MetaboLights

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ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/m_MTBLS14319_LC-MS_negative_hilic_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/m_MTBLS14319_LC-MS_positive_reverse-phase_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/m_MTBLS14319_LC-MS_positive_hilic_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/m_MTBLS14319_LC-MS_negative_reverse-phase_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/s_MTBLS14319.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/a_MTBLS14319_LC-MS_positive_reverse-phase.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/i_Investigation.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/a_MTBLS14319_LC-MS_negative_reverse-phase.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/a_MTBLS14319_LC-MS_positive_hilic.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319/a_MTBLS14319_LC-MS_negative_hilic.txtprimary200OKftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14319An Excel macro was designed to consolidate multiple annotations within a measurement, thereby combining the four datasets for each sample. The values within the matrix reflected the measured intensity of a metabolite. The output file was used for statistical analysis using the MetaboAnalyst 6.0 web application (RRID:SCR_015539). Metabolites with more than 70% missing values in a group were excluded from the dataset. Missing values were replaced with one-fifth of the lowest detected value. Data were transformed to the base-10 logarithm and scaled using the auto-scaling function. MetaboLightsPublicLiquid Chromatography MS - negative - reverse-phaseLiquid Chromatography MS - positive - hilicLiquid Chromatography MS - positive - reverse-phaseLiquid Chromatography MS - negative - hilicThe analysis was performed using an Agilent 1290 Infinity HPLC system (RRID:SCR_019375). Two different columns were used: the Waters ACQUITY UPLC BEH C18 column for non-polar and mild-polar metabolites, and the Agilent InfinityLab Poroshell 120 HILIC-Z for polar metabolites. The mobile phases consisted of water or acetonitrile, each mixed with 0.1% formic acid. The column temperature was set at 30 °C with an injection volume of 10 μL using an autosampler. Between each injection, the needle was washed three times with 70% water/30% acetonitrile/0.1% formic acid.In-depth characterization of the CaV2.2-knockout mouse line. 10.1038/s41598-026-60827-w.Janine KutzscheForschungszentrum JÃ¼lichblood plasmabrainmass spectrometry assay30 mg of each hemisphere were homogenized in methanol/water (4:1) using a Precellys Evolution homogenizer (Bertin Technologies) at 6000 rpm for 3x 20 s, with a 5 s pause between each cycle. The samples were continuously cooled. The homogenates were stored at -20 °C for one hour and then centrifuged (15 min, 16,000 xg, 4 °C). The supernatant was evaporated using a vacuum concentrator 5301 (Eppendorf). The pellet was reconstituted in 100 μL water/methanol (1:1) and sonicated in an ice bath for one minute. The samples were centrifuged at 4 °C and 16,000 xg for 15 min.Plasma samples were thawed on ice and briefly homogenized. 20 μL of plasma were mixed with 80 μL of ice-cold methanol. The mixture was vortexed for 30 s and stored at -20 °C for one hour. The subsequent sample preparation followed the same protocol as the brain samples.Mus musculushttps://www.ebi.ac.uk/metabolights/MTBLS14319Janine Kutzsche. Forschungszentrum Jülich GmbH, Institute of Biological Information Processing, Structural Biochemistry (IBI-7). j.kutzsche@fz-juelich.de.The obtained data were analyzed using the MassHunter Workstation Qualitative Analysis software (version 12.0, Agilent, RRID:SCR_016657). Metabolites were initially identified using Auto-Select Compound Mining. Metabolites with a score above 80 were compared with the METLIN database (RRID:SCR_010500). The resulting dataset contained the identified molecules along with their measured intensity.Cohort defined by study part (1 and 2)SexFrequency of participation in behavioral experimentsGenotypej.kutzsche@fz-juelich.deBrains were dissected and cut into the two hemispheres. The left hemisphere was snap frozen in isopentane and stored at -80 °C until LC-MS/MS analysis.The blood was extracted from each mouse by cardiac puncture. The plasma was collected following a centrifugation at 3,000 xg for 15 min, and stored at -80 °C until LC-MS/MS analysis.Metabolomicshydrophilic interaction chromatographyepilepsyMetabolomicsMus musculusHyperactivityuntargeted analysisAgilent softwareCACNA1BpositiveAgilent 1290 Infinity HPLCbrainexperimental sampleAgilent 6550 iFunnel Q-TOFnegativeB6;129S4-Cacna1b<tm1Ttan>blood plasmareverse phase chromatographyKnock-outhydrophilic interaction chromatographyepilepsyMetabolomicsMus musculusHyperactivityuntargeted analysisAgilent softwareCACNA1BpositivebrainAgilent 1290 Infinity HPLCexperimental sampleAgilent 6550 iFunnel Q-TOFnegativeB6;129S4-Cacna1b<tm1Ttan>blood plasmareverse phase chromatographyKnock-outThe analysis was performed using an Agilent 6550 iFunnel Q-TOF LC/MS (RRID:SCR_019433) and electrospray ionization. The Q-TOF was calibrated prior to analysis in accordance with the manufacturer's recommendations. The MS/MS system used the Auto MS/MS mode. For MS1, the mass range was 50-1000 m/z with an acquisition rate of two spectra per second. MS2 covered a mass range of 30-1000 m/z, with a recording rate of seven spectra per second. The specified collision energies were 10 V, 20 V, and 40 V. falseIn-depth characterization of the CaV2.2-knockout mouse lineABSTRACT The voltage-gated calcium channel CaV2.2 is predominantly located in neuronal synapses, where it plays a role in pain signaling and neurotransmitter release. Furthermore, CaV2.2 is important during early central nervous system development, as it is subsequently replaced by other calcium channels. A number of mouse lines with disrupted CaV2.2 channels have been developed through the utilization of a gene-targeting vector. The CaV2.2-deficient mice primarily exhibited reduced anxiety-linked behavior and response to pain stimuli.In a previous study, we observed hyperactivity and seizures in CaV2.2-knockout mice. Given the absence of prior descriptions of these characteristics, an in-depth characterization was conducted. This characterization included longitudinal behavioral studies, as well as histological, proteomic, and metabolomic analyses. Irrespective of the frequency with which testing was conducted, a variety of behaviors were observed (e.g., hyperactivity and increased exploratory behavior) in comparison with wild-type mice. The frequency and severity of the performed tests demonstrated a positive correlation with the magnitude of the difference in molecular markers between CaV2.2-knockout and wild-type mice. Seizures were observed but were not common.These results suggest that the CaV2.2-knockout mouse lines are susceptible to repeated stress stimuli during experimental studies. The mice are characterized by hyperactivity, explorative behavior, and sometimes experience seizures. 2026-07-092026-04-20MTBLS14319