Multimodal Imaging for Validation and Optimization of Ion Channel-Based Chemogenetics in Nonhuman Primates.

Chemogenetic tools provide an opportunity to manipulate neuronal activity and behavior selectively and repeatedly in nonhuman primates (NHPs) with minimal invasiveness. Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) are one example that is based on mutated muscarinic acetylcholine receptors. Another channel-based chemogenetic system available for neuronal modulation in NHPs uses pharmacologically selective actuator modules (PSAMs), which are selectively activated by pharmacologically selective effector molecules (PSEMs). To facilitate the use of the PSAM/PSEM system, the selection and dosage of PSEMs should be validated and optimized for NHPs. To this end, we used a multimodal imaging approach. We virally expressed excitatory PSAM (PSAM4-5HT3) in the striatum and the primary motor cortex (M1) of two male macaque monkeys, and visualized its location through positron emission tomography (PET) with the reporter ligand [18F]ASEM. Chemogenetic excitability of neurons triggered by two PSEMs (uPSEM817 and uPSEM792) was evaluated using [18F]fluorodeoxyglucose-PET imaging, with uPSEM817 being more efficient than uPSEM792. Pharmacological magnetic resonance imaging (phMRI) showed that increased brain activity in the PSAM4-expressing region began ∼13 min after uPSEM817 administration and continued for at least 60 min. Our multimodal imaging data provide valuable information regarding the manipulation of neuronal activity using the PSAM/PSEM system in NHPs, facilitating future applications.SIGNIFICANCE STATEMENT Like other chemogenetic tools, the ion channel-based system called pharmacologically selective actuator module/pharmacologically selective effector molecule (PSAM/PSEM) allows remote manipulation of neuronal activity and behavior in living animals. Nevertheless, its application in nonhuman primates (NHPs) is still limited. Here, we used multitracer positron emission tomography (PET) imaging and pharmacological magnetic resonance imaging (phMRI) to visualize an excitatory chemogenetic ion channel (PSAM4-5HT3) and validate its chemometric function in macaque monkeys. Our results provide the optimal agonist, dose, and timing for chemogenetic neuronal manipulation, facilitating the use of the PSAM/PSEM system and expanding the flexibility and reliability of circuit manipulation in NHPs in a variety of situations.

Chemogenetic tools to interrogate brain functions.

Elucidating the roles of neuronal cell types for physiology and behavior is essential for understanding brain functions. Perturbation of neuron electrical activity can be used to probe the causal relationship between neuronal cell types and behavior. New genetically encoded neuron perturbation tools have been developed for remotely controlling neuron function using small molecules that activate engineered receptors that can be targeted to cell types using genetic methods. Here we describe recent progress for approaches using genetically engineered receptors that selectively interact with small molecules. Called "chemogenetics," receptors with diverse cellular functions have been developed that facilitate the selective pharmacological control over a diverse range of cell-signaling processes, including electrical activity, for molecularly defined cell types. These tools have revealed remarkably specific behavioral physiological influences for molecularly defined cell types that are often intermingled with populations having different or even opposite functions.

The international food unit: a new measurement aid that can improve portion size estimation.

BACKGROUND: Portion size education tools, aids and interventions can be effective in helping prevent weight gain. However consumers have difficulties in estimating food portion sizes and are confused by inconsistencies in measurement units and terminologies currently used. Visual cues are an important mediator of portion size estimation, but standardized measurement units are required. In the current study, we present a new food volume estimation tool and test the ability of young adults to accurately quantify food volumes. The International Food Unit™ (IFU™) is a 4x4x4 cm cube (64cm3), subdivided into eight 2 cm sub-cubes for estimating smaller food volumes. Compared with currently used measures such as cups and spoons, the IFU™ standardizes estimation of food volumes with metric measures. The IFU™ design is based on binary dimensional increments and the cubic shape facilitates portion size education and training, memory and recall, and computer processing which is binary in nature. METHODS: The performance of the IFU™ was tested in a randomized between-subject experiment (n = 128 adults, 66 men) that estimated volumes of 17 foods using four methods; the IFU™ cube, a deformable modelling clay cube, a household measuring cup or no aid (weight estimation). Estimation errors were compared between groups using Kruskall-Wallis tests and post-hoc comparisons. RESULTS: Estimation errors differed significantly between groups (H(3) = 28.48, p < .001). The volume estimations were most accurate in the group using the IFU™ cube (Mdn = 18.9%, IQR = 50.2) and least accurate using the measuring cup (Mdn = 87.7%, IQR = 56.1). The modelling clay cube led to a median error of 44.8% (IQR = 41.9). Compared with the measuring cup, the estimation errors using the IFU™ were significantly smaller for 12 food portions and similar for 5 food portions. Weight estimation was associated with a median error of 23.5% (IQR = 79.8). CONCLUSIONS: The IFU™ improves volume estimation accuracy compared to other methods. The cubic shape was perceived as favourable, with subdivision and multiplication facilitating volume estimation. Further studies should investigate whether the IFU™ can facilitate portion size training and whether portion size education using the IFU™ is effective and sustainable without the aid. A 3-dimensional IFU™ could serve as a reference object for estimating food volume.

Factors Affecting the Number of Pollen Grains per Male Strobilus in Japanese Cedar (Cryptomeria japonica).

Japanese cedar (Cryptomeria japonica) is the most important timber species in Japan; however, its pollen is the primary cause of pollinosis in Japan. The total number of pollen grains produced by a single tree is determined by the number of male strobili (male flowers) and the number of pollen grains per male strobilus. While the number of male strobili is a visible and well-investigated trait, little is known about the number of pollen grains per male strobilus. We hypothesized that genetic and environmental factors affect the pollen number per male strobilus and explored the factors that affect pollen production and genetic variation among clones. We counted pollen numbers of 523 male strobili from 26 clones using a cell counter method that we recently developed. Piecewise Structural Equation Modeling (pSEM) revealed that the pollen number is mostly affected by genetic variation, male strobilus weight, and pollen size. Although we collected samples from locations with different environmental conditions, statistical modeling succeeded in predicting pollen numbers for different clones sampled from branches facing different directions. Comparison of predicted pollen numbers revealed that they varied >3-fold among the 26 clones. The determination of the factors affecting pollen number and a precise evaluation of genetic variation will contribute to breeding strategies to counter pollinosis. Furthermore, the combination of our efficient counting method and statistical modeling will provide a powerful tool not only for Japanese cedar but also for other plant species.

Practical Considerations for the Use of DREADD and Other Chemogenetic Receptors to Regulate Neuronal Activity in the Mammalian Brain.

Chemogenetics is the process of genetically expressing a macromolecule receptor capable of modulating the activity of the cell in response to selective chemical ligand. This chapter will cover the chemogenetic technologies that are available to date, focusing on the commonly available engineered or otherwise modified ligand-gated ion channels and G-protein-coupled receptors in the context of neuromodulation. First, we will give a brief overview of each chemogenetic approach as well as in vitro/in vivo applications, then we will list their strengths and weaknesses. Finally, we will provide tips for ligand application in each case.Each technology has specific limitations that make them more or less suitable for different applications in neuroscience although we will focus mainly on the most commonly used and versatile family named designer receptors exclusively activated by designer drugs or DREADDs. We here describe the most common cases where these can be implemented and provide tips on how and where these technologies can be applied in the field of neuroscience.