labelling with neurobiotin  (Vector Laboratories)

Journal: Nature Communications

Article Title: Genetic silencing of olivocerebellar synapses causes dystonia-like behaviour in mice

doi: 10.1038/ncomms14912

Figure Lengend Snippet: ( a ) A schematic showing a cerebellar recording in an anaesthetized mouse. Schematic adapted with permission, from drawings published in ref. . ( b ) A schematic depicting a single-unit recording from a Purkinje cell with its climbing fibre. Samples of a simple spike (SS) and a climbing fibre-driven complex spike (CS) are shown. ( c ) Molecular expression showing the one-to-one relationship between a climbing fibre and a Purkinje cell with VGLUT2 marking the climbing fibre terminals. Scale bar, 20 μm. ( d ) A schematic of the juxtacellular labelling process. Current pulses are used to find cells based on increased resistance. Once a cell is found, the recording step begins by loosely patching on the membrane of the cell, and recording its extracellular spiking activity. After recording for a sufficient period of time, the cell is filled by delivering neurobiotin using 500 ms pulses of positive current between 1 and 5 nA for ∼1 h. Schematic adapted with permission, from drawings published in ref. . ( e ) Examples of traces from 3-month-old adult Vglut2 fx/fx and Ptf1a Cre ;Vglut2 fx/fx Purkinje cells either using metal electrodes for extracellular recording or glass electrodes for juxtacellular recording. Complex spikes are labelled with asterisks. Note the lack of complex spikes in the mutant traces. ( f ) Examples of filled Purkinje cells in adult 3–5-month-old Vglut2 fx/fx and Ptf1a Cre ;Vglut2 fx/fx mice. n >4 cells for each genotype. Scale bar, 50 μm. ( g ) Quantification of the number of Purkinje cells with identifiable complex spikes in Vglut2 fx/fx mice compared with recordings from Ptf1a Cre ;Vglut2 fx/fx mice of all ages from postnatal day 19 to 8 months. This analysis includes data from all of our electrophysiology experiments. The Purkinje cells were recorded from all lobules of the vermis and from crusI and II of the hemispheres. gl, granular layer; pcl, Purkinje cell layer; ml, molecular layer; NPY, neuropeptide Y (in this case a transgenic allele drives GFP expression).

Article Snippet: The mice were then perfused and the neurobiotin labelling of single filled cells detected using the VECTASTAIN Elite ABC method (Vector Laboratories Inc.) and visualized with DAB.

Techniques: Single-unit Recording, Expressing, Membrane, Activity Assay, Mutagenesis, Transgenic Assay

Journal: The Journal of Neuroscience

Article Title: Medial Septal GABAergic Neurons Express the Somatostatin sst 2A Receptor: Functional Consequences on Unit Firing and Hippocampal Theta

doi: 10.1523/JNEUROSCI.4619-04.2005

Figure Lengend Snippet: Immunohistochemical identification and electrophysiological characterization of a single juxtacellular labeled PV/sst2A neuron in the rat MS-DB. A, Single confocal section showing a recorded neuron labeled with neurobiotin (red). This neuron was also immunoreactive for parvalbumin (B, blue) and sst2A receptors (C, green) as illustrated in D. E, Extracellular recording from the neurobiotin-labeled neuron shown in A-D. The discharge profile is characterized by a theta-related bursting activity (4.0 Hz) and a high discharge rate (45 spike/s). F, Expanded scale from E indicating the short duration of the spike (0.38 ms). Scale bar: (in D) A-D, 10 μm.

Article Snippet: To characterize the neurochemical identity of recorded neurons, juxtacellular labelings with neurobiotin (Vector Laboratories, Burlingame, CA) were performed in 10 rats.

Techniques: Immunohistochemical staining, Labeling, Activity Assay

Journal: The Journal of Neuroscience

Article Title: Medial Septal GABAergic Neurons Express the Somatostatin sst 2A Receptor: Functional Consequences on Unit Firing and Hippocampal Theta

doi: 10.1523/JNEUROSCI.4619-04.2005

Figure Lengend Snippet: Immunohistochemical identification and electrophysiological characterization of a single juxtacellular labeled GAD67/sst2A neuron in the rat MS-DB. A, Single confocal section showing a recorded neuron labeled with neurobiotin (red). This neuron was also immunoreactive for GAD67 (B, green) and sst2A receptors (C, blue) as illustrated in D. E, Extracellular recording from the neurobiotin-labeled neuron shown in A-D. The discharge profile is characterized by a theta-related bursting activity (4.5 Hz) and a high discharge rate (24 spike/s). F, Expanded scale from E indicating the short duration of the spike (0.39 ms). Scale bar: (in D) A-D, 10 μm.

Article Snippet: To characterize the neurochemical identity of recorded neurons, juxtacellular labelings with neurobiotin (Vector Laboratories, Burlingame, CA) were performed in 10 rats.

Techniques: Immunohistochemical staining, Labeling, Activity Assay

Journal:

Article Title: Contribution of a calcium-activated non-specific conductance to NMDA receptor-mediated synaptic potentials in granule cells of the frog olfactory bulb

doi: 10.1113/jphysiol.2002.024638

Figure Lengend Snippet: OB neurones were distinguished by their anatomical and electrophysiological properties in blind whole-cell recordings. Ai and Bi, reconstructions of neurobiotin-labelled cells from horizontal OB sections (300 μm). GC interneurones (Ai) were characterized by widely branching dendrites projecting rostrally in horizontal sections. GC dendrites were covered in gemmules and long-necked spines, up to 10 μm in length (insets - photomicrographs of the regions delineated by the boxes; arrowheads mark cell somata). MTC dendrites, by contrast, were aspiny. In the MTC shown (Bi) an apical dendrite can be followed rostrally, where it ramifies in an olfactory glomerulus (inset). Aii and Bii, traces of spontaneous synaptic activity from the cells in Ai and Bi show the characteristic differences between GC and MTC electrophysiology at rest. In the nose-brain preparation, large (> 5 mV) spontaneous depolarizing potentials in GCs occurred at rates of ≈5 Hz, while spontaneous synaptic activity was less prominent in MTCs. Rinput, input resistance. Aiii and Biii, lateral olfactory tract (LOT) stimulation evoked an antidromic AP in MTCs, which, in turn, synaptically activated the GC population. The antidromic AP in MTCs rose straight from baseline. Hyperpolarization (lower trace) did not block the AP but reversed the synaptically mediated after-hyperpolarization (the AP is truncated in the lower trace). In GCs, LOT stimulation produced an EPSP that increased at more hyperpolarized levels, while the AP was blocked.

Article Snippet: Neuronal labelling Neurobiotin (Vector Laboratories, Burlingame, CA, USA) was added to the intracellular pipette solution (0.5 % w/v) and cells were filled by passive diffusion.

Techniques: Activity Assay, Blocking Assay, Produced