Journal: Cell reports

Article Title: Cortical regulation of neurogenesis and cell proliferation in the ventral subventricular zone

doi: 10.1016/j.celrep.2023.112783

Figure Lengend Snippet: (A) Schematic: EnvA rVSV-eGFP virus injection into LV of P30 Chat-Cre; R26F-RTT mice. (B) Immunofluorescence: GFP (green) in anterior cingulate cortex of injected side of P33 Chat-Cre; R26F-RTT mice. Scale bar, 100 μm (left) and 30 μm (right). (C) Schematic: AAV-CaMKII-hChR2(E123A)-mCherry virus injection into ACC region of P30 C57BL/6J mice. (D) Immunofluorescence: ChAT (green) and infected projections (red) of the ipsilateral ACC neurons in striatum and SVZ (left), and SVZ (right) of mice in (C). Scale bar, 100 μm (left) and 30 μm (right). (E) Experimental design: pAAV-Ef1a-DIO hChR2(E123T/T159C)-EYFP virus injection into ACC region of P30 Chat-Cre; Ai9 mice. (F) Whole-cell recording: evoked APs from subep-ChAT + neurons upon photo-stimulation for 5 s (left). n = 5 (biological replicates), one-sample t test (right). (G) Whole-cell recordings: evoked EPSCs in response to light stimulation in subep-ChAT + neurons (black) and after blocking with CNQX and AP-5 (red) (left). Mean current amplitude and latency (right). n = 5 (biological replicates), one-sample t test. (H) Experimental design: pAAV-Ef1a-DIO hChR2(E123T/T159C)-mCherry virus injection into ACC region of P30 VGlut1-Cre; ChAT-eGFP mice. (I) Whole-cell recordings: evoked EPSCs in response to light stimulation in subep-ChAT + neurons (black) and after blocking with CNQX and AP-5 (red) (left). Mean current amplitude and latency (right). n = 4 (biological replicates), one-sample t test. (J) Schematic: pAAVrg-hSyn-DIO-mCherry virus injection into LV of P28 Cr-Cre mice. (K) Immunofluorescence: RFP (red) in ACC region of injected side in P50 Cr-Cre mice in (J). Scale bar, 100 μm (left) and 30 μm (right). (L) Electrophysiological recording: CR + excitatory inputs to subep-ChAT + neurons from SVZ wholemount of P30-50 Cr-Cre; ChAT-eGFP; Ai27 mice. Scale bar, 10 μm. (M) Whole-cell recordings: evoked EPSCs in response to light stimulation in subep-ChAT+ neurons (black) and after blocking (red) (left). Mean EPSC amplitude and latency (right). n = 5 (biological replicates), one-sample t test. (N) Experimental design: AAV-EF1a-DIO-hChR2(E123T/T159C)-mCherry virus injection into ACC of P28 Cr-Cre; ChAT-eGFP mice. (O) Whole-cell recordings: EPSCs specifically evoked by ACC CR + input were recorded from subep-ChAT + neurons upon light stimulation (black) and after blocking (red) (left). Mean current amplitude and latency (right). n = 4 (biological replicates), one-sample t test. (P) Schematic summary: ACC-subep-ChAT + circuit regulation of LV NSCs. All blue bars represent the duration of light stimulation. All data presented as mean ± SEM. See also .

Article Snippet: pAAV-Ef1a-DIO hChR2(E123T/T159C)-mCherry , Karl Deisseroth , addgene #35510.

Techniques: Injection, Immunofluorescence, Infection, Blocking Assay

Journal: Cell reports

Article Title: Cortical regulation of neurogenesis and cell proliferation in the ventral subventricular zone

doi: 10.1016/j.celrep.2023.112783

Figure Lengend Snippet: (A) Schematic representation of in vivo optogenetic stimulation experiment in (B) and (J). Immunofluorescence staining for RFP (red) in the ipsilateral (injected) ACC of Cr-Cre mice. Scale bar, 100 μm (right). (B) Experimental design of in vivo optogenetic stimulation for 3 days post AAV-EF1a-DIO-hChR2(E123T/T159C)-mCherry virus injection into ipsilateral ACC and optical fibers implantation into ACC regions of P28 Cr-Cre mice. (C) DCX (green) immunofluorescence staining of ipsilateral (activation) vs. contralateral (control) of SVZ wholemounts from stimulated mice in (B). The yellow dotted circle represents an area where subep-ChAT + neurons are found in the ventral SVZ. Scale bar, 200 μm. (D) Analysis of DCX intensity in ipsilateral (activation) vs. contralateral (control) SVZ wholemounts. p = 0.76 (non-significant), t 3 = 0.32, n = 4 (biological replicates), paired t test. Each dot represents a normalized total DCX protein per SVZ wholemount. (E) DCX (gray) and ChAT (green) immunofluorescence staining of (E′) contralateral (control) vs. (E″) ipsilateral (activation) sides of SVZ from the coronal sections from mice in (B). Red arrows represent DCX + neuroblasts around subep-ChAT + neurons. Scale bar, 10 μm (left and right) and 100 μm (middle). (F) P-S6 intensity analysis of subep-ChAT + neurons in ipsilateral side (activation) vs. contralateral side (control). p = 0.0097, t 4 = 4.65, n = 5 (biological replicates), paired t test. Each dot represents subep-ChAT + neurons per mouse. (G) DCX (gray) immunofluorescence staining of contralateral SVZ (control) vs. ipsilateral SVZ (activation) from coronal section of mice in (E). Blue arrows show DCX + neuroblasts in dorsal domains. Red arrow shows DCX + neuroblasts adjacent to subep-ChAT + neurons in the ventral SVZ. (H) Analysis of DCX + neuroblasts where subep-ChAT + neurons are found on the ipsilateral side (activation) vs. contralateral side (control). ****p < 0.0001, t 4 = 16.3, n = 5 (biological replicates), paired t test. Each dot represents a total of DCX + cells in stimulated coronal sections per mouse. (I) DCX (gray) immunofluorescence staining of contralateral SVZ (control) vs. ipsilateral SVZ (activation) from coronal section of mice in (E) and (G). Blue arrows show DCX + neuroblasts in dorsal domains of SVZ. Scale bar, 20 μm. (J) Experimental design of in vivo optogenetic stimulation for 1 day post AAV-EF1a-DIO-hChR2(E123T/T159C)-mCherry virus injection into ipsilateral ACC and optical fibers implantation into ACC regions of P28 Cr-Cre mice. (K) Schematic representation of the organization of the LV-SVZ. (L) EdU (purple), ChAT (green), and GFAP (gray) immunofluorescence staining of ipsilateral (activation) vs. contralateral (control) SVZ wholemounts from stimulated mice in (J). Yellow arrows show EdU + /GFAP + cells surrounding subep-ChAT + neurons. Scale bar, 10 μm. (M) Analysis of EdU + /GFAP + cells surrounding subep-ChAT + neurons in ipsilateral (activation) V-SVZ vs. contralateral (control) of SVZ wholemounts. p = 0.0015, t 19 = 3.7, n = 20 (biological replicates), paired t test. Data collected from five stimulated Cr-Cre mice. Each dot represents total EdU + /GFAP + cells surrounding a subep-ChAT + neuron. (N) EdU (purple), ChAT (green), and Ki67 (gray) immunofluorescence staining of ipsilateral (activation) vs. contralateral (control) SVZ wholemounts from stimulated mice in (A). Yellow arrows show EdU + /Ki67 + cells surrounding subep-ChAT + neurons. Scale bar, 10 μm. (O) Analysis of EdU + /Ki67 + cells surrounding subep-ChAT + neurons in ipsilateral (activation) V-SVZ vs. contralateral (control) of SVZ wholemounts. p = 0.0002, t 19 = 4.5, n = 20 (biological replicates), paired t test. Data collected from five stimulated Cr-Cre mice. Each dot represents total EdU + /Ki67 + cells surrounding a subep-ChAT + neuron. All data presented as mean ± SEM. See also and .

Article Snippet: pAAV-Ef1a-DIO hChR2(E123T/T159C)-mCherry , Karl Deisseroth , addgene #35510.

Techniques: In Vivo, Immunofluorescence, Staining, Injection, Activation Assay

Journal: Cell reports

Article Title: Cortical regulation of neurogenesis and cell proliferation in the ventral subventricular zone

doi: 10.1016/j.celrep.2023.112783

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: pAAV-Ef1a-DIO hChR2(E123T/T159C)-mCherry , Karl Deisseroth , addgene #35510.

Techniques: Recombinant, Staining, In Vivo, Software