Journal: Science signaling

Article Title: A role for corticotrophin releasing factor signaling in the lateral habenula and its modulation by early life stress *

doi: 10.1126/scisignal.aan6480

Figure Lengend Snippet: MD increased the excitability of LHb neurons. A. Top: Representative spontaneous AP recordings of LHb neurons from non-MD and MD rats using cell-attached voltage clamp recordings. Bottom: Percent of spontaneously active LHb neurons (left) and mean AP frequency (right) from each group (n= 33 cells from 18 rats or 29 cells from 19 rats, respectively). B. Representative traces (top) and number of APs (bottom) recorded from LHb neurons using whole cell patch clamp recording, with synaptic transmission intact, in slices from non-MD and MD rats in response to depolarizing current steps (calibration bar: 20mV/1s) [n= 24 cells from 11 rats (non-MD) or 24 cells from 9 rats (MD), F(1,460)=1.69]C. As described in (B), with fast synaptic transmission blocked [n= 25 cells from 17 rats (non-MD) or 31 cells from 18 rats (MD), F(1,600)=35.84]. D: Average amplitude of fAHP, mAHP and Rin derived from AP recordings in (C). E: Representative Western blots and quantitation of SK2 abundance (β-actin: loading control) in LHb tissue homogenates from non-MD and MD rats (n= 6 biological replicates). Data are means ± SEM. *P<0.05, **P<0.01 by unpaired Student's t-test; ****P<0.0001 by two-way ANOVA.

Article Snippet: Separated proteins were transferred onto nitrocellulose membranes, blocked with casein-based blocking reagent (I-Block, Life Technologies) for 60 minutes at room temperature and then incubated overnight at 4°C with rabbit antibodies recognizing SKs (Antibody against KCa2.1 for SK1, Antibody against KCa2.2 for SK2, and Antibody against KCa2.3 (C-term) for SK3,1:300, Alomone Labs), antibody against PKA regulatory β2 subunit (1:5000, Abcam ab75993), antibody against BK slo-1 (α1 subunit of BK) (1:1000, Millipore MADN70) and GAPDH (1:1000, Cell signaling 2118) or antibody against actin (1:10,000, Abcam, ab6276).

Techniques: Patch Clamp, Transmission Assay, Derivative Assay, Western Blot, Quantitation Assay

Journal: Experimental neurology

Article Title: Targeting histone deacetylation for recovery of maternal deprivation-induced changes in BDNF and AKAP150 expression in the VTA

doi: 10.1016/j.expneurol.2018.08.002

Figure Lengend Snippet: MD induces GABAergic metaplasticity in VTA DA neurons through disruption of AKAP150 signaling that renders GABAergic synapses more susceptible to LTD. Our recent findings show that MD increases HDAC2 activity in VTA DA neurons and reduces Ac-H3K9 in the VTA. MD also increases synaptic levels of AKAP150 protein in the VTA with accompanying decreases in synaptic levels of PKA and the levels of mBDNF protein in the VTA. In vivo systemic injection with a selective class I HDAC inhibitor is sufficient to reverse MD-induced histone hypoacetylation and to normalize the levels of mBDNF and AKAP150 proteins in the VTA at 24h after the injection. Our model suggests that HDAC2-mediated targeting of mBDNF could decrease mBDNF signaling through its receptor tropomyosin receptor kinase B (TrkB). We hypothesize that the reduced local BDNF signaling in turn decreases AKAP150-PKA association and AKAP-dependent PKA anchoring at GABAergic synapses while enhancing AKAP150-CaN association that favors the induction of LTD in VTA DA neurons. These epigenetic and synaptic modifications induced by MD could then affect DA neuronal excitability and DA release in VTA projection areas.→ means excitation and ┤ means inhibition.

Article Snippet: Separated proteins were transferred onto nitrocellulose membranes, blocked with casein-based blocking reagent (I-Block, Life Technologies) for 60 minutes at room temperature and then incubated overnight at 4°C with antibodies recognizing HH3 (1:10,000, Abcam ab1791), antibody against PKA regulatory β2 subunit (1:5000, Abcam, ab75993), antibody against PSD95 (1:500, Cell signaling 362333), antibody against ac-H3K9 (1:1,000 cell signaling 3649), antibody against calcineurin subunit A (1:2,000, Abcam ab3673), antibody against AKAP150 (1:500, Santa Cruz Sc-6445), antibody against mature BDNF(mBDNF, 1:1000 ab108319), antibody against vinculin (1:1,000, Abcam ab129002) and antibody against β-actin (1:10,000, Abcam, ab6276).

Techniques: Disruption, Activity Assay, In Vivo, Injection, Inhibition

Journal: Experimental neurology

Article Title: Targeting histone deacetylation for recovery of maternal deprivation-induced changes in BDNF and AKAP150 expression in the VTA

doi: 10.1016/j.expneurol.2018.08.002

Figure Lengend Snippet: (A) Representative Western blots and quantitative data of AKAP150, PSD95 (postsynaptic marker) and β actin (control) in synaptosomal membrane fractions (LP1) of VTA homogenates from non-MD and MD rats. (B) Representative Western blots and quantitative data of synaptic (LP1) and total levels of PKA-RIIβ in LP1 fractions or total homogenates of VTA from non-MD and MD rats. (C) Representative Western blots and quantitative data of synaptic (LP1) and total levels of CaN A subunit in LP1 fractions or total homogenates of VTA from non-MD and MD rats.

Article Snippet: Separated proteins were transferred onto nitrocellulose membranes, blocked with casein-based blocking reagent (I-Block, Life Technologies) for 60 minutes at room temperature and then incubated overnight at 4°C with antibodies recognizing HH3 (1:10,000, Abcam ab1791), antibody against PKA regulatory β2 subunit (1:5000, Abcam, ab75993), antibody against PSD95 (1:500, Cell signaling 362333), antibody against ac-H3K9 (1:1,000 cell signaling 3649), antibody against calcineurin subunit A (1:2,000, Abcam ab3673), antibody against AKAP150 (1:500, Santa Cruz Sc-6445), antibody against mature BDNF(mBDNF, 1:1000 ab108319), antibody against vinculin (1:1,000, Abcam ab129002) and antibody against β-actin (1:10,000, Abcam, ab6276).

Techniques: Western Blot, Marker, Control, Membrane

Journal: Experimental neurology

Article Title: Targeting histone deacetylation for recovery of maternal deprivation-induced changes in BDNF and AKAP150 expression in the VTA

doi: 10.1016/j.expneurol.2018.08.002

Figure Lengend Snippet: Top: Examples of brain sections stained with antibodies to TH (red), and AKAP150 (green) with the merged panels, which show the expression of AKAP150 in TH+ neurons in the VTA of non-MD and MD rats injected with either vehicle or CI-994 (i.p. injection of 10mg/kg) at 24h post-injection. Scale bar, 20μm. Bottom: Graph shows the averaged levels of AKAP150 expression (pooled and averaged data at three AP levels for each group) from non-MD and MD rats in each group.

Article Snippet: Separated proteins were transferred onto nitrocellulose membranes, blocked with casein-based blocking reagent (I-Block, Life Technologies) for 60 minutes at room temperature and then incubated overnight at 4°C with antibodies recognizing HH3 (1:10,000, Abcam ab1791), antibody against PKA regulatory β2 subunit (1:5000, Abcam, ab75993), antibody against PSD95 (1:500, Cell signaling 362333), antibody against ac-H3K9 (1:1,000 cell signaling 3649), antibody against calcineurin subunit A (1:2,000, Abcam ab3673), antibody against AKAP150 (1:500, Santa Cruz Sc-6445), antibody against mature BDNF(mBDNF, 1:1000 ab108319), antibody against vinculin (1:1,000, Abcam ab129002) and antibody against β-actin (1:10,000, Abcam, ab6276).

Techniques: Staining, Expressing, Injection