Journal: eLife

Article Title: ExSTED microscopy reveals contrasting functions of dopamine and somatostatin CSF-c neurons along the lamprey central canal

doi: 10.7554/eLife.73114

Figure Lengend Snippet: ( A–C ) Stimulated emission depletion (STED) images of dopamine-containing dense-core vesicles (DCVs) in the soma of CSF-c neurons in normal (pH 7.4), acidic (pH 6.5), and alkaline (pH 8.5) extracellular solution. Scale bar, 1 µm. ( D ) Quantification of the number of dopamine DCVs number density in cell area (µm −2 ) in the different conditions ( n = 10). Student’s t -test: non-significant (n.s.) between pH 7.4 and 6.5 (p = 0.27, t 9 = 1.12), and 7.4 and 8.5 (p = 0.29, t 9 = 1.08). ( E ) Whole-cell patch recording of a CSF-c neuron, showing firing spontaneous action potentials in control (pH 7.4), acidic (p H 6.5), and alkaline (pH 8.5) conditions in the presence of gabazine (20 mM) and kynurenic acid (2 mM). ( F–H ) Photomicrographs of the CSF-c neurons recorded in ( E ) intracellularly filled with Neurobiotin (arrow) during recording. The labelled cell showed immunoreactivity to tyrosine hydroxylase (TH, arrow). Scale bar, 10 µm. ( I ) Action potential frequency during 1 min in CSF-c neurons at pH 7.4, 6.5, and 6.8, respectively ( n = 15). Student’s paired t -test: non-significant difference (n.s.) between pH 7.4 and 6.5 (p = 0.24, t 14 = −1.22), and 7.4 and 8.5 (p = 0.1, t 14 = −1.75). The bar graph data are represented as the means, with error bars representing standard deviation (SD). cc, central canal. Figure 4—source data 1. Effect of acidic or alkaline pH on dopamine dense-core vesicles (DCVs) number density in cell area of dopaminergic cerebrospinal fluid-contacting (CSF-c) neurons. Quantification of dopamine DCVs number density in cell area (µm −2 ) in dopaminergic CSF-c neurons in the different pH. Figure 4—source data 2. Effect of acidic and alkaline pH on action potential frequency in dopaminergic cerebrospinal fluid-contacting (CSF-c) neurons. Quantification of action potential frequency in dopaminergic CSF-c neurons at different pH conditions during 1 min.

Article Snippet: Neurons were intracellularly labelled by injection of 0.5% Neurobiotin (Vector Laboratories) during whole-cell recordings.

Techniques: Standard Deviation

Journal: eLife

Article Title: ExSTED microscopy reveals contrasting functions of dopamine and somatostatin CSF-c neurons along the lamprey central canal

doi: 10.7554/eLife.73114

Figure Lengend Snippet:

Article Snippet: Neurons were intracellularly labelled by injection of 0.5% Neurobiotin (Vector Laboratories) during whole-cell recordings.

Techniques: Isolation, Recombinant, Plasmid Preparation, Injection, Software, Labeling, In Situ Hybridization

Journal:

Article Title: Cell-type specific expression of ATP-sensitive potassium channels in the rat hippocampus

doi: 10.1111/j.1469-7793.1999.315ae.x

Figure Lengend Snippet: Top row, hippocampal slice stained with DAPI. The white frames enclose the regions in which the somata of CA1 pyramidal cells (A), interneurones from stratum radiatum CA1 (B) and dentate granule cells (C) were located. Second row, morphology of a pyramidal cell (A), an interneurone (B) and a granule cell (C). The cells were filled with neurobiotin, coupled to avidin-fluorescein, and imaged using a two-photon microscope. Axons are marked by white arrows. Note that the interneurone (B) is shown at a higher magnification and that the distal dendrites of this cell were cut during slicing. Third row, spontaneous action potentials recorded in the three cell types. Fourth row, trains of action potentials evoked by 400 ms current pulses of 50 pA, as indicated by the bars.

Article Snippet: To label the cells for morphological studies, 1 % neurobiotin tracer (Vector Labs, Burlingame, CA, USA) was added to the internal solution.

Techniques: Staining, Avidin-Biotin Assay, Microscopy

Journal: bioRxiv

Article Title: Functional development of eye movements and visuomotor circuits in lampreys

doi: 10.1101/2023.09.06.556551

Figure Lengend Snippet: The colored rectangles group results belonging to the developmental stage indicated under the schematic of the experimental preparation (blue: stage 5 metamorphic; green: stage 7 metamorphic/postmetamorphic). (A) Schematic drawing (left) indicating the location of the photomicrograph (right) showing retrogradely labeled neurons in the pretectum (PT) of a stage 5 metamorphic lamprey after a Neurobiotin injection in the middle rhombencephalic reticulospinal nucleus (MRRN). Projection neurons can be observed both in the periventricular region (dashed line oval), and in lateral aspects (arrows). (B) Extracellular responses in the MRRN after stimulation of the PT (red trace) and optic tectum (OT, green trace). The onset of the extracellular activity is indicated by a dashed red line for PT stimulation, and a dashed green line for OT stimulation. (C) Photomicrograph showing that no retrogradely labeled neurons can be seen in the OT (indicated by a dashed line) of a stage 5 metamorphic lamprey after a Neurobiotin injection in the MRRN. (D) Graph showing that the onsets of MRRN responses evoked by PT stimulation (red) were significantly shorter than those evoked by OT stimulation (green; unpaired t-test). (E-F) Graphs showing the mean responses evoked in the MRRN of a stage 5 metamorphic animal evoked by stimulation of the PT (E) and OT (F) in response to 4 pulses (10 Hz). Values are normalized to the first local field potential (LFP). (G) Schematic drawing (left) indicating the location of the photomicrograph (right) showing a few retrogradely labeled neurons from the MRRN (arrows) in the OT of a late metamorphic animal (stage 7/recent postmetamorphic). (H) Extracellular responses in the MRRN of a late metamorphic animal (stage 7/recent postmetamorphic) in response to OT stimulation (4 pulses, 10 Hz). A two-components response can be observed: a fast onset weak response (indicated by a dashed line rectangle) followed by a stronger signal. In the electrophysiological traces, stimulation artifacts were removed for clarity. In all graphs, data are shown as mean ± s.d. Abbreviations: nMLF Nucleus of the Medial Longitudinal Fasciculus, pc Posterior commissure, SNc Substantia Nigra pars compacta, TS Torus Semicircularis, nIII Oculomotor Nucleus, M5 Retinopetal Nucleus of Schöber. Scale bar = 100 µm in A, G and M; 200 µm in C and I.

Article Snippet: To detect the Neurobiotin tracer, sections were incubated in Cy2-conjugated streptavidin (Jackson ImmunoResearch) 1:1000 in blocking solution (1 % bovine serum albumin, 10 % sheep serum, 0.1 % sodium azide and 0.3 % Triton X-100 in PBS).

Techniques: Labeling, Injection, Activity Assay

Journal: Physiological reports

Article Title: Hierarchical organization of long-range circuits in the olfactory cortices.

doi: 10.14814/phy2.12550

Figure Lengend Snippet: Figure 4. Whole-cell recordings of LRMGP-EPSCs in pPC AAV-ChR2-injected mice. (A) Representative experiment with AAV-ChR2 injection in the pPC; a, confocal image of 300-lm thick horizontal slice showing the pPC injection site (white arrow) and the recording sites (red boxes); b, neurobiotin-labeled neurons; c, monosynaptic AMPA (downward) and AMPA + NMDA (upward) current traces recorded from neurons in three individual cortices. Note the differences of scale bars. (B, C) Comparisons of normalized monosynaptic AMPA and NMDA current strength. (D) a, Bright-field image of an exemplar pair of IN and PYR recorded in LEC with neurobiotin labeling; b-d, three summary plots showing the comparisons of AMPA, NMDA, and their ratios, respectively, between INs and nearby PYRs recorded in aPC and LEC for pPC injection experiments. *P < 0.05, **P < 0.01.

Article Snippet: Recorded neurons were typically located 50–100 lm below the surface of the recorded slice and had relatively intact dendritic arbors, verified by post hoc neurobiotin reconstruction (antibodies used from the Santa Cruz Biotechnology, Dallas, TX: goat anti-rabbit HRP, sc-2030; Rb HRP, sc-2749; donkey anti-goat HRP, sc2056).

Techniques: Injection, Labeling

Journal: Physiological reports

Article Title: Hierarchical organization of long-range circuits in the olfactory cortices.

doi: 10.14814/phy2.12550

Figure Lengend Snippet: Figure 5. Whole-cell recordings of LRMGP-EPSCs in LEC AAV-ChR2-injected mice. (A) Representative experiment with AAV-ChR2 injection in the LEC; a, bright-field image of 300-lm thick horizontal slice showing the LEC injection site (white arrow) and the recording sites (red boxes). The fluorescence of the recorded slice was quenched after DAB staining for neurobiotin, but an adjacent slice in the inset without staining shows that the injection site is located exactly in LEC; b, neurobiotin-labeled neurons; c, monosynaptic AMPA (downward) and AMPA + NMDA (upward) current traces recorded from neurons in three individual cortices. Note the differences of scale bars. (B, C) Comparisons of normalized monosynaptic AMPA and NMDA current strength. (D) a, Bright-field image of an exemplar pair of IN and PYR; b-d, three summary plots showing the comparisons of AMPA, NMDA, and their ratios, respectively, between INs and nearby PYRs recorded in aPC and pPC for LEC injection experiments. *P < 0.05, **P < 0.01.

Article Snippet: Recorded neurons were typically located 50–100 lm below the surface of the recorded slice and had relatively intact dendritic arbors, verified by post hoc neurobiotin reconstruction (antibodies used from the Santa Cruz Biotechnology, Dallas, TX: goat anti-rabbit HRP, sc-2030; Rb HRP, sc-2749; donkey anti-goat HRP, sc2056).

Techniques: Injection, Staining, Labeling