TY - JOUR

T1 - Brainstem transcriptomic changes in male Wistar rats after acute stress, comparing the use of duplex specific nuclease (DSN)

AU - Lanshakov, Dmitriy A

AU - Sukhareva, Ekaterina V

AU - Bulygina, Veta V

AU - Khozyainova, Anna A

AU - Gerashchenko, Tatiana S

AU - Denisov, Evgeny V

AU - Kalinina, Tatyana S

N1 - Studies was supported by joint research project FWNR-2022-0002. RNA-sequencing was carried out in The Core Facility «Medical genomics»(Tomsk NRMC) and the Tomsk Regional Common Use Center. Bioinformatics analysis was carried out at the Information and Computing Center of Novosibirsk State University © 2024. The Author(s).

PY - 2024/12

Y1 - 2024/12

N2 - In this work, we have analyzed the transcriptomic changes in the brainstem of male Wistar rats 2 h after an acute stress exposure. We performed duplex-specific nuclease normalization of cDNA libraries and compared the results back-to-back for the first time. Based on our RNAseq data, we selected reference genes for RT-qPCR that are best suited for acute stress experiments. Most genes were upregulated. We detected a massive shift in neuropeptide Crh, Trh,Cga, Tshb, Uts2b, Tac4, Lep and neuropeptide receptor Hcrtr1, Sstr5, Bdkrb2, Crhr2 signaling, as well as glutamate Grin3b, Grm2 and GABA Gpr156, acetylcholine Chrm4,Chrne, adrenergic Adra2b receptors expression. A strong increase in the expression of intermediate filaments Krt83/Krt86/Krt80/Krt84/Krt87/Krt4/Krt76 and motor proteins Myo7a, Klc3 was detected. Remarkably, in the absence of astrocyte activation, we also observed signs of microglial activation at this time point. Both expression of anti-inflammatory cytokines Il13, Ccl24 and pro-inflammatory cytokine receptors Il9r, Il12rb1, Tnfrsf14, Tnfrsf13c, Tnfrsf25, Tnfrsf1b were increased. In the Wnt signaling pathway, we observed increased expression of ligands-receptors Wnt1, Wnt11, Ror2 and also negative regulators Notum, Sfrp5, Sost. RNAseq results after DSN treatment correlated at a high level with RNAseq results without DSN, but there was a proportion of genes that shifted their logFC values. They are mostly rare transcripts TPM 1-10 with higher 0.5-0.9 GC content.

AB - In this work, we have analyzed the transcriptomic changes in the brainstem of male Wistar rats 2 h after an acute stress exposure. We performed duplex-specific nuclease normalization of cDNA libraries and compared the results back-to-back for the first time. Based on our RNAseq data, we selected reference genes for RT-qPCR that are best suited for acute stress experiments. Most genes were upregulated. We detected a massive shift in neuropeptide Crh, Trh,Cga, Tshb, Uts2b, Tac4, Lep and neuropeptide receptor Hcrtr1, Sstr5, Bdkrb2, Crhr2 signaling, as well as glutamate Grin3b, Grm2 and GABA Gpr156, acetylcholine Chrm4,Chrne, adrenergic Adra2b receptors expression. A strong increase in the expression of intermediate filaments Krt83/Krt86/Krt80/Krt84/Krt87/Krt4/Krt76 and motor proteins Myo7a, Klc3 was detected. Remarkably, in the absence of astrocyte activation, we also observed signs of microglial activation at this time point. Both expression of anti-inflammatory cytokines Il13, Ccl24 and pro-inflammatory cytokine receptors Il9r, Il12rb1, Tnfrsf14, Tnfrsf13c, Tnfrsf25, Tnfrsf1b were increased. In the Wnt signaling pathway, we observed increased expression of ligands-receptors Wnt1, Wnt11, Ror2 and also negative regulators Notum, Sfrp5, Sost. RNAseq results after DSN treatment correlated at a high level with RNAseq results without DSN, but there was a proportion of genes that shifted their logFC values. They are mostly rare transcripts TPM 1-10 with higher 0.5-0.9 GC content.

KW - Animals

KW - Male

KW - Rats, Wistar

KW - Rats

KW - Transcriptome

KW - Brain Stem/metabolism

KW - Gene Expression Profiling

KW - Endonucleases/metabolism

KW - Stress, Physiological

UR - https://www.scopus.com/record/display.uri?eid=2-s2.0-85204511969&origin=inward&txGid=720a489d51181b91b2a8c161ce467385

U2 - 10.1038/s41598-024-73042-2

DO - 10.1038/s41598-024-73042-2

M3 - Article

C2 - 39300279

VL - 14

JO - Scientific Reports

JF - Scientific Reports

SN - 2045-2322

IS - 1

M1 - 21856

ER -