#METABOLOMICS WORKBENCH MingxinZhang_20250214_002539 DATATRACK_ID:5625 STUDY_ID:ST003739 ANALYSIS_ID:AN006139 PROJECT_ID:PR002325 VERSION 1 CREATED_ON February 15, 2025, 5:03 pm #PROJECT PR:PROJECT_TITLE Serum neurotransmitter targeted metabolome in sleep-deprived mice PR:PROJECT_SUMMARY This project investigates the changes in serum neurotransmitter levels before PR:PROJECT_SUMMARY and after 2 days of sleep deprivation in mice, exploring the alterations in PR:PROJECT_SUMMARY serum neurotransmitter release caused by sleep deprivation. Metabolomic results PR:PROJECT_SUMMARY indicate that in mice subjected to 2 days of sleep deprivation, serum PR:PROJECT_SUMMARY neurotransmitters such as 5-HTP, Kyn, and KynA were significantly reduced, while PR:PROJECT_SUMMARY Ach, Gln, Trp, and 5-HT were significantly increased. PR:INSTITUTE China Agriculture University PR:LAST_NAME Zhang PR:FIRST_NAME Mingxin PR:ADDRESS 2 West Yuanmingyuan Road, Beijing, Beijing, 100193, China PR:EMAIL zmx15310686122@163.com PR:PHONE 18800152903 #STUDY ST:STUDY_TITLE Serum neurotransmitter targeted metabolome in sleep-deprived (SD) mice ST:STUDY_SUMMARY Sleep disturbances associate with pathogenesis of numerous chronic disorders, ST:STUDY_SUMMARY including chronic gastrointestinal diseases. However, the mechanism that ST:STUDY_SUMMARY transmits sleep disturbance-induced aberrant neural signaling from the brain to ST:STUDY_SUMMARY the gut remains elusive. We show that acute sleep deprivation (SD) impairs ST:STUDY_SUMMARY intestinal stem cell function, leading to shortening of crypt-villus ST:STUDY_SUMMARY architecture and Paneth cell loss. We identified dorsal motor nucleus of vagus ST:STUDY_SUMMARY (DMV) as the SD-sensitive central nervous system center that transmits sleep ST:STUDY_SUMMARY effects to the gut. SD aberrantly activates DMV neurons, driving excessive ST:STUDY_SUMMARY acetylcholine release from the vagus nerve into the gut. Acetylcholine triggers ST:STUDY_SUMMARY 5-hydroxytryptamine (5-HT) release by enterochromaffin cells and suppresses its ST:STUDY_SUMMARY reuptake via muscarinic receptors, thereby causing a spike in 5-HT levels. ST:STUDY_SUMMARY Elevated 5-HT induces excessive oxidative stress in intestinal stem cells ST:STUDY_SUMMARY through its receptor HTR4, promoting gut pathologies. Overall, we reveal ST:STUDY_SUMMARY SD-responsive neural circuit that controls intestinal stem cells and identify ST:STUDY_SUMMARY therapeutic strategies for mitigating SD-related gut diseases. ST:INSTITUTE China Agriculture University ST:LAST_NAME Zhang ST:FIRST_NAME Mingxin ST:ADDRESS 2 West Yuanmingyuan Road, Beijing, Beijing, 100193, China ST:EMAIL zmx15310686122@163.com ST:PHONE 18800152903 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENDER Female #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS 1 SD0d_1 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FZTM220028237-1A SUBJECT_SAMPLE_FACTORS 2 SD0d_2 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FZTM220028238-1A SUBJECT_SAMPLE_FACTORS 3 SD0d_3 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FZTM220028239-1A SUBJECT_SAMPLE_FACTORS 4 SD0d_4 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FZTM220028240-1A SUBJECT_SAMPLE_FACTORS 5 SD0d_5 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FZTM220028241-1A SUBJECT_SAMPLE_FACTORS 6 SD2d_1 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FZTM220028242-1A SUBJECT_SAMPLE_FACTORS 7 SD2d_2 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FZTM220028243-1A SUBJECT_SAMPLE_FACTORS 8 SD2d_3 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FZTM220028244-1A SUBJECT_SAMPLE_FACTORS 9 SD2d_4 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FZTM220028245-1A SUBJECT_SAMPLE_FACTORS 10 SD2d_5 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FZTM220028246-1A #COLLECTION CO:COLLECTION_SUMMARY Fresh blood from mice was collected via cardiac puncture before and after sleep CO:COLLECTION_SUMMARY deprivation. After allowing the blood to stand for 1 hour, it was centrifuged at CO:COLLECTION_SUMMARY 3000g for 10 minutes to obtain 400 µL of serum, which was then stored at CO:COLLECTION_SUMMARY -80°C.The samples (100 μL) were taken respectively and homogenized with 200 CO:COLLECTION_SUMMARY μL of acetonitrile (80%) which contained mixed internal standards and CO:COLLECTION_SUMMARY centrifuged at 12,000 rpm for 10 min. Finally, the supernatant (2 μL) was CO:COLLECTION_SUMMARY injected into the LC-MS/MS system for analysis. CO:SAMPLE_TYPE Blood (serum) #TREATMENT TR:TREATMENT_SUMMARY Serum samples were obtained immediately after mice were subjected to 48 hours of TR:TREATMENT_SUMMARY continuous sleep deprivation(SD). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The stock solution of individual neurotransmitter was mixed and prepared in SP:SAMPLEPREP_SUMMARY neurotransmitter -free matrix to obtain a series of neurotransmitter SP:SAMPLEPREP_SUMMARY calibrators. Certain concentrations of L-Tyrosine-d4 was mixed as Internal SP:SAMPLEPREP_SUMMARY Standard (IS). The stock solution of all of these and working solution were SP:SAMPLEPREP_SUMMARY stored in refrigerator of -20°C #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME AB Sciex QTRAP 6500+ CH:COLUMN_NAME Waters XSelect HSS T3 XP (150 x 3mm,2.5um) CH:SOLVENT_A 0.1% formic acid in 5mM ammonium acetate CH:SOLVENT_B 0.1% formic acid in acetonitrile CH:FLOW_GRADIENT initial 5% B, 2min; 5-40% B, 4min; 40-70% B, 7 min; 70-100% B, 7.5min; 100% B, CH:FLOW_GRADIENT 9min; 100-5% B, 9.1min; 5% B, 11min. CH:FLOW_RATE 0.30 mL/min CH:COLUMN_TEMPERATURE 45°C #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6520 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Curtain Gas：35psi； Collision Gas：Medium；IonSpray Voltage：4500V； MS:MS_COMMENTS Temperature：550℃； Ion Source Gas 1：60； Ion Source Gas 2：60 #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS ng/ml MS_METABOLITE_DATA_START Samples SD0d_1 SD0d_2 SD0d_3 SD0d_4 SD0d_5 SD2d_1 SD2d_2 SD2d_3 SD2d_4 SD2d_5 Factors Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d 4-Aminobutyric acid 5.742199069 3.415319607 7.540027207 5.929117503 5.183642436 7.77146814 3.329923441 3.35123182 5.133495725 6.664622139 Histamine 23.62847791 27.27129173 18.50553212 46.33843489 36.07635922 53.44962299 71.30917222 56.21072554 34.87532514 39.55313135 Picolinic acid 22.76652806 17.00048162 21.04621304 24.6896427 39.25567618 17.27294212 26.14260891 29.26757864 20.97139076 29.36382211 Acetylcholine chloride 3.399086941 1.747054004 2.763207561 2.123492317 2.090690677 2.496528753 3.187686496 3.386654177 2.214818262 2.140686182 Hydroxytyramine hydrochloride 52.19328944 35.21163179 52.22130572 76.17821761 129.8764203 41.10972997 41.26284001 56.37347221 69.09149211 28.14775878 Tryptamine 0.405885878 0.821243608 0.468027649 0.397635993 1.074413414 2.173243472 1.955404745 1.747366951 1.473860388 3.039985076 Noradrenaline hydrochloride 55502.29661 53341.97685 68859.57017 83349.9607 59755.87662 61356.4521 62404.25808 64055.71288 57608.62225 59780.27904 Serotonin hydrochloride 1614.910186 1250.111892 1819.045041 2573.006068 2167.672026 2721.62831 2323.251075 1883.213887 2057.066407 2811.166491 L-Tyrosine 7451.442683 6087.301584 3788.007313 9590.061789 8495.781519 9826.914581 7825.584693 6977.087331 7513.964701 9052.665349 Kynurenic acid 39.52750643 16.99437351 30.16941231 36.8864081 41.48811907 23.36078625 23.41040668 23.37897282 19.7233528 24.38170105 5-Hydroxyindole-3-acetic acid 131.9974315 106.9930878 109.9895812 160.6151153 147.1357488 165.7213829 145.7548672 167.907975 145.7308692 172.7796213 L-Tryptophan 7933.917645 7197.575404 7396.568514 12334.88589 11143.25824 12650.50835 11621.54163 10916.43889 12350.71827 11528.10879 Xanthurenic acid 14.41982972 7.9862103 8.569418305 12.84603598 8.100464518 8.366194813 8.124952393 9.663699356 5.098197142 7.98828149 DL-Kynurenine 1392.389393 708.0417667 919.4042681 979.1688748 1010.914576 755.5002434 681.6501739 746.8369603 626.6502582 626.1652636 L-Glutamic acid 56046.53934 45628.3943 103663.769 77891.69018 66310.72511 72015.50159 59861.98044 54868.63699 60678.809 74655.6867 L-Histidine 265.4795219 233.2453245 149.9650439 345.3604887 312.7196858 280.6802494 231.7512738 281.4122072 247.9883685 186.8959679 L-Glutamine 367972.0996 293006.3941 216073.5875 349066.3141 470264.9073 444635.5504 406148.9431 391618.0378 428052.8394 427198.3824 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name 4-Aminobutyric acid Histamine Picolinic acid Acetylcholine chloride Hydroxytyramine hydrochloride Tryptamine Noradrenaline hydrochloride Serotonin hydrochloride L-Tyrosine Kynurenic acid 5-Hydroxyindole-3-acetic acid L-Tryptophan Xanthurenic acid DL-Kynurenine L-Glutamic acid L-Histidine L-Glutamine METABOLITES_END #END