Journal: Scientific Reports

Article Title: IL-4 Protects the Mitochondria Against TNFα and IFNγ Induced Insult During Clearance of Infection with Citrobacter rodentium and Escherichia coli

doi: 10.1038/srep15434

Figure Lengend Snippet: Immunohistochemical staining using antibodies for ( A ) MTND6 (complex I) ( B ) SDHA (complex-II) ( C ) CYC1 (complex-III) ( D ) CCO-VIc (complex-IV). Statistics: unpaired t test. * P < 0.05, ** P < 0.01, *** P < 0.001 vs. control, n = 5–6 mice/group. The infection experiments were performed twice, and each time point contain results pooled from 5–6 mice. Scale bar 100 μm, magnification × 200.

Article Snippet: Rabbit polyclonal primary antibodies for subunits of the mitochondrial respiratory enzyme complexes were selected for complex I (anti-NADH dehydrogenase subunit 6, MTND6, orb6548, Biorbyt), complex-II (anti-Succinate dehydrogenase subunit A, SDHA, ab86932, Abcam), complex-III (anti-Ubiquinol-cytochrome c reductase complex cytochrome c1 subunit, CYC1, NBP1-86872, Novus Biologicals), complex-IV (anti-Cytochrome c oxidase subunit VIc ,CCO-VIc, ab150422, Abcam), Caspase-3 (activated form, ab4051, Abcam) and 3-Nitrotyrosine (AB5411, Millipore) at optimal dilutions of 1:2000, 1:200, 1:2000, 1:2000, 1:500 and 1:500 respectively, in antibody diluent (Dako, S0809).

Techniques: Immunohistochemical staining, Staining, Control, Infection

Journal: Scientific Reports

Article Title: IL-4 Protects the Mitochondria Against TNFα and IFNγ Induced Insult During Clearance of Infection with Citrobacter rodentium and Escherichia coli

doi: 10.1038/srep15434

Figure Lengend Snippet: Immunohistochemical staining using antibodies for ( A ) MTND6 (complex I) ( B ) SDHA (complex-II) ( C ) CYC1 (complex-III) ( D ) CCO-VIc (complex-IV). Values are mean ± S.E.M. Statistics: ANOVA with Student Newman-Keuls Multiple Comparison post hoc test: * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: Rabbit polyclonal primary antibodies for subunits of the mitochondrial respiratory enzyme complexes were selected for complex I (anti-NADH dehydrogenase subunit 6, MTND6, orb6548, Biorbyt), complex-II (anti-Succinate dehydrogenase subunit A, SDHA, ab86932, Abcam), complex-III (anti-Ubiquinol-cytochrome c reductase complex cytochrome c1 subunit, CYC1, NBP1-86872, Novus Biologicals), complex-IV (anti-Cytochrome c oxidase subunit VIc ,CCO-VIc, ab150422, Abcam), Caspase-3 (activated form, ab4051, Abcam) and 3-Nitrotyrosine (AB5411, Millipore) at optimal dilutions of 1:2000, 1:200, 1:2000, 1:2000, 1:500 and 1:500 respectively, in antibody diluent (Dako, S0809).

Techniques: Immunohistochemical staining, Staining, Comparison

Journal: Scientific Reports

Article Title: IL-4 Protects the Mitochondria Against TNFα and IFNγ Induced Insult During Clearance of Infection with Citrobacter rodentium and Escherichia coli

doi: 10.1038/srep15434

Figure Lengend Snippet: Immunohistochemical staining using antibodies for ( A ) MTND6 (complex I) ( B ) SDHA (complex-II) ( C ) CYC1 (complex-III) ( D ) CCO-VIc (complex-IV). Values are mean ± S.E.M. Statistics: ANOVA with Student Newman-Keuls Multiple Comparison post hoc test: * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: Rabbit polyclonal primary antibodies for subunits of the mitochondrial respiratory enzyme complexes were selected for complex I (anti-NADH dehydrogenase subunit 6, MTND6, orb6548, Biorbyt), complex-II (anti-Succinate dehydrogenase subunit A, SDHA, ab86932, Abcam), complex-III (anti-Ubiquinol-cytochrome c reductase complex cytochrome c1 subunit, CYC1, NBP1-86872, Novus Biologicals), complex-IV (anti-Cytochrome c oxidase subunit VIc ,CCO-VIc, ab150422, Abcam), Caspase-3 (activated form, ab4051, Abcam) and 3-Nitrotyrosine (AB5411, Millipore) at optimal dilutions of 1:2000, 1:200, 1:2000, 1:2000, 1:500 and 1:500 respectively, in antibody diluent (Dako, S0809).

Techniques: Immunohistochemical staining, Staining, Comparison

Journal: Scientific Reports

Article Title: IL-4 Protects the Mitochondria Against TNFα and IFNγ Induced Insult During Clearance of Infection with Citrobacter rodentium and Escherichia coli

doi: 10.1038/srep15434

Figure Lengend Snippet: Immunohistochemical staining using antibodies for ( A ) MTND6 (complex I) ( B ) SDHA (complex-II) ( C ) CYC1 (complex-III) ( D ) CCO-VIc (complex-IV). Statistics: ANOVA Student Newman-Keuls Multiple Comparison post hoc test: * P < 0.05, ** P < 0.01, *** P < 0.001. vs. control.

Article Snippet: Rabbit polyclonal primary antibodies for subunits of the mitochondrial respiratory enzyme complexes were selected for complex I (anti-NADH dehydrogenase subunit 6, MTND6, orb6548, Biorbyt), complex-II (anti-Succinate dehydrogenase subunit A, SDHA, ab86932, Abcam), complex-III (anti-Ubiquinol-cytochrome c reductase complex cytochrome c1 subunit, CYC1, NBP1-86872, Novus Biologicals), complex-IV (anti-Cytochrome c oxidase subunit VIc ,CCO-VIc, ab150422, Abcam), Caspase-3 (activated form, ab4051, Abcam) and 3-Nitrotyrosine (AB5411, Millipore) at optimal dilutions of 1:2000, 1:200, 1:2000, 1:2000, 1:500 and 1:500 respectively, in antibody diluent (Dako, S0809).

Techniques: Immunohistochemical staining, Staining, Comparison, Control

Journal: bioRxiv

Article Title: Reuniens transiently synchronizes memory networks at beta frequencies

doi: 10.1101/2022.06.21.497087

Figure Lengend Snippet: A) A linear track was used with odor ports located at opposite ends where two separate four-odor sequences (A, B, C, D or W, X, Y, Z) were presented. Rats had to correctly identify the odor as either InSeq (hold ≥ 1 s) or OutSeq (hold < 1 s). B) SMI was not significantly different between Seq 1 and Seq 2. Individual rat performance is indicated by circles. C) Average running speeds less than 10 cm/s represent 80% of the data. D) Representative spectrogram with corresponding filtered LFP from theta and beta bands in prefrontal cortex (mPFC, blue) and hippocampus (HC, red) during both sequences interleaved with a short running bout. High power beta is aligned to the odor sampling period. E) Sample raw LFP (two samples from each rat within the experiment) in prefrontal cortex (blue) and hippocampus (red). Each rat is indicated with a different shade of color. Beta bursts are highlighted in gray and can be seen only during the trial (after poke in). F i ) Prefrontal-hippocampal coherence was significantly different between running periods (non-memory) and memory (odor-trials). Inset shows a difference between memory and running. Beta synchrony increases during memory while theta was lower (close to zero). F ii ) AUC shows that theta coherence was not significantly different between running and memory, while beta coherence was significantly different between running and memory. G i ) Sample bandpass beta filtered from prefrontal and hippocampal sites with a sample of 10 trials shows closely matched high amplitude beta ∼100ms after the poke in. G ii ) A zoomed in trial shows the bursty properties of beta. H) The probability of prefrontal and hippocampal beta burst occurrence are not significantly different. I) Average duration of a beta burst was not significantly different between prefrontal and hippocampal sites. J) The latency to the first beta burst was not significantly different between prefrontal and hippocampal sites. K) Prefrontal-hippocampal coherence separated based on sequential context (InSeq vs OutSeq). L) Coherence between InSeq and OutSeq trials separated based on accuracy. M) Beta AUC was significantly different across the four trail types. InSeq correct trials was significantly higher compared to InSeq incorrect , OutSeq correct , and OutSeq incorrect . Theta AUC was not significantly different across the four trial types. Abbreviations: mPFC, medial prefrontal cortex; HC, hippocampus, InSeq, in-sequence; OutSeq, out-of-sequence; SMI, sequence memory index; Seq, sequence; LFP, local field potential; AUC, area under the curve; InSeq correct , in-sequence correct, InSeq incorrect , in-sequence incorrect, OutSeq correct , out-of-sequence correct, OutSeq incorrect , out-of-sequence incorrect; θ, theta; β, beta; ns, not significant.

Article Snippet: We injected bilaterally 600μL channelrhodopsin-infused retrograde adeno associated virus pAAV-Syn-ChR2(H134R)-GFP (Addgene) in ventral hippocampus (A/P: -5.6, M/L: ±5.85; D/V: -7.4, 200μL, -6.8, 200 μL & -6.2, 100 μL) at a speed of 1.0 nL/sec (Nanoject III, Drummund Scientific).

Techniques: Sampling, Sequencing

Journal: bioRxiv

Article Title: Reuniens transiently synchronizes memory networks at beta frequencies

doi: 10.1101/2022.06.21.497087

Figure Lengend Snippet: A) SMI was not significantly different between Seq 1 and Seq 2. Individual rat performance is indicated by circles. B) Representative spectrogram with corresponding filtered LFP from theta and beta bands in reuniens (RE; purple) during both sequences with a sample running bout in between. High power beta is aligned to the odor sampling period. C i ) Reuniens-hippocampal coherence was significantly different between running periods (non-memory) and memory (odor-trials). Inset shows a difference (memory – running) and show that beta synchrony increases during memory while theta was lower (close to zero). C ii ) AUC shows that theta coherence was not significantly different between running and memory, while beta coherence was significantly different between running and memory. D) Sample raw LFP (two samples from each rat within the experiment) in reuniens (purple) and hippocampus (red). Each rat is indicated with a different shade of color. Beta bursts are highlighted in gray and can be seen only during the trial (after poke in). E i ) Sample bandpass beta filtered from reuniens and hippocampal sites with a sample of 10 trials shows reuniens beta occurs earlier than hippocampus beta. E ii ) A zoomed in trial shows the bursty properties of beta with reuniens beta occurring earlier in the trial. F) The probability of reuniens and hippocampal beta burst occurrence are significantly different with reuniens occurring earlier indicated with purple arrow. G) Average duration of a beta burst was significantly different longer in reuniens than hippocampal sites. H) The latency to the first beta burst was significantly earlier in reuniens than hippocampal sites. I) Reuniens-hippocampal coherence separated based on sequential context (InSeq vs OutSeq). J) Coherence between InSeq and OutSeq trials separated based on accuracy. K) Beta AUC was significantly different across the four trail types. InSeq correct trials were significantly higher compared to OutSeq correct and OutSeq correct trials were significantly lower than InSeq incorrect . Theta AUC was not significantly different across the four trial types. Abbreviations: RE, nucleus reuniens; HC, hippocampus, SMI, sequence memory index; Seq, sequence; LFP, local field potential; AUC, area under the curve; InSeq, in-sequence; OutSeq, out-of-sequence; InSeq correct , in-sequence correct, InSeq incorrect , in-sequence incorrect, OutSeq correct , out-of-sequence correct, OutSeq incorrect , out-of-sequence incorrect; θ, theta; β, beta; ns, not significant.

Article Snippet: We injected bilaterally 600μL channelrhodopsin-infused retrograde adeno associated virus pAAV-Syn-ChR2(H134R)-GFP (Addgene) in ventral hippocampus (A/P: -5.6, M/L: ±5.85; D/V: -7.4, 200μL, -6.8, 200 μL & -6.2, 100 μL) at a speed of 1.0 nL/sec (Nanoject III, Drummund Scientific).

Techniques: Sampling, Sequencing

Journal: bioRxiv

Article Title: Reuniens transiently synchronizes memory networks at beta frequencies

doi: 10.1101/2022.06.21.497087

Figure Lengend Snippet: A) A sample of the upper envelope for both beta and theta for all three regions (prefrontal cortex, reuniens, and hippocampus). B) Beta amplitudes in reuniens start rising just prior to the onset of trials indicated with an arrow. Prefrontal cortex and hippocampus beta simultaneously rise after the trial. Overall showing a main effect of region, time and region x time. Inset plot shows that the hippocampal recordings from both data sets were not significantly different. C i ) Calculated slopes show that in prefrontal cortex beta significantly rose ∼300 ms after the trial and then decreased after the poke out ( p ’s < 0.01, indicated with a star). C ii ) In reuniens beta slopes significantly rose ∼100 ms before the onset of the trial and continued to rise during the trial ( p ’s < 0.01, indicated with a star). C iii ) In hippocampus beta slopes shows similar patterns to prefrontal cortex, rising ∼300 ms after the trial ( p ’s < 0.01, indicated with a star). D) Theta amplitudes in reuniens and hippocampus show phasic increases before a trial followed by a phasic increase in prefrontal cortex at the start of the trial. During the trial there is a decrease in theta indicated with an arrow. Overall showing a main effect of region, time and region x time. Inset plot shows that the hippocampal recordings from both data sets were not significantly different. E i ) Calculated slopes show that in prefrontal cortex theta significantly rose ∼300 ms before the start of the trial and ∼200 ms before the poke out (end of trial), while decreasing during the trial ( p ’s < 0.01, indicated with a star). E ii ) In reuniens theta slopes significantly rose ∼500 ms before the onset of the trial and decreased during the trial ( p ’s < 0.01, indicated with a star). E iii ) In hippocampus theta slopes shows similar patterns to reuniens and prefrontal cortex where theta slopes decrease during the trial ( p ’s < 0.01, indicated with a star). F i ) A sample of prefrontal cortex beta voltage (blue) superimposed with theta voltage (dark blue) shows that beta and theta are weakly coupled. Example phase-amplitude plot from a rat showing beta amplitude was weakly modulated by theta phase. F ii ) A sample of prefrontal cortex beta voltage (blue) superimposed with delta voltage (dark blue) shows that beta and delta are strongly coupled. Example phase-amplitude plot from a rat showing beta amplitude was more modulated by delta phase. G i ) A sample of reuniens beta voltage (purple) superimposed with theta voltage (dark purple) shows that beta and theta are weakly coupled. Example phase-amplitude plot from a rat showing beta amplitude was weakly modulated by theta phase. G ii ) A sample of reuniens beta voltage (purple) superimposed with delta voltage (dark purple) shows that beta and delta are strongly coupled. Example phase-amplitude plot from a rat showing beta amplitude was more modulated by delta phase. H i ) A sample of hippocampal beta voltage (red) superimposed with theta voltage (dark red) shows that beta and theta are coupled. Example phase-amplitude plot from a rat showing beta amplitude was modulated by theta phase. H ii ) A sample of hippocampal beta voltage (red) superimposed with delta voltage (dark red) shows that beta and delta are coupled. Example phase-amplitude plot from a rat showing beta amplitude was modulated by delta phase. Yellow waveforms represent the sine waves fitted to the amplitude– phase distributions. Abbreviations: mPFC, medial prefrontal cortex; RE, nucleus reuniens; HC, hippocampus; Exp, experiment; θ, theta; δ, delta; β, beta.

Article Snippet: We injected bilaterally 600μL channelrhodopsin-infused retrograde adeno associated virus pAAV-Syn-ChR2(H134R)-GFP (Addgene) in ventral hippocampus (A/P: -5.6, M/L: ±5.85; D/V: -7.4, 200μL, -6.8, 200 μL & -6.2, 100 μL) at a speed of 1.0 nL/sec (Nanoject III, Drummund Scientific).

Techniques:

Journal: bioRxiv

Article Title: Reuniens transiently synchronizes memory networks at beta frequencies

doi: 10.1101/2022.06.21.497087

Figure Lengend Snippet: A i ) Bilateral injections of retrograde pAAV-Syn-ChR2(H134R)-GFP (experimental) or AAV-CAGGFP (controls) were delivered in vCA1 for the retrograde expression of RE neurons. An optic fiber above reuniens stimulated channelrhodopsin or fluorescence protein expressing neurons. A ii ) Confocal image (60x) showing retrograde expression of ChR2 in RE neurons (indicated by white arrows). B) LED based blue light pulse or sinewave stimulations at 1,2,4,8, 20 or 50 Hz were used to activate RE-ChR2 transduced and control rats. C) Averaged evoked responses in ventral hippocampus resulting from reuniens 8 Hz pulse blue light stimulations (60 ms each) in a control and an experimental subject. As illustrated, stimulating RE-ChR2 expressing neurons resulted in two consecutive negative deflections suggestive of a direct monosynaptic response in vCA1 (in red). This effect was not seen in controls (black). Mild to moderate LFP changes in prefrontal activity were also observed (blue) which may be the result of stimulating putative reuniens mPFC-HC dual projecting neurons expressing ChR2. D) Sample averaged raw traces in prefrontal cortex (blue) and hippocampus (red; two samples from each experimental rat) showing a predominant beta rhythm one second prior and one second after the last optogenetic pulse (two samples from each rat). Gray boxes highlight areas of high beta activity. E) z -scored mPFC-HC coherence in RE-ChR2 and controls across three time points: E i ) one min before where all rats have similar theta and beta amplitude values, E ii ) during blue light stimulation where there are marked increases in the theta and beta band and a decrease in theta in experimental rats but not controls, and E iii ) one min after stimulations where coherence is similar to E ii . Notably, stimulating RE-ChR2 neurons with a 20 Hz sine resulted in a large increase in beta (as previously described) but with an additional large peak around the 20Hz frequency that surpassed the amplitude of delta and theta bands ( E ii inset, black arrow), this effect was not seen before or after stimulations ( E i & E iii inset). F) Theta AUC coherence was significantly different across conditions (control, pulse, sine) and time windows (before, during, after). Notably, theta coherence in control was similar across all time points. G) Beta AUC coherence was significantly different across conditions and time windows. In controls, beta coherence was similar across all time points in controls. Abbreviations: AUC, area under the curve; ChR2, channelrhodopsin, HC, hippocampus; min, minute; ms, milliseconds; mPFC, medial prefrontal cortex; RE, reuniens; vCA1, ventral CA1.

Article Snippet: We injected bilaterally 600μL channelrhodopsin-infused retrograde adeno associated virus pAAV-Syn-ChR2(H134R)-GFP (Addgene) in ventral hippocampus (A/P: -5.6, M/L: ±5.85; D/V: -7.4, 200μL, -6.8, 200 μL & -6.2, 100 μL) at a speed of 1.0 nL/sec (Nanoject III, Drummund Scientific).

Techniques: Expressing, Fluorescence, Control, Activity Assay