Journal: Advanced Science

Article Title: Gonadotropin‐Inhibitory Hormone (GnIH) and Its Receptor GPR147 Mediates Acute Stress‐Induced Activation of the Locus Coeruleus‐SAM Axis

doi: 10.1002/advs.202518996

Figure Lengend Snippet: Primer sequences for quantitative real‐time PCR.

Article Snippet: The following primary antibodies were used: Tyrosine Hydroxylase (TH; Cat. sc‐25269, Santa Cruz, 1:100 dilution; immunogen: human TH amino acids 1–196; species reactivity: mouse; label dopaminergic cells); GPR147 (Cat. Orb157323, Biorbyt, 1:250 dilution; immunogen: keyhole limpet hemocyanin (KLH)‐conjugated synthetic peptide derived from human GPR147/NPFF1R; species reactivity: mouse); GnIH (raised in our laboratory, 1:500 dilution; immunogen: KLH‐conjugated synthetic peptide derived from mouse NPVF/RFRP amino acids); choline O‐acetyltransferase (ChAT; Cat. AMAB91130, Sigma, 1:1000 dilution; immunogen: human ChAT; species reactivity: mouse); neuropeptide Y (NPY; Cat. YP‐Ab‐12776, UpingBio, 1:100 dilution; immunogen: synthetic peptide derived from human NPY amino acids 45–94; species reactivity: mouse); and phenylethanolamine N‐methyltransferase (PNMT; Cat. sc‐393995, Santa Cruz, 1:100 dilution; immunogen: the internal region of human PNMT, amino acids 164–212; species reactivity: mouse; label adrenergic cells).

Techniques: Real-time Polymerase Chain Reaction

Journal: Advanced Science

Article Title: Gonadotropin‐Inhibitory Hormone (GnIH) and Its Receptor GPR147 Mediates Acute Stress‐Induced Activation of the Locus Coeruleus‐SAM Axis

doi: 10.1002/advs.202518996

Figure Lengend Snippet: Activation of the GnIH/GPR147 System in the LC‐SAM Axis across Acute Restraint Stress Paradigms. (A to D): the concentration of CORT (A), NE (B), EPI (C), and GnIH (D) in the serum of mice under restraint stress (RS). (E and F): (E) The representative immunohistochemistry images and (F) mean densitometric values of GnIH or GPR147 in the locus coeruleus in acute restraint stress paradigms. (G and H): GnIH , GPR147 , TH mRNA (G) and TH protein (H) levels in the locus coeruleus across acute restraint stress paradigms. (I and J): (I) Representative immunohistochemistry images and (J) mean densitometric values of GnIH or GPR147 in the adrenal gland in acute restraint stress paradigms. (K and L): GnIH , GPR147 , TH , PNMT mRNA (K) and TH protein (L) in the locus coeruleus across diverse acute stress paradigms, including RS. (M) Optogenetic experiments demonstrated the expression of TH and GnIH neurons in the LC after the activation of norepinephrine neurons. Scale bar, 200 µm. (N and O) the concentration of GnIH (N) and NE (O) in the serum of mice after the activation of norepinephrine neurons. CORT: corticosterone; NE: norepinephrine; EPI: epinephrine; LC: locus coeruleus; TH: tyrosine hydroxylase; PNMT: phenylethanolamine N‐methyltransferase. GAPDH and β‐tubulin was used as an internal control. Data are mean ± SEM (n = 10 biological replicates per group). Comparisons between two groups were performed using two‐sided unpaired Student's t ‐test with α = 0.05. For multiple comparisons, one‐way ANOVA followed by Tukey's post‐hoc test was used. Effect sizes (Cohen's d) were calculated for all significant comparisons. * p < 0.05, ** p < 0.01, compared to the control group.

Article Snippet: The following primary antibodies were used: Tyrosine Hydroxylase (TH; Cat. sc‐25269, Santa Cruz, 1:100 dilution; immunogen: human TH amino acids 1–196; species reactivity: mouse; label dopaminergic cells); GPR147 (Cat. Orb157323, Biorbyt, 1:250 dilution; immunogen: keyhole limpet hemocyanin (KLH)‐conjugated synthetic peptide derived from human GPR147/NPFF1R; species reactivity: mouse); GnIH (raised in our laboratory, 1:500 dilution; immunogen: KLH‐conjugated synthetic peptide derived from mouse NPVF/RFRP amino acids); choline O‐acetyltransferase (ChAT; Cat. AMAB91130, Sigma, 1:1000 dilution; immunogen: human ChAT; species reactivity: mouse); neuropeptide Y (NPY; Cat. YP‐Ab‐12776, UpingBio, 1:100 dilution; immunogen: synthetic peptide derived from human NPY amino acids 45–94; species reactivity: mouse); and phenylethanolamine N‐methyltransferase (PNMT; Cat. sc‐393995, Santa Cruz, 1:100 dilution; immunogen: the internal region of human PNMT, amino acids 164–212; species reactivity: mouse; label adrenergic cells).

Techniques: Activation Assay, Concentration Assay, Immunohistochemistry, Expressing, Control