In vivo Effects of Salicornia herbacea and Calystegia soldanella Extracts for Memory Improvement.

The global elderly population, aged 65 and over, reached approximately 10% in 2020, and this proportion is expected to continue rising. Therefore, the prevalence of neurodegenerative diseases such as Parkinson's disease (PD), which are characterized by declining memory capabilities, is anticipated to increase. In a previous study, we successfully restored the diminished memory capabilities in a fruit fly model of PD by administering an omija extract. To identify functional ingredients that can enhance memory akin to the effects of the omija extract, we conducted screenings by administering halophyte extracts to the PD model. Halophytes are plants that thrive in high-salt environments, and given Korea's geographic proximity to the sea on three sides, it serves as an optimal hub for the utilization of these plants. Upon examining the effects of the oral administration of 12 halophyte extracts, Salicornia herbacea and Calystegia soldanella emerged as potential candidates for ameliorating memory loss in PD patients. Moreover, our findings suggested that C. soldanella, but not S. herbacea, can mitigate oxidative stress in DJ-1ß mutants.

A single pair of pharyngeal neurons functions as a commander to reject high salt in Drosophila melanogaster.

Salt (NaCl), is an essential nutrient for survival, while excessive salt can be detrimental. In the fruit fly, Drosophila melanogaster, internal taste organs in the pharynx are critical gatekeepers impacting the decision to accept or reject a food. Currently, our understanding of the mechanism through which pharyngeal gustatory receptor neurons (GRNs) sense high salt are rudimentary. Here, we found that a member of the ionotropic receptor family, Ir60b, is expressed exclusively in a pair of GRNs activated by high salt. Using a two-way choice assay (DrosoX) to measure ingestion volume, we demonstrate that IR60b and two co-receptors IR25a and IR76b are required to prevent high salt consumption. Mutants lacking external taste organs but retaining the internal taste organs in the pharynx exhibit much higher salt avoidance than flies with all taste organs but missing the three IRs. Our findings highlight the vital role for IRs in a pharyngeal GRN to control ingestion of high salt.

A single pair of pharyngeal neurons functions as a commander to reject high salt in Drosophila melanogaster.

Salt is an essential nutrient for survival, while excessive NaCl can be detrimental. In the fruit fly, Drosophila melanogaster, internal taste organs in the pharynx are critical gatekeepers impacting the decision to accept or reject a food. Currently, our understanding of the mechanism through which pharyngeal gustatory receptor neurons (GRNs) sense high salt are rudimentary. Here, we found that a member of the ionotropic receptor family, Ir60b, is expressed exclusively in a pair of GRNs activated by high salt. Using a two-way choice assay (DrosoX) to measure ingestion volume, we demonstrate that IR60b and two coreceptors IR25a and IR76b, are required to prevent high salt consumption. Mutants lacking external taste organs but retaining the internal taste organs in the pharynx exhibit much higher salt avoidance than flies with all taste organs but missing the three IRs. Our findings highlight the vital role for IRs in a pharyngeal GRN to control ingestion of high salt.

Protocol for binary food choice assays using Drosophila melanogaster.

Food preference is a fundamental behavior for animals to choose nutritious foods while rejecting foods containing toxins. Here, we describe binary food choice assays using Drosophila melanogaster, which are straightforward approaches for the characterization of two-way choice tastants. We detail the preparation of flies and dye-containing food, followed by the binary-choice feeding assays and the determination of the preference index (PI). This protocol is simple, sensitive, and reproducible in qualitatively detecting attractive or aversive characteristics toward any two-way choice tastants. For complete details on the use and execution of this protocol, please refer to Aryal et al. (2022).

Ionotropic receptors mediate nitrogenous waste avoidance in Drosophila melanogaster.

Ammonia and its amine-containing derivatives are widely found in natural decomposition byproducts. Here, we conducted biased chemoreceptor screening to investigate the mechanisms by which different concentrations of ammonium salt, urea, and putrescine in rotten fruits affect feeding and oviposition behavior. We identified three ionotropic receptors, including the two broadly required IR25a and IR76b receptors, as well as the narrowly tuned IR51b receptor. These three IRs were fundamental in eliciting avoidance against nitrogenous waste products, which is mediated by bitter-sensing gustatory receptor neurons (GRNs). The aversion of nitrogenous wastes was evaluated by the cellular requirement by expressing Kir2.1 and behavioral recoveries of the mutants in bitter-sensing GRNs. Furthermore, by conducting electrophysiology assays, we confirmed that ammonia compounds are aversive in taste as they directly activated bitter-sensing GRNs. Therefore, our findings provide insights into the ecological roles of IRs as a means to detect and avoid toxic nitrogenous waste products in nature.

Cucurbitacin B Suppresses Hyperglycemia Associated with a High Sugar Diet and Promotes Sleep in Drosophila melanogaster.

Secondary metabolites enable plants to protect themselves from herbivorous insects. Among these, cucurbitacin B (cuc-B) is a bitter-tasting compound with promising pharmacological potential. Dietary exposure to cuc-B lowered the hemolymph glucose levels of Drosophila melanogaster fed with a high carbohydrate diet, which is homologous to high blood glucose in humans, and its effect was comparable to that of metformin, a well-known glucose-lowering drug. Furthermore, cuc-B reduced tissue sugar levels and glycogen levels, as well as triacylglycerol levels. Our results thus highlight the potential applicability of this compound to treat chronic metabolic diseases such as diabetes and obesity. Additionally, we analyzed sleep quality and taste-associative memory enhancement after cuc-B and metformin treatment. Both supplements increased nighttime bout length and metformin increased memory consolidation. Therefore, discarded shell of Cucurbitaceae could be processed into health supplements.

Critical enzymes for biosynthesis of cucurbitacin derivatives in watermelon and their biological significance.

Various cucurbitacins have been isolated, and their structures have been elucidated. Owing to their economic potential and importance as active pharmacological compounds, their cytotoxicity in various cancer cells has been assessed. Here, we mined several candidate genes with potential involvement in cucurbitacin biosynthesis in watermelon (Citrullus lanatus) and performed in vitro enzymatic assays and instrumental analyses using various substrates to identify cucurbitacin functions and products. Enzymatic activities of two acetyltransferases (ACTs) and one UDP-glucosyltransferase (UGT) against cucurbitacins were confirmed, resulting in the synthesis of novel cucurbitacins in vivo and/or in vitro to our knowledge. As ACTs and UGT are involved in the dynamic conversion of cucurbitacins by catalyzing acetylation and glucosylation at moieties in the cucurbitacins skeleton, these findings improve our knowledge on how these genes contribute to the diversity of cucurbitacins.

Cucurbitacin B Activates Bitter-Sensing Gustatory Receptor Neurons via Gustatory Receptor 33a in Drosophila melanogaster.

The Gustatory system enables animals to detect toxic bitter chemicals, which is critical for insects to survive food induced toxicity. Cucurbitacin is widely present in plants such as cucumber and gourds that acts as an anti-herbivore chemical and an insecticide. Cucurbitacin has a harmful effect on insect larvae as well. Although various beneficial effects of cucurbitacin such as alleviating hyperglycemia have also been documented, it is not clear what kinds of molecular sensors are required to detect cucurbitacin in nature. Cucurbitacin B, a major bitter component of bitter melon, was applied to induce action potentials from sensilla of a mouth part of the fly, labellum. Here we identify that only Gr33a is required for activating bitter-sensing gustatory receptor neurons by cucurbitacin B among available 26 Grs, 23 Irs, 11 Trp mutants, and 26 Gr-RNAi lines. We further investigated the difference between control and Gr33a mutant by analyzing binary food choice assay. We also measured toxic effect of Cucurbitacin B over 0.01 mM range. Our findings uncover the molecular sensor of cucurbitacin B in Drosophila melanogaster. We propose that the discarded shell of Cucurbitaceae can be developed to make a new insecticide.

Mechanism of Acetic Acid Gustatory Repulsion in Drosophila.

The decision to consume or reject a food based on the degree of acidity is critical for animal survival. However, the gustatory receptors that detect sour compounds and influence feeding behavior have been elusive. Here, using the fly, Drosophila melanogaster, we reveal that a member of the ionotropic receptor family, IR7a, is essential for rejecting foods laced with high levels of acetic acid. IR7a is dispensable for repulsion of other acidic compounds, indicating that the gustatory sensation of acids occurs through a repertoire rather than a single receptor. The fly's main taste organ, the labellum, is decorated with bristles that house dendrites of gustatory receptor neurons (GRNs). IR7a is expressed in a subset of bitter GRNs rather than GRNs dedicated to sour taste. Our findings indicate that flies taste acids through a repertoire of receptors, enabling them to discriminate foods on the basis of acid composition rather than just pH.

Gustatory receptor 28b is necessary for avoiding saponin in Drosophila melanogaster.

Saponins function as a natural self-defense mechanism for plants to deter various insects due to their unpleasant taste and their toxicity. Here, we provide evidence that saponin from Quillaja saponaria functions as an antifeedant as well as an insecticide to ward off insects in both the larval and the adult stages. Using a behavioral screen of 26 mutant fly lines, we show that the Gr28b gene cluster plays a role in saponin avoidance in the labellum. The Gr28b mutant does not avoid saponin and exhibits increased lethality when fed saponin-mixed food. Tissue-specific rescue experiments with five different Gr28b isoforms revealed that only the Gr28b.c isoform is required for saponin sensation. We propose that in contrast to sensing many other bitter compounds, saponin sensing does not require the function of core taste receptors, such as GR32a, GR33a, and GR66a. Our results reveal a novel role for GR28b in taste. In addition, the ability of saponin to act as insecticides as well as antifeedants suggests its potential application in controlling insect pests.