Tardigrades and Their Emergence as Model Organisms

Experimentally tractable organisms like C. elegans, Drosophila, zebrafish, and mouse are popular models for addressing diverse questions in biology. In 1997, two of the most valuable invertebrate model organisms to date – C. elegans and Drosophila – were found to be much more closely related to each other than expected. C. elegans and Drosophila belong to the nematodes and arthropods respectively, and these two phyla and six other phyla make up a clade of molting animals referred to as the Ecdysozoa. The other ecdysozoan phyla could be valuable models for comparative biology, taking advantage of the rich and continual sources of research findings as well as tools from both C. elegans and Drosophila. But when the Ecdysozoa was first recognized, few tools were available for laboratory studies in any of these six other ecdysozoan phyla. In 1999 I began an effort to develop tools for studying one such phylum, the tardigrades. Here, I describe how the tardigrade species Hypsibius exemplaris and tardigrades more generally have emerged over the past two decades as valuable new models for answering diverse questions. To date, these questions have included how animal body plans evolve and how biological materials can survive some remarkably extreme conditions.

Caenorhabditis Elegans Exhibits Morphine Addiction-like Behavior via the Opioid-like Receptor NPR-17

Addiction has become a profound societal problem worldwide, and few effective treatments are available. The nematode Caenorhabditis elegans (C. elegans) is an excellent invertebrate model to study neurobiological disease states. C. elegans reportedly developed a preference for cues that had previously been paired with addictive drugs, similar to place conditioning findings in rodents. Moreover, several recent studies discovered and reported the existence of an opioid-like system in C. elegans. Still unclear, however, is whether C. elegans exhibits addictive-like behaviors for opioids, such as morphine. In the present study, we found that C. elegans exhibited dose-dependent preference for morphine using the conditioned chemosensory-cue preference (CCP) test. This preference was blocked by co-treatment with the opioid receptor antagonist naloxone. C. elegans also exhibited aversion to naloxone-precipitated withdrawal from chronic morphine exposure. The expression of morphine-induced CCP and morphine withdrawal were abolished in worms that lacked the opioid-like receptor NPR-17. Dopamine-deficient mutant (cat-2 (e1112)) worms also did not exhibit morphine-induced CCP. These results indicate that the addictive function of the opioid system exists in C. elegans, which may serve as a useful model of opioid addiction.

Functional Conservation and Genetic Divergence of Chordate Glycinergic Neurotransmission: Insights from Amphioxus Glycine Transporters

Glycine is an important neurotransmitter in vertebrates, performing both excitatory and inhibitory actions. Synaptic levels of glycine are tightly controlled by the action of two glycine transporters, GlyT1 and GlyT2, located on the surface of glial cells and neurons, respectively. Only limited information is available on glycinergic neurotransmission in invertebrates, and the evolution of glycinergic neurotransmission is poorly understood. Here, by combining phylogenetic and gene expression analyses, we characterized the glycine transporter complement of amphioxus, an important invertebrate model for studying the evolution of chordates. We show that amphioxus possess three glycine transporter genes. Two of these (GlyT2.1 and GlyT2.2) are closely related to GlyT2 of vertebrates, whereas the third (GlyT) is a member of an ancestral clade of deuterostome glycine transporters. GlyT2.2 expression is predominantly non-neural, whereas GlyT and GlyT2.1 are widely expressed in the amphioxus nervous system and are differentially expressed, respectively, in neurons and glia. Vertebrate glycinergic neurons express GlyT2 and glia GlyT1, suggesting that the evolution of the chordate glycinergic system was accompanied by a paralog-specific inversion of gene expression. Despite this genetic divergence between amphioxus and vertebrates, we found strong evidence for conservation in the role glycinergic neurotransmission plays during larval swimming, the implication being that the neural networks controlling the rhythmic movement of chordate bodies may be homologous.

Novel Pathogenic Mucorales Identified Using the Silkworm Infection Model

Mucormycosis, a rare but highly fatal infection, is caused by fungi of the order Mucorales. Due to their ubiquitous nature, reduced susceptibility to antifungals, acid tolerance, and ability to infect immunocompromised patients through rapid dissemination, these fungi have been frequently reported to infect the COVID-19 patients. In order to develop strategies to overcome mucormycosis, it is essential to understand and identify novel Mucorales present in the environment. In this study, we report the identification of four novel pathogenic Mucorales using the silkworm (Bombyx mori) model. The strains’ phylogeny was analyzed using the genome sequence of the large subunit ribosomal ribonucleic acid (LSU rRNA) and the internal transcribed spacer (ITS) region, where strains 1-3, 5-3, and S286-1101 claded with Mucor orantomantidis, and strain 827-14 claded with Backusella lamprospora. All the strains had a cold-sensitive phenotype with their inability to grow prominently at 4 °C. Mucor sp. 1-3 and 5-3 were characterized by their filamentous and yeast-like growth under aerobic and anaerobic conditions, respectively. The yeast colonies of Mucor sp. 5-3 had multipolar budding cells often observed with cleaved cell surfaces under a scanning electron microscope. We further found that these strains were able to kill immunocompromised mice suggesting their pathogenicity to mammals. Our study established an invertebrate model-based screening system to identify novel pathogenic Mucorales from the natural environment and provided a clue towards the rapid increase in COVID-19 related mucormycosis.

Functional Conservation and Genetic Divergence of Chordate Glycinergic Neurotransmission: Insights from Amphioxus Glycine Transporters

Glycine is an important neurotransmitter in vertebrates, performing both excitatory and inhibitory actions. Synaptic levels of glycine are tightly controlled by the action of two glycine transporters, GlyT1 and GlyT2, located on the surface of glial cells and glycinergic or glutamatergic neurons, respectively. Glycinergic neurotransmission in invertebrates has so far only been investigated in a very limited number of species, and, although it was suggested that its functions are to some extent conserved with vertebrates, the evolution of glycinergic neurotransmission remains very poorly understood. Here, by combining phylogenetic and gene expression analyses, we characterized the glycine transporter complement of amphioxus, an important invertebrate model for studying the evolution of chordates. We show that amphioxus possesses three glycine transporter genes, two of which (GlyT2.1 and GlyT2.2) are closely related to GlyT2 of vertebrates, while the other (GlyT) is a member of an ancestral clade of deuterostome glycine transporters. While expression of GlyT2.2 is predominantly non-neural, GlyT and GlyT2.1 are widely expressed in the amphioxus nervous system and are characterized by differential expression in neurons and glia, respectively. However, in vertebrates, glycinergic neurons express GlyT2 and glia GlyT1, suggesting that the evolution of the chordate glycinergic system was accompanied by complex genetic remodeling leading to the paralog-specific inversion of gene expression. Albeit this genetic divergence between amphioxus and vertebrates, we found strong evidence for a general conservation of the role of glycinergic neurotransmission during larval swimming, allowing us to hypothesize that the neural networks controlling the rhythmic movement of chordate bodies are homologous.

Targeted Propolis-Loaded Poly (Butyl) Cyanoacrylate Nanoparticles: An Alternative Drug Delivery Tool for the Treatment of Cryptococcal Meningitis

In this study, we describe a nano-carrier system for propolis that is able to cross an in vitro model of the blood-brain barrier (BBB) and effectively reduce the virulence of Cryptococcus neoformans in animal models. Antimicrobial properties of propolis have been widely studied. However, propolis applications are limited by its low water solubility and poor bioavailability. Therefore, we recently formulated novel poly (n-butyl cyanoacrylate) nanoparticles (PBCA-NP) containing propolis. PBCA-NP are biocompatible, biodegradable and have been shown to effectively cross the BBB using apolipoprotein E (ApoE) as a ligand. Prepared nanoparticles were characterized for particle size, zeta potential, propolis entrapment efficiency and in vitro release. Additionally, the PBCA-NP were functionalized with polysorbate 80, which then specifically adsorbs ApoE. Using an in vitro BBB model of human brain microvascular endothelial cells hCMEC/D3, it was shown that fluorescence labelled ApoE-functionalized PBCA-NP were internalized by the cells and translocated across the cell monolayer. Propolis-loaded PBCA-NP had in vitro, antifungal activity against C. neoformans, which causes meningitis. To utilize the invertebrate model, Galleria mellonella larvae were infected with C. neoformans and treated with propolis-loaded PBCA-NP. The larvae exhibited normal behavior in toxicity testing, and treatment with propolis-loaded PBCA-NP increased survival in the C. neoformans-infected larvae group. In addition, following cryptococcal infection and then 7 days of treatment, the tissue fungal burden of mice treated with propolis-loaded PBCA-NP was significantly lower than control groups. Therefore, our ApoE-functionalized propolis-loaded PBCA-NP can be deemed as a potential targeted nanoparticle in the therapeutic treatment of cerebral cryptococcosis.

Benefits of Using the Invertebrate Model Organism C. Elegans in an Inquiry-Based Laboratory Activity

Background: The goals of laboratory experiences include developing knowledge base, research skills, and scientific communication abilities. Objective: The aim was to assess an inquiry-based laboratory activity using the model organism Caenorhabditis elegans in relation to learning goals. Method: Students in a Biopsychology laboratory course worked in groups to test the effect of various drugs (e.g., nicotine, ethanol, fluoxetine, and melatonin) on C. elegans behavior. The activity included literature review, experimental design, and a final lab report. A cumulative final exam included a synaptic communication question related to the content of the activity. Results: Students showed better retention of laboratory-related content compared to other topics from the course, as demonstrated through performance on the final exam and were able to replicate previous research demonstrating effects of drug on locomotion. However, students did not improve writing ability compared to performance on a previous American Psychological Association style lab report. Conclusion: This study demonstrates that using a student-designed, multi-week laboratory assignment in an undergraduate Biopsychology course supports the growth of psychology knowledge and the development of research skills. Teaching Implications: Instructors should consider using the described laboratory activity for biopsychology or behavioral neuroscience classes or consider similarly designed laboratory formats for other courses in Psychology.