of hypothalamus and midbrain are mainly VGLUT mRNA‐expressing, whereas those in regions derived from cerebral nuclei are mainly VGAT mRNA‐expressing, suggesting a functional segregation of glutamate/GABA co‐transmission with AVP. A newly identified Slc17a7 and Slc17a6 (but not Slc32a1) expressing vasopressinergic cell group was found in layer II‐III neurones of the central entorhinal cortex, which projects to the hippocampus. These data support the notion of a complex role for AVP with respect to modulating multiple central circuits controlling behaviour in specific ways depending on co‐transmission with glutamate or GABA, potentially giving rise to a functional classification of AVPergic neurones in the central nervous system.     

Innate immunity in cutaneous melanoma

The skin immune system is composed of a vast network of immune cells, including lymphocytes, macrophages, neutrophils, dendritic cells and Langerhans cells, which not only are involved in inflammatory responses but also contribute to homeostatic function and may participate in the various steps of carcinogenesis. Many studies support the notion that innate immunity has a key role in the development, growth and prognosis of cutaneous malignant melanoma (MM), through the release of pro‐ and/or anti‐inflammatory cytokines and tumour growth factors. The tumour environment in a major subset of cutaneous MM shows evidence of a T cell‐infiltrated phenotype, but there is less known about the presence and the phenotype of other immune system cells. Response to immunotherapy is largely correlated with the presence of T cells in the tumour microenvironment, while the regulation exerted by stromal components such as macrophages and mast cells has been less investigated. In the current report, we review the recent literature, focusing our attention on the role of macrophages, dendritic cells, mast cells and natural killer cells in orchestrating MM progression, to better understand tumour immunobiology. The identification of new therapeutic targets and the application of approaches aimed at modulating crosstalk between immune and tumour cells, could have a crucial impact on immunotherapy and result in better clinical outcome. We hope this review will be helpful in cutaneous MM research.     

Epstein–Barr virus and skin manifestations in childhood

Epstein–Barr virus (EBV) is a human B‐lymphotropic herpes virus and one of the most common viruses in humans. Specific skin signs related to EBV infection are the exanthem of mononucleosis, which is observed more frequently after ingestion of amoxicillin, and oral hairy leukoplakia, a disease occurring mostly in immunocompromised subjects with HIV infection. Other more uncommon cutaneous disorders that have been associated with EBV infection include virus‐related exanthems or diseases such as Gianotti–Crosti syndrome, erythema multiforme, and acute genital ulcers. Other skin manifestations, not correlated to virus infection, such as hydroa vacciniforme and drug‐induced hypersensitivity syndrome have also been linked to EBV. The putative involvement of EBV in skin diseases is growing similarly to other areas of medicine, where the role of EBV infection is being investigated in potentially debilitating inflammatory diseases. The prognosis of EBV infection in healthy, immunocompetent individuals is excellent. However, lifelong infection, which is kept in check by the host immune system, determines an unpredictable risk of pathologic unpredictable scenarios. In this review, we describe the spectrum of non‐tumoral dermatological manifestations that can follow EBV primary infection or reactivation of EBV in childhood.     

Rosetting of human T lymphocytes with goat red blood cells: Effect of treatment with 2-aminoethylisothiouronium bromide (AET) and comparison with AET treated sheep red blood cells

In vitro treatment of goat red blood cells (GRBCs) with the sulphydryl compound 2-aminoethylisothiouronium bromide (AET) increases their specific reactivity with human T lymphocytes without affecting the specificity of the reaction. AET-GRBCs bind to only part of T lymphocytes rosetting with AET-sheep red blood cells (SRBCs): the receptors for both types of RBCs are very similar if not identical, but display higher affinity for AET-SRBCs than for AET-GRBCs. Rosetting of T lymphocytes with AET-GRBCs may be useful to enumerate T lymphocyte subsets in patients with abnormality of the immune system and to fractionate T lymphocyte subpopulations.     

Effectiveness of extracorporeal shock wave therapy in bone marrow edema syndrome of the hip

There is no gold standard for treatment of bone marrow edema syndrome of the hip (BMESH). Usually, treatment is conservative, owing to the favorable and self-limiting prognosis. In musculoskeletal disorders, the effectiveness of extracorporeal shock wave therapy (ESWT) has been widely recognized and recent research supports its use in the treatment of the first stages of avascular osteonecrosis of the proximal femur and in other conditions where bone marrow edema is present. On this basis, we performed a prospective study to evaluate the effectiveness of ESWT in normalizing the symptoms and imaging features of BMESH. Twenty consecutive symptomatic patients underwent two treatments of high-energy ESWT and were followed-up at 2, 3 and 6 months, with a final clinical follow-up at mean 15.52 ± 1.91 months. Patients underwent magnetic resonance imaging of the hip and were evaluated according to the Harris hip score. The mean improvement in HHS over the course of the study was of 58.5 ± 14.9 points (p < 0.0001), and the mean edema area reduced from 981.9 ± 453.2 mm² pre-treatment to 107.8 ± 248.1 mm² at 6 months. ESWT seems to be a powerful, non-pharmacological tool that produces rapid pain relief and functional improvement and aids the normalization of the vascular and metabolic impairments which characterize BMESH.