anti coup tfii  (R&D Systems)

Journal: bioRxiv

Article Title: Cerebral Organoids Uncover Mechanisms of Neural Activity Changes in Epileptogenesis

doi: 10.1101/2025.08.26.672285

Figure Lengend Snippet: (A and B), Immunostainings on isogenic control organoids (A) and TSC patient organoids (B) from patient 1 at day 126 show excitatory neurons marked by TBR1, CAMKII and SATB2. Organoids are positive for COUP-TFII and Calretinin, and negative for NKX2.1, SST and Parvalbumin, marking GABAergic inhibitory interneurons from the CGE and not the medial ganglionic eminence (MGE). DAPI+ nuclei are shown in blue. Scale bars: 50 µm. (C to H) Pharmacological perturbations of network activity affect event occurrences and MUA firing rates in control and TSC organoids. Representative raster plots showing that blocking transmission through gap junctions with CBX (200 µM) did not affect synchronous network activity (C) and MUA firing rates (G). In contrast, CNQX (10 µM) application blocking AMPA receptors (D) and 0.5 µM TTX (E) significantly reduces synchronous network event activity and firing rates (G). Application of 20 µM of GABA to TSC and control brain organoids abolishes synchronous network activity (F) and firing activity (H). Partial recovery during washout validates the role of GABAergic transmission. Network events are highlighted in orange. (I) Top: Representative LFP signals from a single electrode channel during network events in control (n = 31 network events) and TSC (n = 24 network events) organoids between 6-7 months, presented mean ± SEM. Bottom: Corresponding time-frequency spectrograms of mean LFP signals highlight dominant power in the delta and theta range (white arrow), especially related to trailing oscillatory subpeaks. (J) (Top and bottom left) Immunohistochemistry overview of SCGN from two samples from the left front seizure focus of a 20-month TSC patient. (Top right panels) 20x magnification of the first sample shows SCGN+ interneurons within the front seizure focus that co-localize with Sp8 and COUP-TFII, confirming CGE identity. (Bottom right panels) 63x magnification of the second sample highlights one SCGN+ interneuron within the frontal seizure focus that exhibits pathological dendritic beading. This interneuron also co-localizes with Sp8 and COUP-TFII, confirming CGE identity. Scale bar: A: 50 µm; B: 50µm (K) In organoids, immunohistochemistry of SCGN in TSC organoids show beading morphology in the SCGN+ interneurons and co-localization with MAP2, marking dendrites. Yellow arrowheads mark dendritic beads that SCGN+/MAP2+.

Article Snippet: The following antibodies were used on human brain samples: COUP-TFII (PP-H7147-00; Ms; Perseus Proteomics), Sp8 (sc-104661; Gt; Santa Cruz Biotechnology), SCGN (HPA006641; Rb; Sigma-Aldrich).

Techniques: Control, Activity Assay, Blocking Assay, Transmission Assay, Immunohistochemistry