Journal: Science Advances

Article Title: Neural circuitry mediating hypercapnia-induced microarousal and impaired sleep depth

doi: 10.1126/sciadv.aei1892

Figure Lengend Snippet: ( A ) Immunofluorescence staining for c-Fos (magenta) and AVP (green) in the PVN of mice exposed to NA (top) or HA (bottom). ( B ) RNAscope in situ hybridization for Fos (magenta) and Crh (green) mRNAs in the PVN of mice exposed to NA (top) or HA (bottom). ( C ) Top: Experimental schematic. Bottom: Representative images showing the recorded PVN neuron (biocytin, magenta; CRH, red). ( D ) oEPSC traces under pharmacological conditions. Gray traces indicate individual recordings, and black traces indicate mean traces ( n = 6 cells from five mice). ( E ) Time course and quantification of oEPSC amplitude normalized to the 10-min baseline. ( F ) Schematic of viral injection for chemogenetic inhibition of PVN CRH neurons and representative viral expression. ( G ) Effect of saline or CNO (1 mg/kg) treatment on HA-induced arousal. Left: Latency to wakefulness following HA onset. Right: Proportion of trials in which sleep was maintained over time after HA onset ( n = 8). ( H ) Representative recordings after saline or CNO (1 mg/kg) injection, showing EEG spectrogram, EEG, EMG, and respiratory signals during HA exposure. ( I ) Distribution of Crh (magenta) and Slc17a6 (green) mRNAs in the PVN. ( J ) Proportion of Crh + cells coexpressing Slc17a6 ( n = 17 sections from seven mice). ( K ) Top: Schematic of CRISPR-mediated gene knockdown in the PVN of CRH-Cre mice. Bottom: Representative viral expression and Slc17a6 RNAscope signals. ( L ) Representative recordings from the sgScramble and sgSlc17a6 mice during HA exposure. ( M ) Effect of Slc17a6 knockdown in CRH neurons on wake latency and sleep-maintenance probability after HA onset (sgScramble, n = 5; sgSlc17a6, n = 6). Orange shading indicates HA exposure. Data are presented as the means ± SEM. ns, not significant. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: Optogenetically evoked EPSCs (oEPSCs) were triggered by 2-ms pulses of 470-nm blue light (CoolLED pE-300 ultra) delivered every 20 s. To verify monosynaptic connections, pharmacological dissection was performed by sequential perfusion of TTX (1 μM; Mreda, Beijing, China, catalog no. M046335-1 mg), 4-AP (500 μM; MedChemExpress, catalog no. HY-B0604), and a cocktail of NBQX (10 μM; MedChemExpress, catalog no. HY-15068) and D-AP5 (50 μM; MedChemExpress, catalog no. HY-100714A).

Techniques: Immunofluorescence, Staining, RNAscope, In Situ Hybridization, Injection, Inhibition, Expressing, Saline, CRISPR, Knockdown

Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: Relative HSP abundances in whole skeletal muscle homogenates from young adults and older adults pre and post HIT exercise. Representative Westen blots of (A) HSP72, HSP27, and αB-crystallin and (B) phosphorylated HSP27 Ser15 (pHSP27 Ser15) and pαB-crystallin Ser59 in whole muscle homogenates from the vastus lateralis of the same individuals. Calibration curves of mixed muscle homogenates are indicated and were used to determine the relative number of given proteins (see Methods). Stain-free gels are indicative of total protein loading, and molecular weights are indicated by markers collected under white light capture without moving the membrane between that and chemiluminescence detection. Relative abundances of (C) HSP72, (D) HSP27, (E) pHSP27 Ser15, (F) αB-crystallin, and (G) pαB-crystallin Ser59 from young (circle) and older adults Pre (square) and older adults Post (triangle) HIT exercise are shown relative to average Old (pre) on a given gel (data are presented as mean ± SD). Individuals indicated by the number of symbols ( n : 5–7), with the same color assigned to the same individual and consistent across all graphs. * p ≤ 0.05 indicates Brown-Forsye and Welch’s and post hoc analysis using Games-Horwell. HIT = high-intensity training; HSP = heat shock protein; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques: Staining, Membrane

Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: HSP abundances in type I and II skeletal muscle fibers from young and older adults. (A, C, and F) The MHC isoform present was determined in individual muscle fiber segments from the vastus lateralis and, following pooling into type I and type II groups from a given biopsy, were analyzed by Westen blotting. Westen blots of (A) HSP72, (C) HSP27 and pHSP27 Ser15, (F) αB-crystallin and pαB-crystallin Ser59, with MHC isoforms in groups of fibers. Stain-free gels are indicative of total protein loading, and molecular weights are indicated by markers collected under white light capture without moving the membrane between that and chemiluminescence detection. Calibration curves of mixed muscle homogenates are indicated. Relative protein abundances of (B) HSP72, (D) HSP27, (E) pHSP27 Ser15, (G) αB-crystallin, and (H) pαB-crystallin Ser59 in fibers from young (circle) and older adults (square) type I fibers (no outline) and type II fibers (outline). All fibers are expressed relative to the average older adult’s type I fibers. The same color is assigned to the same individual and is consistent with (data are presented as mean ± SD). * p < 0.05 and ** p < 0.01, mixed effect model Univariant using either Tukey’s or Games-Horwell’s multiple comparison test (see Methods). HIT = high-intensity training; HSP = heat shock protein; MHC = myosin heavy chain; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques: Staining, Membrane, Comparison

Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: HSP abundances in type I and II skeletal fibers from older adults pre- and post HIT exercise. Relative protein abundances of (A and B) HSP72, (C and D) HSP27, (E and F) pHSP27 Ser15, (G and H) αB-crystallin, and (I and J) pαB-crystallin Ser59 in fibers from old pre and old post HIT exercise. All fibers are expressed relative to the average old pre type I fibers or relative pre type II depending on fiber type. The same color is assigned to the same Individual, consistent in both graphs and all figures. * p < 0.05 and ** p < 0.01 indicated significant difference in paired t -test (except pHSP27 Ser15 Wilcoxon match-pair rank test). Representative blots are shown in . HIT = high-intensity training; HSP = heat shock protein; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques: