recombinant aav vectors  (Obio Technology Corp Ltd)

Journal: bioRxiv

Article Title: An ASXL3–thyroid hormone axis in parvalbumin interneurons controls autism-like behaviors

doi: 10.64898/2026.05.22.727319

Figure Lengend Snippet: (A) Illustration of the molecular strategy for the expression of the intact ASXL3 protein facilitated by the Cfa GEP split intein. (B) Diagrammatic depiction of the retro-orbital injection technique applied to neonatal mice at postnatal day 3. (C) and (D) Immunodetection and quantitative analysis of total ASXL3 protein expression levels. AAV-GFP, injection of AAV-PHP.eB encoding GFP. AAV-Asxl3, injection of AAV-PHP.eB encoding Flag-ASXL3 N -Intein N and Intein C -ASXL3 C -HA. (E) - (H) Representative images and quantification of ASXL3 and PV expression in the RSC following AAV delivery. (I) - (K) Examination and quantification of ASXL3 and PV expression in the hippocampus after AAV administration. (L) and (M) Immunofluorescence staining of neurons in the second and third cortical layers, accompanied by quantitative analysis of neuronal counts. (N) and (O) Immunofluorescence staining of the fifth layer of cortical neurons and its number statistics. Data are presented as mean ± SD, n = 9 slices from 3 mice for each group. Statistical analysis was performed using one-way ANOVA, with significance denoted by * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The AAV9-hSyn-Flag-Asxl3 N -Intein N (1×1014 vg/mL) and AAV9-hSyn-Intein C -Asxl3 C -3HA (1×1014 vg/mL), also sourced from PackGene Biotech, were administered to cynomolgus monkeys via intrathecal injection.

Techniques: Expressing, Injection, Immunodetection, Immunofluorescence, Staining

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Subtle changes in intrinsic neuronal excitability in Cln3 Δex7/8 mice. ( a, b ) Representative voltage responses from DG-GC to 300 ms current pulses: − 50 pA to 50 pA current steps in 10 pA increments and voltage-current relations (4-months: left, n = 36, 43 cells; N = 5, 5 mice [WT, Cln3 Δex7/8 ]; 3-weeks: right, n = 38, 33 cells; N = 5, 4 mice [WT, Cln3 Δex7/8 ]). ( c, d ) firing pattern of DG-GCs in response to 1 second sub-threshold and hyperpolarizing current pulses: number of action potentials (AP) plotted against increasing current steps (4-months: left, n = 26, 30 cells; N = 4, 5 mice [WT, Cln3 Δex7/8 ]; 3-weeks: right, n = 27, 24 cells; N = 5, 4 mice [WT, Cln3 Δex7/8 ]). Data are mean ± SEM, ** p < 0.01, * p < 0.05; multiple Mann Whitney test (multiple comparisons FDR: two-stage set-up (Benjamini, Krieger and Yakutieli))

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: MANN-WHITNEY

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Impaired synaptic transmission in Cln3 Δex7/8 mice. ( a, b ) Representative mEPSC traces from DG-GCs with summary bar plots of frequency and amplitude (4-months: left, n = 17, 26 cells; N = 3, 3 mice [WT, Cln3 Δex7/8 ]; 3-weeks: right, n = 28, 31 cells; N = 3, 3 mice [WT, Cln3 Δex7/8 ]). ( c, d ) Representative mIPSC traces from DG-GCs with summary bar plots of frequency and amplitude (4-months: left, n = 31,40 cells; N = 5, 5 mice [WT, Cln3 Δex7/8 ]; 3-weeks: right, n = 26, 30 cells; N = 4, 5 mice [WT, Cln3 Δex7/8 ]). Data are median [IQR]; ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Transmission Assay

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Loss of Cln3 impairs synaptic strength in Cln3 Δex7/8 mice. ( a ) Schematic of perforant pathway stimulation (rec., recording pipette; Stim., stimulation electrode). ( b ) Example traces of evoked AMPA- and NMDA- receptor-mediated currents at − 70 mV and +40 mV, respectively; stimulus artifacts removed (replaced by a black line) and summary bar plot of AMPA/NMDA ratio (4-months: n = 33, 42 cells; N = 5, 6 mice [WT, Cln3 Δex7/8 ]). ( c ) Representative traces of evoked NMDA receptor-mediated currents at + 40 mV and peak amplitudes of NMDAR-mediated currents plotted against increasing stimulus intensities (μA). ( d ) Summary bar plot of decay time constant (τ) for NMDA receptor-mediated currents ( n = 35, 40 cells; N = 4, 4 mice [WT, Cln3 Δex7/8 ]). Data are mean [95% CI]; ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Transferring

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Hippocampal neurons of Cln3 Δex7/8 mice have reduced spine number and dendritic complexity. ( a ) Confocal images of dendritic spines in the medial molecular layer of DG (inset scale bar, 1 μm). ( b ) Representative dendritic arbors of DG granule cells from WT and Cln3 Δex7/8 . Left: confocal images with dendrites visualized in green. Right: corresponding reconstructed tracings used for Sholl analysis. Scale bar: 50 μm. ( c ) Summary bar plots of total spine density and ( d ) spine subtype counts per 10 μm of dendritic length (4-months: n = 43, 39 cells; N = 8, 5 mice [WT, Cln3 Δex7/8 ]). ( e ) Sholl analysis quantifying dendritic intersections and ( f ) summary bar plot of total dendritic length ( n = 17, 26 cells; N = 3, 5 mice [WT, Cln3 Δex7/8 ]). Data are mean [95% CI] in ( c ) and median [IQR] in ( d, f ); ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques:

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Synapses in Cln3 Δex7/8 mice have reduced vesicle release probability. ( a ) Representative trace of evoked EPSCs in response to 30 Hz perforant pathway stimulation (black vertical lines above the traces indicate stimulus times). ( b ) Cumulative EPSC plot with linear fit (black line) of the last 6 points (steady state). ( c ) Summary bar plots of release probability (P r ), readily releasable pool (RRP) size, and ( d ) slope (recovery rate) (4-months: n = 31, 25 cells; N = 4, 3 mice [WT, Cln3 Δex7/8 ]). Data are mean [95% CI] for P r and median [IQR] for RRP size and slope; ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques:

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Hippocampal long-term potentiation (LTP) is intact in Cln3 Δex7/8 mice. ( a ) Schematic of LTP induction in the DG via stimulation of the perforant pathway (MPP, medial perforant pathway, induced; LPP, lateral perforant pathway, control). ( b ) Example traces of evoked synaptic currents at baseline (grey) and after LTP induction for WT and Cln3 (colored). ( c ) Time course of normalized EPSC peaks showing LTP induced at the MPP-GC synapse at time 0 (black arrow), while LPP synapses show heterosynaptic depression. Data are mean ± SEM. ( d ) Summary bar plots of induced and control EPSC peaks normalized to baseline for the first and last 10 minutes (4-months: n = 13, 14 cells; N = 6, 5 mice [WT, Cln3 Δex7/8 ]). Data are mean [95% CI]; ns, not significant; linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Control

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Re-expression of CLN3 in the presynaptic neurons of Cln3 Δex7/8 mice improves defective synaptic release. ( a ) Schematic of AAV injections into the entorhinal cortex (EC) to re-introduce CLN3 in presynaptic neurons, combined with optogenetics for selective activation of CLN3 -expressing perforant pathway axons. ( b ) Representative fluorescence images showing CLN3-EGFP and ChR2-mCherry expression in the EC and perforant pathway; merged image shows their colocalization in the molecular layer (ML) of DG (inset, scale bar: 100 µm). Sub, subiculum; GCL, granule cell layer. ( c ) Representative trace of evoked EPSCs in response to 30 Hz optical stimulation of the perforant pathway (blue vertical lines above the traces mark stimulus times). The eEPSC traces in the dotted box shows a zoomed-in portion of the high-frequency optical stimulation period. ( d ) Cumulative EPSC plot with linear fit (black line) of the last 10 points (steady state). ( e ) Summary bar plots of release probability (P r ), readily releasable pool (RRP) size, and slope (4-months: n = 22, 20, 26 cells; N = 3, 3, 4 mice [WT, Cln3 Δex7/8 , Cln3 Δex7/8 + AAV9. CLN3 ]). Data are median [IQR]; ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Expressing, Introduce, Optogenetics, Activation Assay, Fluorescence

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Postsynaptic re-expression of CLN3 normalizes synaptic strength in Cln3 Δex7/8 mice. ( a ) Schematic of AAV injections into the dentate gyrus to re-introduce CLN3 in postsynaptic DG granule cells. ( b ) DIC-IR image showing CLN3-EGFP expression and positions of stimulation (Stim. E) and recording (rec. GFP+) electrodes during electrophysiological recordings. ( c ) Post hoc verification of patched cells by immunostaining after biocytin filling via patch pipette; merged image shows colocalization of CLN3-EGFP and biocytin, confirming successful targeting. ( d ) Example traces of evoked AMPA and NMDA receptor-mediated currents at holding potentials of − 70 mV and +40 mV, respectively. Stimulus artifacts are removed for clarity (replaced with black vertical line). ( e ) Summary bar plots of AMPA/NMDA ratio (4-months: n = 33, 45 cells; N = 5, 5 mice [ Cln3 Δex7/8 , Cln3 Δex7/8 + AAV9. CLN3 ]) and ( f ) mEPSC frequency and amplitude (4-months: n = 25, 22 cells; N = 4, 3 mice [ Cln3 Δex7/8 , Cln3 Δex7/8 + AAV9. CLN3 ]). Horizontal dashed line in blue above the data bars indicate WT values. Data are median [IQR] for AN ratio and mEPSC frequency and mean [95% CI] for mEPSC amplitude; ns, not significant; p -values from linear mixed-effects model

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Expressing, Introduce, Immunostaining, Transferring

Journal: Journal of Translational Medicine

Article Title: Reversible synaptic deficits in early-stage batten disease

doi: 10.1186/s12967-026-08304-w

Figure Lengend Snippet: Timelines of disease progression in humans and mice. The schematic shows disease progression in JNCL patients (top, red) and Cln3 -deficient mice (bottom, cyan). In mice, early functional deficits emerge during the presymptomatic and early symptomatic stages, before extensive neurodegeneration at late stages (14–24 months). AAV-mediated CLN3 rescue at 4 months (green), after onset of synaptic dysfunction, reverses established deficits, demonstrating functional reversibility. Human disease milestones are aligned above to illustrate approximate clinical correspondence, with the 4-month mouse timepoint mapping to the post-diagnosis, pre-motor-decline period in children

Article Snippet: For presynaptic rescue experiments, a 400 nL cocktail of AAV9-hSyn-h CLN3 -IRES-EGFP (Packgene) and AAV9-hSyn-hChR2(H134R)-mCherry (Addgene # 26,976-AAV9; deposited by Karl Deisseroth), containing 2.5 × 10 9 genome copies of each virus, was bilaterally injected into the entorhinal cortex (AP = − 5.2 mm; ML = ± 3.4 mm; DV = − 3.5 mm).

Techniques: Biomarker Discovery, Functional Assay