Summary of Study ST002237

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001427. The data can be accessed directly via it's Project DOI: 10.21228/M88D9X This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST002237
Study TitleMetabolomic analysis of brain cortex from neuronal specific Depdc5 knockout in fed and fasting state
Study TypeDepdc5 KO fed vs. fasted comparison
Study SummaryCaloric restriction and acute fasting are known to reduce seizures but through unclear mechanisms. In this study, we demonstrate that mTORC1 signaling is reduced after acute fasting of mice. In neurons, mTORC1 is most sensitive to withdrawal of leucine, arginine, and glutamine, which is dependent on DEPDC5. We performed metabolomic analysis of brain cortex from neuronal specific Depdc5 knockout in fed and fasting state. The Depdc5 neuronal specific knockout mice are resistant to sensing significant fluctuations in brain amino acid levels after fasting. These results establish that acute fasting reduces seizure susceptibility in a DEPDC5-dependent manner.
Institute
Northwestern University, Feinberg School of Medicine
DepartmentMedicine
LaboratoryChandel Lab
Last NameChandel
First NameNavdeep
Address303 E Superior St, Chicago, Illinois, 60611, USA
Emailnav@northwestern.edu
Phone3125032549
Submit Date2022-07-27
Num Groups4
Total Subjects40
Num Males20
Num Females20
PublicationsDEPDC5-dependent mTORC1 signaling mechanisms are critical for the anti-seizure effects of acute fasting
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2022-08-15
Release Version1
Navdeep Chandel Navdeep Chandel
https://dx.doi.org/10.21228/M88D9X
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001427
Project DOI:doi: 10.21228/M88D9X
Project Title:Metabolomic analysis of brain cortex from neuronal specific Depdc5 knockout in fed and fasting states
Project Type:LCMS based comprehensive hydrophilic metabolites profiling
Project Summary:We performed a metabolomic analysis of the brain cortex from neuronal-specific Depdc5 knockout in fed and fasting states.
Institute:Northwestern University Feinberg School of Medicine
Department:Medicine
Laboratory:Chandel Lab
Last Name:Chandel
First Name:Navdeep
Address:303 E Superior St, Chicago, Illinois, 60611, USA
Email:nav@northwestern.edu
Phone:3125032549
Funding Source:NIA
Publications:DEPDC5-dependent mTORC1 signaling mechanisms are critical for the anti-seizure effects of acute fasting
Contributors:Christopher J. Yuskaitis, Jinita B. Modasia, Sandra Schroetter, Leigh-Ana Rossitto, Karenna Groff, Christopher Morici, Divakar S. Mithal, Ram P. Chakrabarty, Navdeep S. Chandel, Brendan D. Manning, Mustafa Sahin

Subject:

Subject ID:SU002323
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Genotype Treatment
SA213107600-1Depdc5 KO Control
SA213108609-6Depdc5 KO Control
SA213109248-7Depdc5 KO Control
SA213110192-2Depdc5 KO Control
SA213111192-4Depdc5 KO Control
SA213112192-5Depdc5 KO Control
SA213113189-2Depdc5 KO Control
SA213114167-7Depdc5 KO Control
SA213115993-10Depdc5 KO Control
SA213116625-1Depdc5 KO Control
SA213117986-5Depdc5 KO Fasted
SA213118192-6Depdc5 KO Fasted
SA213119986-3Depdc5 KO Fasted
SA213120611-5Depdc5 KO Fasted
SA213121627-7Depdc5 KO Fasted
SA213122601-5Depdc5 KO Fasted
SA213123248-6Depdc5 KO Fasted
SA213124166-6Depdc5 KO Fasted
SA213125192-1Depdc5 KO Fasted
SA213126169-3Depdc5 KO Fasted
SA213127993-1Depdc5 WT Control
SA213128189-3Depdc5 WT Control
SA213129169-2Depdc5 WT Control
SA213130169-5Depdc5 WT Control
SA213131601-1Depdc5 WT Control
SA213132166-3Depdc5 WT Control
SA213133166-4Depdc5 WT Control
SA213134191-3Depdc5 WT Control
SA213135191-1Depdc5 WT Control
SA213136993-4Depdc5 WT Control
SA213137993-5Depdc5 WT Fasted
SA213138166-5Depdc5 WT Fasted
SA213139190-4Depdc5 WT Fasted
SA213140600-7Depdc5 WT Fasted
SA213141248-1Depdc5 WT Fasted
SA213142600-8Depdc5 WT Fasted
SA213143601-2Depdc5 WT Fasted
SA213144190-5Depdc5 WT Fasted
SA213145601-4Depdc5 WT Fasted
SA213146627-6Depdc5 WT Fasted
Showing results 1 to 40 of 40

Collection:

Collection ID:CO002316
Collection Summary:Mouse cerebral cortex was rapidly dissected and flash frozen.
Sample Type:Brain
Storage Conditions:-80℃

Treatment:

Treatment ID:TR002335
Treatment Summary:Group housed mice, 6-9 weeks old Depdc5cc+ mice and their littermate controls, were randomized to fed or fasted conditions. Mice were weighed, followed by food removal at 1 pm for 24hr with ad lib water access. Twenty-four hours post-fasting, mice were weighed, followed by experimentation.

Sample Preparation:

Sampleprep ID:SP002329
Sampleprep Summary:Mouse cortical lysate samples were rapidly isolated, and flash frozen in liquid nitrogen. Soluble metabolites were extracted directly from tissue using cold methanol/water (80/20, v/v) at approximately 1μL per 50μg of tissue, followed by ultrasonication (Branson Sonifier 250) for 15 s. Homogenized samples were incubated at −80 °C to precipitate proteins and subsequently centrifuged at 18,000xg for 20 min at 4 °C to pellet the debris. The supernatants containing soluble metabolites were collected in new tubes and evaporated to dryness using a SpeedVac concentrator (Thermo Savant). Next, the metabolites were reconstituted in acetonitrile/water (60/40, v/v), vortex-mixed, and centrifuged at 18,000xg for 30 min at 4°C to remove debris.

Combined analysis:

Analysis ID AN003650
Analysis type MS
Chromatography type HILIC
Chromatography system Thermo Dionex Ultimate 3000
Column Waters Xbridge Amide (100 x 3mm,3.5um)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Q Exactive Orbitrap
Ion Mode UNSPECIFIED
Units Peak area

Chromatography:

Chromatography ID:CH002704
Chromatography Summary:Samples were analyzed by High-Performance Liquid Chromatography and High-Resolution Mass Spectrometry and Tandem Mass Spectrometry (HPLC-MS/MS). Specifically, system consisted of a Thermo Q-Exactive in line with an electrospray source and an Ultimate3000 (Thermo) series HPLC consisting of a binary pump, degasser, and auto-sampler outfitted with a Xbridge Amide column (Waters; dimensions of 3.0 mm × 100 mm and a 3.5 µm particle size). The mobile phase A contained 95% (vol/vol) water, 5% (vol/vol) acetonitrile, 10 mM ammonium hydroxide, 10 mM ammonium acetate, pH = 9.0; B was 100% Acetonitrile. The gradient was as following: 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1-18 min, 75% A; 18-25 min, 15% A with a flow rate of 150 μL/min. The capillary of the ESI source was set to 275 °C, with sheath gas at 35 arbitrary units, auxiliary gas at 5 arbitrary units and the spray voltage at 4.0 kV. In positive/negative polarity switching mode, an m/z scan range from 60 to 900 was chosen and MS1 data was collected at a resolution of 70,000. The automatic gain control (AGC) target was set at 1 × 106 and the maximum injection time was 200 ms. The top 5 precursor ions were subsequently fragmented, in a data-dependent manner, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. Besides matching m/z, metabolites are identified by matching either retention time with analytical standards and/or MS2 fragmentation pattern. Data acquisition and analysis were carried out by Xcalibur 4.1 software and Tracefinder 4.1 software, respectively (both from Thermo Fisher Scientific).
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Waters Xbridge Amide (100 x 3mm,3.5um)
Flow Gradient:0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1-18 min, 75% A; 18-25 min, 15% A
Flow Rate:150 µL/min
Solvent A:95% water/5% acetonitrile; 10 mM ammonium hydroxide; 10 mM ammonium acetate, pH 9.0
Solvent B:100% acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS003401
Analysis ID:AN003650
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:The capillary of the ESI source was set to 275 °C, with sheath gas at 35 arbitrary units, auxiliary gas at 5 arbitrary units and the spray voltage at 4.0 kV. In positive/negative polarity switching mode, an m/z scan range from 60 to 900 was chosen and MS1 data was collected at a resolution of 70,000. The automatic gain control (AGC) target was set at 1 × 106 and the maximum injection time was 200 ms. The top 5 precursor ions were subsequently fragmented, in a data-dependent manner, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. Besides matching m/z, metabolites are identified by matching either retention time with analytical standards and/or MS2 fragmentation pattern. Data acquisition and analysis were carried out by Xcalibur 4.1 software and Tracefinder 4.1 software, respectively (both from Thermo Fisher Scientific).
Ion Mode:UNSPECIFIED
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