recombinant probdnf  (Alomone Labs)

Journal: Science Advances

Article Title: BDNF-driven synaptic plasticity requires autocrine matrix metalloproteinase–9 activity

doi: 10.1126/sciadv.adx2369

Figure Lengend Snippet: ( A ) Scheme of the TrkB FRET sensor. Created in BioRender. Kalita, K. (2025) https://BioRender.com/qaducze . ( B ) Example FLIM images showing TrkB activation. Warmer colors represent higher TrkB activity. Yellow cross indicates uncaging spot. Scale bar, 1 μm. ( C ) Averaged TrkB activation changes (Δ binding fraction) in dendritic spines following uncaging in the presence of DMSO or Inhibitor I. Data are means ± SEM. Gray box indicates uncaging period. ( D ) Statistical analysis of (C). Averaged TrkB activation in stimulated spines during transient (1 to 3 min) and sustained phase (9 to 11 min). Gray dots, individual spines; bars, means ± SEM. DMSO (blue; n = 70 spines, 27 cells, 16 animals) and Inhibitor I (red; n = 49 spines, 21 cells, 10 animals). Repeated-measures ANOVA: Time ( P = 0.0010); Inhibitor ( P = 0.0013); Time × Inhibitor ( P = 0.7403), followed by Šídák’s multiple comparisons test ( P values indicated on the graph). ( E ) Averaged TrkB activation changes in dendritic spines following uncaging in WT or MMP-9 KO slices. All markings as in (C). ( F ) Statistical analysis of (E). All markings as in (D). WT (blue, n = 66 spines; 22 cells, 10 animals), MMP-9 KO (yellow, n = 73 spines; 25 cells, 11 animals). Repeated-measures ANOVA: Time ( P = 0.0257); MMP-9 KO ( P = 0.0054); Time × MMP-9 KO ( P = 0.7238), followed by Šidák’s multiple comparison test ( P values indicated on the graph). ( G ) Example immunoblot of digestion reaction of proBDNF incubated with either active MMP-9, inactive MMP-9 (E402A), or the reaction buffer. Bands correspond to proBDNF (~26 kDa) and mBDNF (~14 kDa). ( H ) Quantification of immunoblots of three digestion reactions. Gray dots, individual values of mBDNF band intensity in separate experiments; bars, mean ± SEM. One-way ANOVA ( P = 0.0021) followed by Tukey’s multiple comparisons test ( P values indicated on the graph).

Article Snippet: Twenty nanograms of recombinant proBDNF (Alomone Labs) was incubated with 50 ng of recombinant MMP-9 (Calbiochem) or 50 ng of recombinant, human, inactive MMP-9 (E402A) in total volume of 20 μl.

Techniques: Activation Assay, Activity Assay, Binding Assay, Comparison, Western Blot, Incubation

Journal: Science Advances

Article Title: BDNF-driven synaptic plasticity requires autocrine matrix metalloproteinase–9 activity

doi: 10.1126/sciadv.adx2369

Figure Lengend Snippet: Activation of NMDAR (1) leads to the release of MMP-9 and BDNF (2), which might be released in its either pro-form or mature form with a propeptide. (3) tPA activates plasminogen to plasmin, which can also activate proMMP-9. (4) Plasmin and MMP-9 can extracellularly process proBDNF to mBDNF, which activates its receptor—TrkB (5). TrkB activation, together with other intracellular signaling, leads to the LTP cascade, including actin polymerization and cytoskeleton remodeling causing spine enlargement. (6) ProBDNF and BDNF propeptide, which is also co-released with mBDNF, can activate p75 TNR , leading to LTD. (7) It is possible that MMP-9 can reduce bioactive BDNF propeptide and promote a competing TrkB activation. (8) Both plasmin and MMP-9 are blocked by their inhibitors, which control their action. Created in BioRender. Kalita, K. (2025) https://BioRender.com/nbak5x7 .

Article Snippet: Twenty nanograms of recombinant proBDNF (Alomone Labs) was incubated with 50 ng of recombinant MMP-9 (Calbiochem) or 50 ng of recombinant, human, inactive MMP-9 (E402A) in total volume of 20 μl.

Techniques: Activation Assay, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Maternal immune activation-induced proBDNF-mediated neural information processing dysfunction at hippocampal CA3-CA1 synapses associated with memory deficits in offspring

doi: 10.3389/fcell.2022.1018586

Figure Lengend Snippet: The performance in the conditioning and fear memory test. PolyI:C-treated offspring were intra -CA1 or -CA3 infused with anti-proBDNF antibody or p75 NTR inhibitor TAT-Pep5 30 min before the testing. The freezing levels of polyI:C-treated offspring during the training (A) and the memory test (B) are lower while blocking the activation of proBDNF/p75 NTR signaling can effectively reversed the impaired memory behavior. Data are presented as mean ± SEM. * p < 0.05, Control + ACSFCA1 vs. other groups in A and the group vs. Control + ACSFCA1, PolyI:C + AntiCA1 and PolyI:C + Pep5CA1 in B. Training phase: Control + ACSFCA1: n = 6; PolyI:C + ACSFCA1: n = 6; PolyI:C + AntiCA1: n = 6; PolyI:C + AntiCA3: n = 6; PolyI:C + Pep5CA1: n = 6. Memory test: Control + ACSFCA1: n = 6; PolyI:C + ACSFCA1: n = 4; PolyI:C + AntiCA1: n = 7; PolyI:C + AntiCA3: n = 7; PolyI:C + Pep5CA1: n = 7.

Article Snippet: The cleavage-resistant proBDNF (2 ng/ml; Cat#B257 Alomone Labs), TAT-Pep5 (4 ng/μl; Cat#506181, EMD Millipore), or artificial CSF (ACSF) was infused bilaterally at a rate of 0.5 μl/min/side for 2 min.

Techniques: Blocking Assay, Activation Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Maternal immune activation-induced proBDNF-mediated neural information processing dysfunction at hippocampal CA3-CA1 synapses associated with memory deficits in offspring

doi: 10.3389/fcell.2022.1018586

Figure Lengend Snippet: (A) The changes in EPSC of prymidal CA1 neurons. Typical consecutive sample traces of sEPSCs from each group (Top). The frequency of sEPSC (B) is not altered but the amplitude of sEPSC (C) is decreased in polyI:C-treated offspring. Incubation with anti-proBDNF antibody or TAT-Pep5 inhibitor can significantly enhance the declined amplitude. Data are presented as mean ± SEM. * p < 0.05, vs. Control + ACSFCA1, PolyI:C + AntiCA1 and PolyI:C + Pep5CA1. Control + ACSFCA1: n = 5; PolyI:C + ACSFCA1: n = 5; PolyI:C + AntiCA1: n = 7; PolyI:C + AntiCA3: n = 7; PolyI:C + Pep5CA1: n = 6.

Article Snippet: The cleavage-resistant proBDNF (2 ng/ml; Cat#B257 Alomone Labs), TAT-Pep5 (4 ng/μl; Cat#506181, EMD Millipore), or artificial CSF (ACSF) was infused bilaterally at a rate of 0.5 μl/min/side for 2 min.

Techniques: Incubation

Journal: Frontiers in Cellular Neuroscience

Article Title: ProBDNF and Brain-Derived Neurotrophic Factor Prodomain Differently Modulate Acetylcholine Release in Regenerating and Mature Mouse Motor Synapses

doi: 10.3389/fncel.2022.866802

Figure Lengend Snippet: proBDNF in low concentration (1 nM) decreases the rate of spontaneous ACh release at newly formed NMJs but has no effect on evoked ACh release at newly formed NMJs. (A) Representative recordings of MEPPs in control (left) and upon application of proBDNF (right). (B) Mean MEPP amplitude, cumulative probability plots, frequency and time-course parameters (left to right) in control ( n = 27) and upon application of proBDNF ( n = 30). (C) Representative recordings of EPPs during a short (1 s) high-frequency (50 Hz) train in control (above) and upon application of proBDNF (below). (D) Changes in the EPP amplitude (above) and in the quantal content of EPPs (below) in control ( n = 19) and in the presence of proBDNF ( n = 24). Inset shows MEPP amplitudes.

Article Snippet: We used recombinant human proBDNF (its cleavable form) and BDNF prodomain (purchased from Alomone Labs, Jerusalem, Israel); tertiapin-Q as a selective blocker of inward-rectifier K + channels, iberiotoxin as a selective blocker of the big conductance Ca 2+ -activated K + channels, nitrendipine as a L-type calcium channel blocker, Y-27632 dihydrochloride as a selective inhibitor of ROCK, and (±)-Vesamicol hydrochloride as a direct inhibitor of vesicular ACh transport (all purchased from Tocris, Bio-Techne, Minneapolis, MN, United States); TAT-Pep5 as a p75 receptor signaling inhibitor (purchased from Sigma-Aldrich, United States).

Techniques: Concentration Assay, Control

Journal: Frontiers in Cellular Neuroscience

Article Title: ProBDNF and Brain-Derived Neurotrophic Factor Prodomain Differently Modulate Acetylcholine Release in Regenerating and Mature Mouse Motor Synapses

doi: 10.3389/fncel.2022.866802

Figure Lengend Snippet: Inhibition of GIRK channels and TrkB receptors but not Rho-GDI-associated signaling of p75 receptors, abolish proBDNF-induced decrease in MEPP frequency newly formed NMJs. (A) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 21) and upon application of GIRK blocker tertiapin-Q (100 nM, n = 23). (B) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 17) and upon application of proBDNF (1 nM) in the presence of tertiapin-Q (100 nM, n = 26). (C) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 23) and upon application of proBDNF (1 nM) in the presence of Rho-GDI-associated p75 signaling inhibitor TAT-Pep5 (1 μM, n = 31). (D) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 15) and upon application of TrkB antagonist cyclotraxin B (100 nM, n = 19). (E) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 19) and upon application of proBDNF (1 nM) in the presence of cyclotraxin B (100 nM, n = 28).

Article Snippet: We used recombinant human proBDNF (its cleavable form) and BDNF prodomain (purchased from Alomone Labs, Jerusalem, Israel); tertiapin-Q as a selective blocker of inward-rectifier K + channels, iberiotoxin as a selective blocker of the big conductance Ca 2+ -activated K + channels, nitrendipine as a L-type calcium channel blocker, Y-27632 dihydrochloride as a selective inhibitor of ROCK, and (±)-Vesamicol hydrochloride as a direct inhibitor of vesicular ACh transport (all purchased from Tocris, Bio-Techne, Minneapolis, MN, United States); TAT-Pep5 as a p75 receptor signaling inhibitor (purchased from Sigma-Aldrich, United States).

Techniques: Inhibition, Control

Journal: Frontiers in Cellular Neuroscience

Article Title: ProBDNF and Brain-Derived Neurotrophic Factor Prodomain Differently Modulate Acetylcholine Release in Regenerating and Mature Mouse Motor Synapses

doi: 10.3389/fncel.2022.866802

Figure Lengend Snippet: BDNF prodomain (1 nM) but not proBDNF (1 nM), induces strong inhibition of spontaneous end evoked ACh release at mature NMJs. (A) Mean MEPP amplitude, cumulative probability plots, frequency, and time-course parameters (left to right) in control ( n = 19) and upon application of proBDNF ( n = 26). (B) Representative recordings of MEPPs (top left) and mean MEPP amplitude, cumulative probability plots, frequency, (top right) and their time-course parameters (bottom) in control ( n = 23) and upon application of BDNF prodomain ( n = 33). (C) Representative recordings of EPPs during a short (1 s) high-frequency (50 Hz) train in control (above) and upon application of BDNF prodomain (below). (D) Changes in the EPP amplitude (above) and in the quantal content of EPPs (below) in control ( n = 22) and in the presence of proBDNF ( n = 21). Inset shows MEPP amplitudes.

Article Snippet: We used recombinant human proBDNF (its cleavable form) and BDNF prodomain (purchased from Alomone Labs, Jerusalem, Israel); tertiapin-Q as a selective blocker of inward-rectifier K + channels, iberiotoxin as a selective blocker of the big conductance Ca 2+ -activated K + channels, nitrendipine as a L-type calcium channel blocker, Y-27632 dihydrochloride as a selective inhibitor of ROCK, and (±)-Vesamicol hydrochloride as a direct inhibitor of vesicular ACh transport (all purchased from Tocris, Bio-Techne, Minneapolis, MN, United States); TAT-Pep5 as a p75 receptor signaling inhibitor (purchased from Sigma-Aldrich, United States).

Techniques: Inhibition, Control