synapsin ii antibody  (ABclonal Biotechnology)

Journal: bioRxiv

Article Title: GABA-induced Ca 2+ signaling in the primary cilium of neurons

doi: 10.1101/2025.05.26.656109

Figure Lengend Snippet: A. Immunofluorescence images from naïve or 5HT6-GGECO1-expressing cultured mouse neurons (from cortex and CA1) showing the distribution of GABA-B1 receptors (magenta) in primary cilia positive for AC3 (green). Intensity profiles from lines drawn along the cilia are shown to the right and further show the confinement of GABA-B1 receptors to the cilia base of cultured neurons. B. Quantification of GABA-B1 receptor enrichment at the cilia base in naïve cultured neurons or cultured cortical and hippocampal neurons expressing 5HT6-GGECO1 (naïve 2,76±0,37; Cx 8,18±0,91; Hipp 4,94±0,44 arbitrary units). GABA-B1 positive segment length was measured for each group and found significantly different (naïve, Cx and Hipp, in µm: 2,78±0,37; 8,18±0,91; 5,03±0,39; mean±SEM; Kruskal-Wallis ANOVA followed by multiple comparisons, naïve n=12, Cx n=22, Hipp n=38, for A & B). C. Simultaneous recording of cytoplasmic (grey) and ciliary (pink) calcium activities from a neuron before and after stimulation with 100nM baclofen. Activation of metabotropic GABA receptors was without effect on the frequency of somatic events while ciliary signaling was increased by the agonist. Positive deviations from baseline were computed over time to produce ciliary activity count traces (black, lower panel). D. Averaged activity counts corresponding to neurons in control (blue) baclofen (100nM, black) and TTX (1 µM) + baclofen (100nM, grey) showing an increase of activity after stimulation with baclofen. E. Quantification of activity changes was performed in a pair wise fashion, comparing the averaged activity during initial vs final period (control) and baseline vs baclofen (either in the absence of presence of TTX). Time lacked an effect on activity (control P=0,4332) while baclofen stimulation led to a significant increase in both - /+ TTX conditions (respectively P=0,0002 and P=0,0039; two tailed Wilcoxon matched-pairs signed rank test) F. To test whether abolition of action potentials had an impact on spontaneous ciliary activity, recordings in control and TTX (initial 20 minutes both groups) were compared. Statistical analysis showed no difference (P=0,1468 Mann-Whitney two tailed t-test). G. Immunofluorescence images showing the distribution of primary cilia (AC3; green) and synapses (Synapsin II; magenta) in cultured cortical neurons. Boxed areas are magnified to the right. Scatter plot to the left shows Synapsin II fluorescence intensity at primary cilia and random locations within the same sample (P=0,3696 Wilcoxon matched pairs signed rank test).

Article Snippet: The following primary antibodies (1/300 dilutions) were used: ARL13b (Abcam ab136648), AC3 (Abcam ab277619), AC3 (Alomone AAR-043), NeuN (Sigma-Aldrich ABN90), GFAP (Synaptic System 173–004), Synapsin II (Alomone ANR-015), GABA-B1 (Alomone AGB-001), MCHR1 (Thermo Fischer PA5-77492), Patched (Abcam ab53715), EP4 (Santa Cruz Biotechnology sc-55596), ANKS6 (Sigma HPA008355), NPHP3 (Proteintech 22026-1-AP), NEK8 (kind gift from Prof. David R. Beier, Seattle Children’s Hospital, USA).

Techniques: Immunofluorescence, Expressing, Cell Culture, Activation Assay, Activity Assay, Control, Two Tailed Test, MANN-WHITNEY, Fluorescence

Journal: Neural Regeneration Research

Article Title: Nerve root magnetic stimulation regulates the synaptic plasticity of injured spinal cord by ascending sensory pathway

doi: 10.4103/NRR.NRR-D-24-00628

Figure Lengend Snippet: NRMS upregulates transcriptional levels of genes related to synaptic plasticity in damaged region of SCI rats. (A) The functional classification of different genes in the SCI + NRMS and SCI + SS groups by GO enrichment analysis. (B) The functional classification of different genes between the SCI + NRMS and SCI + SS groups by KEGG database. (C–F) The relative levels of Syn2 , Syp , GAP43 , and MAP2 mRNA normalized by the Sham group. The bold lines, upper boundaries, and lower boundaries represent the medians, 75 th percentiles, and 25 th percentiles, respectively. Whiskers extend 1.5 times interquartile range ( n = 4 per group). * P < 0.05, vs . Sham group; # P < 0.05, vs . SCI + SS group. GAP43: Growth-associated protein 43; GO: Gene Ontology; KEGG: Kyoto Encyclopedia of Genes and Genomes; MAP2: microtubule-associated protein 2; NRMS: nerve root magnetic stimulation; SCI: spinal cord injury; SS: sham stimulation; Syn2: synapsin II; Syp: synaptophysin.

Article Snippet: The following antibodies were used: postsynaptic density protein-95 (PSD95; rabbit, 1:1000, Cell Signaling Technology, Danvers, MA, USA, Cat# 3450S, RRID: AB_2292883), Synapsin I (rabbit, 1:1000, Abmart, Shanghai, China, Cat# 334286), GAP43 (mouse, 1:400, Santa Cruz, Biotechnology, Dallas, TX, USA, Cat# sc-17790, RRID: AB_627660), Synapsin II (rabbit, 1:1000, Abclonal, Wuhan, China, Cat# A19542), and GAPDH (mouse, 1:20 000, Sigma-Aldrich, Cat# G8795, RRID: AB_1078991).

Techniques: Functional Assay

Journal: Neural Regeneration Research

Article Title: Nerve root magnetic stimulation regulates the synaptic plasticity of injured spinal cord by ascending sensory pathway

doi: 10.4103/NRR.NRR-D-24-00628

Figure Lengend Snippet: NRMS alters expression of proteins associated with synaptic plasticity in the injured spinal cord of rats. (A) Representative western blots show the expression levels of PSD95, GAP43, Synapsin I and Synapsin II collected from the damaged area of spinal cord on day 22 after SCI. (B–E) The semi-quantified results of PSD95, GAP43, Synapsin I, and Synapsin II protein levels. All experiments were repeated three times. Data are expressed as mean ± SD ( n = 5 per group). ** P < 0.01, *** P < 0.001, vs. Sham group; # P < 0.05, ## P < 0.01, vs . SCI + SS group. GAP43: Growth-associated protein 43; PSD95: postsynaptic density protein-95; NRMS: nerve root magnetic stimulation; SCI: spinal cord injury; SS: sham stimulation.

Article Snippet: The following antibodies were used: postsynaptic density protein-95 (PSD95; rabbit, 1:1000, Cell Signaling Technology, Danvers, MA, USA, Cat# 3450S, RRID: AB_2292883), Synapsin I (rabbit, 1:1000, Abmart, Shanghai, China, Cat# 334286), GAP43 (mouse, 1:400, Santa Cruz, Biotechnology, Dallas, TX, USA, Cat# sc-17790, RRID: AB_627660), Synapsin II (rabbit, 1:1000, Abclonal, Wuhan, China, Cat# A19542), and GAPDH (mouse, 1:20 000, Sigma-Aldrich, Cat# G8795, RRID: AB_1078991).

Techniques: Expressing, Western Blot

Journal: Brain, Behavior, & Immunity - Health

Article Title: Synapsin autoantibodies during pregnancy are associated with fetal abnormalities

doi: 10.1016/j.bbih.2023.100678

Figure Lengend Snippet: High prevalence of synapsin-I autoantibodies in pregnant women. (A) Representative examples of immunofluorescence stainings with human sera at 1:300 dilution (green) with (top row) or without binding (bottom row) to HEK293 cells overexpressing human synapsin-Ib. Protein expression is confirmed with a commercial synapsin-I/II antibody (red). Nuclei are stained with DAPI (blue). Scale bar = 20 μm. (B) Frequencies of synapsin-Ib autoantibodies in pregnant women and control cohorts, as determined by CBA. CBA scores: 0 = no binding, 1 = unspecific signal, 2 = intensive binding (positive); vertical dotted line represents cut-off for positivity. (C) Representative immunoblots of wild type (wt) and Syn1/2/3 triple knock out (TKO) mice cortex homogenates with a commercial synapsin-I/II antibody as positive control and sera from CBA-positive pregnant women (1:200 dilution). The strong band at ∼90 kDa corresponding to the molecular weight of synapsin-Ia/Ib was detected in wild type but not in TKO mouse tissue. Another band at ∼50 kDa likely represents the synapsin-IIb isoform or breakdown products of synapsin-I. Detection of GAPDH served as loading control. (D) Immunofluorescence staining on rat hippocampal neurons demonstrated strong IgG binding of human sera (green) in a characteristic synaptic pattern (insert shows high magnification, MAP2 [blue] staining for better visualization of neuronal processes). The punctate staining completely overlapped with a commercial synapsin antibody (red, insert for high magnification).

Article Snippet: For co-stainings, commercial rabbit synapsin-I/II antibody (Synaptic Systems, #106002) and anti-rabbit IgG AF594-antibody (Jackson IR, #111-585-003) were used.

Techniques: Immunofluorescence, Binding Assay, Expressing, Staining, Western Blot, Knock-Out, Positive Control, Molecular Weight

Journal: Brain, Behavior, & Immunity - Health

Article Title: Synapsin autoantibodies during pregnancy are associated with fetal abnormalities

doi: 10.1016/j.bbih.2023.100678

Figure Lengend Snippet: Strong, titer-dependent association of synapsin-I autoantibodies with abnormalities of fetal development during pregnancy. (A-B) Several ultrasound parameters and status of having previous children with neuropsychiatric disorders were significantly associated with the presence of synapsin-I autoantibodies in serum of pregnant mothers (A), with odds ratios of 3–7 (B). Statistical analysis used Fisher's exact test when expected frequencies were <5 or Chi-square test with expected frequencies ≥5, * representing p < 0.05, **p < 0.01 and ***p < 0.001. (C) Synapsin autoantibodies reached the fetal circulation and were present with similar titers (±1 titer step, in one case 2 titer steps) in umbilical cord blood. (D) Synapsin autoantibodies were of IgG1 and/or IgG3 subclass in almost all pregnant women. (E) While the autoantibody titer alone did not discriminate between pathological and normal findings during fetal development (left), the combination of IgG1 subclass with high-level titers (≥1:30,000) was significantly associated with abnormal measurements (right).

Article Snippet: For co-stainings, commercial rabbit synapsin-I/II antibody (Synaptic Systems, #106002) and anti-rabbit IgG AF594-antibody (Jackson IR, #111-585-003) were used.

Techniques:

Journal: Brain, Behavior, & Immunity - Health

Article Title: Synapsin autoantibodies during pregnancy are associated with fetal abnormalities

doi: 10.1016/j.bbih.2023.100678

Figure Lengend Snippet: Detection of synapsin-I autoantibodies by ELISA, immunoblotting and microarray-based epitope mapping. (A) ELISA-based screening of all human sera using recombinant synapsin-I correlated with autoantibody binding in the CBA. (B) Immunoblots of synapsin-Ib-transfected HEK293 cells confirm that some human sera (1:200 dilution) strongly detect the synapsin-Ib band (middle), while in others binding to non-conformational epitopes is lost (right). Commercial synapsin-I/II antibody served as positive control (left, GAPDH is exemplarily shown; UT = untransfected cells). (C) An overlapping peptide library containing 20-mer peptides with an off-set of three amino acids was designed to fully cover synapsin-I primary sequence. Microarrays were probed with autoantibody-positive and -negative serum. The representative example shows detection of IgG epitope 2 within the synapsin-I slides and a representative negative control serum. Heat map visualization: IgG binding intensities are displayed in grayscale (0 = no binding, 1 = maximal binding). The core shared motif is highlighted in red. (D) Synapsin-I domain architecture together with a cartoon representation of the alpha-fold model of synapsin-I visualized in Pymol. Mapped synapsin-I autoantibody epitopes (1–5, marked in red) are located in domain A-B (epitope 1, peptide sequence: 28 PQPPPPPPGAH 38 ), domain C (epitope 2, 124 TDWAKYFKGKK 134 ) and domain D (epitope 3: 472 PGPQRQGPPLQ 482 ; 4: 583 GGQQRQGPPQK 592 ; 5: 610 VPRTGPPTTQQPRP 623 ).

Article Snippet: For co-stainings, commercial rabbit synapsin-I/II antibody (Synaptic Systems, #106002) and anti-rabbit IgG AF594-antibody (Jackson IR, #111-585-003) were used.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Microarray, Recombinant, Binding Assay, Transfection, Positive Control, Sequencing, Negative Control