Choroid plexus (ChP), the brain structure primarily responsible for cerebrospinal fluid production, contains a robust circadian clock, whose role remains to be elucidated. The aim of our study was to [1] identify rhythmically controlled cellular processes in the mouse ChP and [2] assess the role and nature of signals derived from the master clock in the suprachiasmatic nuclei (SCN) that control ChP rhythms. To accomplish this goal, we used various mouse models (WT, mPer2Luc, ChP-specific Bmal1 knockout) and combined multiple experimental approaches, including surgical lesion of the SCN (SCNx), time-resolved transcriptomics, and single cell luminescence microscopy. In ChP of control (Ctrl) mice collected every 4 h over 2 circadian cycles in darkness, we found that the ChP clock regulates many processes, including the cerebrospinal fluid circadian secretome, precisely times endoplasmic reticulum stress response, and controls genes involved in neurodegenerative diseases (Alzheimer's disease, Huntington's disease, and frontotemporal dementia). In ChP of SCNx mice, the rhythmicity detected in vivo and ex vivo was severely dampened to a comparable extent as in mice with ChP-specific Bmal1 knockout, and the dampened cellular rhythms were restored by daily injections of dexamethasone in mice. Our data demonstrate that the ChP clock controls tissue-specific gene expression and is strongly dependent on the presence of a functional connection with the SCN. The results may contribute to the search for a novel link between ChP clock disruption and impaired brain health.

• Keywords
• mPer2 Luc mouse, Choroid plexus, Circadian clock, Circadian transcriptome, Glucocorticoid, Mouse, Suprachiasmatic nuclei,
• MeSH
• Circadian Clocks * physiology   MeSH
• Circadian Rhythm physiology   MeSH
• Mice, Inbred C57BL MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Suprachiasmatic Nucleus * metabolism   physiology   MeSH
• Choroid Plexus * metabolism   physiology   MeSH
• ARNTL Transcription Factors metabolism   genetics   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Bmal1 protein, mouse MeSH Browser
• ARNTL Transcription Factors MeSH

Liquid chromatography with mass spectrometry (LC-MS)-based metabolomics detects thousands of molecular features (retention time-m/z pairs) in biological samples per analysis, yet the metabolite annotation rate remains low, with 90% of signals classified as unknowns. To enhance the metabolite annotation rates, researchers employ tandem mass spectral libraries and challenging in silico fragmentation software. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) may offer an additional layer of structural information in untargeted metabolomics, especially for identifying specific unidentified metabolites that are revealed to be statistically significant. Here, we investigate the potential of hydrophilic interaction liquid chromatography (HILIC)-HDX-MS in untargeted metabolomics. Specifically, we evaluate the effectiveness of two approaches using hypothetical targets: the post-column addition of deuterium oxide (D2O) and the on-column HILIC-HDX-MS method. To illustrate the practical application of HILIC-HDX-MS, we apply this methodology using the in silico fragmentation software MS-FINDER to an unknown compound detected in various biological samples, including plasma, serum, tissues, and feces during HILIC-MS profiling, subsequently identified as N1-acetylspermidine.

• Keywords
• hydrogen/deuterium exchange, liquid chromatography, mass spectrometry, metabolomics, unknown identification,
• MeSH
• Chromatography, Liquid methods   MeSH
• Deuterium MeSH
• Hydrophobic and Hydrophilic Interactions MeSH
• Metabolomics * methods   MeSH
• Hydrogen Deuterium Exchange-Mass Spectrometry * MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Deuterium MeSH