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Authorship：Lead author,　Corresponding author   Publishing type：Research paper (scientific journal)   Publisher：Society for Neuroscience  

Neuronal activity is modulated not only by inputs from other neurons but also by various factors, such as bioactive substances. Noradrenergic (NA) neurons in the locus coeruleus (LC-NA neurons) are involved in diverse physiological functions, including sleep/wakefulness and stress responses. Previous studies have identified various substances and receptors that modulate LC-NA neuronal activity through techniques including electrophysiology, calcium imaging, and single-cell RNA sequencing. However, many substances with unknown physiological significance have been overlooked. Here, we established an efficient screening method for identifying substances that modulate LC-NA neuronal activity through intracellular calcium ([Ca²⁺]_i) imaging using brain slices. Using both sexes of mice, we screened 53 bioactive substances, and identified five novel substances: gastrin-releasing peptide, neuromedin U, and angiotensin II, which increase [Ca²⁺]_i, and pancreatic polypeptide and prostaglandin D₂, which decrease [Ca²⁺]_i. Among them, neuromedin U induced the greatest response in female mice. In terms of the duration of [Ca²⁺]_ichange, we focused on prostaglandin E₂(PGE₂), since it induces a long-lasting decrease in [Ca²⁺]_ivia the EP₃receptor. Conditional knock-out of the receptor in LC-NA neurons resulted in increased depression-like behavior, prolonged wakefulness in the dark period, and increased [Ca²⁺]_iafter stress exposure. Our results demonstrate the effectiveness of our screening method for identifying substances that modulate a specific neuronal population in an unbiased manner and suggest that stress-induced prostaglandin E₂can suppress LC-NA neuronal activity to moderate the behavioral response to stressors. Our screening method will contribute to uncovering previously unknown physiological functions of uncharacterized bioactive substances in specific neuronal populations.

SIGNIFICANCE STATEMENTBioactive substances modulate the activity of specific neuronal populations. However, since only a limited number of substances with predicted effects have been investigated, many substances that may modulate neuronal activity have gone unrecognized. Here, we established an unbiased method for identifying modulatory substances by measuring the intracellular calcium signal, which reflects neuronal activity. We examined noradrenergic (NA) neurons in the locus coeruleus (LC-NA neurons), which are involved in diverse physiological functions. We identified five novel substances that modulate LC-NA neuronal activity. We also found that stress-induced prostaglandin E₂(PGE₂) may suppress LC-NA neuronal activity and influence behavioral outcomes. Our screening method will help uncover previously overlooked functions of bioactive substances and provide insight into unrecognized roles of specific neuronal populations.

DOI： 10.1523/jneurosci.0353-23.2023

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Authorship：Lead author   Publishing type：Research paper (scientific journal)   Publisher：Elsevier BV  

DOI： 10.1016/j.neures.2022.12.009

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Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)   Publisher：Springer Science and Business Media LLC  

Abstract

Optogenetics is a powerful tool for manipulating neuronal activity by light illumination with high temporal and spatial resolution. Anion-channelrhodopsins (ACRs) are light-gated anion channels that allow researchers to efficiently inhibit neuronal activity. A blue light-sensitive ACR2 has recently been used in several in vivo studies; however, the reporter mouse strain expressing ACR2 has not yet been reported. Here, we generated a new reporter mouse strain, LSL-ACR2, in which ACR2 is expressed under the control of Cre recombinase. We crossed this strain with a noradrenergic neuron-specific driver mouse (NAT-Cre) to generate NAT-ACR2 mice. We confirmed Cre-dependent expression and function of ACR2 in the targeted neurons by immunohistochemistry and electrophysiological recordings in vitro, and confirmed physiological function using an in vivo behavioral experiment. Our results show that the LSL-ACR2 mouse strain can be applied for optogenetic inhibition of targeted neurons, particularly for long-lasting continuous inhibition, upon crossing with Cre-driver mouse strains. The LSL-ACR2 strain can be used to prepare transgenic mice with homogenous expression of ACR2 in targeted neurons with a high penetration ratio, good reproducibility, and no tissue invasion.

DOI： 10.1038/s41598-023-30907-2

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Other Link： https://www.nature.com/articles/s41598-023-30907-2

Publishing type：Research paper (scientific journal)   Publisher：Oxford University Press (OUP)  

Abstract

Melanin-concentrating hormone-producing neurons (MCH neurons), found mainly in the lateral hypothalamus and surrounding areas, play essential roles in various brain functions, including sleep and wakefulness, reward, metabolism, learning, and memory. These neurons coexpress several neurotransmitters and act as glutamatergic neurons. The contribution of glutamate from MCH neurons to memory- and metabolism-related functions has not been fully investigated. In a mouse model, we conditionally knocked out Slc17a6 gene, which encodes for vesicular glutamate transporter 2 (vGlut2), in the MCH neurons exclusively by using two different methods: the Cre recombinase/loxP system and in vivo genome editing using CRISPR/Cas9. Then, we evaluated several aspects of memory and measured metabolic rates using indirect calorimetry. We found that mice with MCH neuron-exclusive vGlut2 ablation had higher discrimination ratios between novel and familiar stimuli for novel object recognition, object location, and three-chamber tests. In contrast, there was no significant change in body weight, food intake, oxygen consumption, respiratory quotient, or locomotor activity. These findings suggest that glutamatergic signaling from MCH neurons is required to regulate memory, but its role in regulating metabolic rate is negligible.

DOI： 10.1093/pnasnexus/pgae275

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Other Link： https://academic.oup.com/pnasnexus/article-pdf/3/7/pgae275/58598945/pgae275.pdf

Publishing type：Research paper (scientific journal)   Publisher：Springer Science and Business Media LLC  

Abstract

We previously reported that enhanced corticotropin-releasing factor (CRF) signaling in the bed nucleus of the stria terminalis (BNST) caused the aversive responses during acute pain and suppressed the brain reward system during chronic pain. However, it remains to be examined whether chronic pain alters the excitability of CRF neurons in the BNST. In this study we investigated the chronic pain-induced changes in excitability of CRF-expressing neurons in the oval part of the BNST (ovBNST^CRF neurons) by whole-cell patch-clamp electrophysiology. CRF-Cre; Ai14 mice were used to visualize CRF neurons by tdTomato. Electrophysiological recordings from brain slices prepared from a mouse model of neuropathic pain revealed that rheobase and firing threshold were significantly decreased in the chronic pain group compared with the sham-operated control group. Firing rate of the chronic pain group was higher than that of the control group. These data indicate that chronic pain elevated neuronal excitability of ovBNST^CRF neurons.

DOI： 10.1186/s13041-024-01094-6

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Other Link： https://link.springer.com/article/10.1186/s13041-024-01094-6/fulltext.html

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Event date： 2023.9

Language：English   Presentation type：Poster presentation  

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Event date： 2022.6 - 2022.7

Language：Japanese   Presentation type：Oral presentation (general)  

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Event date： 2022.3

Language：English   Presentation type：Poster presentation  

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Event date： 2021.9

Language：Japanese   Presentation type：Oral presentation (invited, special)  

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Event date： 2021.7

Language：English   Presentation type：Poster presentation  

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