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A multimodal micro-optrode combining field and single unit recording, multispectral detection and photolabeling capabilities.

Dufour S, Lavertu G, Dufour-Beauséjour S, Juneau-Fecteau A, Calakos N, Deschênes M, Vallée R, De Koninck Y - PLoS ONE (2013)

Bottom Line: Here, we describe a, aluminum-coated, fibre optic-based glass microprobe with multiple electrical and optical detection capabilities while retaining tip dimensions that enable single cell measurements (diameter ≤10 µm).It also enables color conversion of photoswitchable fluorescent proteins, which can be used for post-hoc identification of the recorded cells.The extended range of functionalities provided by the same microprobe thus opens several avenues for multidimensional structural and functional interrogation of single cells and their surrounding deep within the intact nervous system.

View Article: PubMed Central - PubMed

Affiliation: Unité de neurosciences cellulaires et moléculaires, Institut universitaire en santé mentale de Québec, Québec, Québec, Canada.

ABSTRACT
Microelectrodes have been very instrumental and minimally invasive for in vivo functional studies from deep brain structures. However they are limited in the amount of information they provide. Here, we describe a, aluminum-coated, fibre optic-based glass microprobe with multiple electrical and optical detection capabilities while retaining tip dimensions that enable single cell measurements (diameter ≤10 µm). The probe enables optical separation from individual cells in transgenic mice expressing multiple fluorescent proteins in distinct populations of neurons within the same deep brain nucleus. It also enables color conversion of photoswitchable fluorescent proteins, which can be used for post-hoc identification of the recorded cells. While metal coating did not significantly improve the optical separation capabilities of the microprobe, the combination of metal on the outside of the probe and of a hollow core within the fiber yields a microelectrode enabling simultaneous single unit and population field potential recordings. The extended range of functionalities provided by the same microprobe thus opens several avenues for multidimensional structural and functional interrogation of single cells and their surrounding deep within the intact nervous system.

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Multispectral detection allows identifying D1 and D2 expressing neurons.A) Schema of the shutters state and incident signals on the detecting PMTs as the probe passes by a green and a red cell in succession. B) (Left) Micrograph of a GFP fluorescent cell and a microprobe (highlighted with gray contour). Scale bar is 10 µm. (Right) Collected fluorescence in the PMT detector 1 (green dots), and 2 (red dots) as the probe is moved transversally in front of the cell shown in left panel. C) Same result is shown as the probe pass by a tdTomato fluorescent cell. Scale bar is 10 µm. Arrows in (B) and (C) represent the probe displacement in front of the cell. D) Striatal section of a transgenic mouse showing cells expressing fluorescent proteins under the control of D1 (red) or D2 (green) receptor (green) promoters. Some cells co-express both fluorescent proteins (see arrow). E) Average density of the different cell types in the striatum (n = 5 striatal sections). Cells were counted and densities were estimated with the help of the software Image J. F) Histogram of the optically detected cells. Fluorescent cell detections were accompanied by a rise (signal-to-noise >2) and a decay of green and/or red fluorescence as described previously. A total of 29 cells were detected and computed in a histogram according to their fluorescence colour. (G-H) Examples of in vivo simultaneous optical and electrical recordings (inset) as the probe pass by a green (G) or a red (H) cell.
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pone-0057703-g002: Multispectral detection allows identifying D1 and D2 expressing neurons.A) Schema of the shutters state and incident signals on the detecting PMTs as the probe passes by a green and a red cell in succession. B) (Left) Micrograph of a GFP fluorescent cell and a microprobe (highlighted with gray contour). Scale bar is 10 µm. (Right) Collected fluorescence in the PMT detector 1 (green dots), and 2 (red dots) as the probe is moved transversally in front of the cell shown in left panel. C) Same result is shown as the probe pass by a tdTomato fluorescent cell. Scale bar is 10 µm. Arrows in (B) and (C) represent the probe displacement in front of the cell. D) Striatal section of a transgenic mouse showing cells expressing fluorescent proteins under the control of D1 (red) or D2 (green) receptor (green) promoters. Some cells co-express both fluorescent proteins (see arrow). E) Average density of the different cell types in the striatum (n = 5 striatal sections). Cells were counted and densities were estimated with the help of the software Image J. F) Histogram of the optically detected cells. Fluorescent cell detections were accompanied by a rise (signal-to-noise >2) and a decay of green and/or red fluorescence as described previously. A total of 29 cells were detected and computed in a histogram according to their fluorescence colour. (G-H) Examples of in vivo simultaneous optical and electrical recordings (inset) as the probe pass by a green (G) or a red (H) cell.

Mentions: Multispectral detection is used in microscopy to discriminate cells labeled with proteins having different emission spectra. To test the compatibility of multispectral detection with our microprobe, we injected light from two different excitation sources in the probe optical channel. The detection system proposed here (see schematics in Fig. 2a) was successfully used to differentiate two cell populations labeled with two different fluorescent markers in vitro and in vivo. Recorded fluorescence signals as a function of the probe position in front of a fixed GFP cell is shown in Figure 2b. The same result is shown for a tdTomato cell (Fig. 2c). Fluorescence increase and decrease only in the corresponding PMT signal as the probe passed the cell. To test the ability of the probe to differentiate between two labeled populations of cells in vivo, we performed vertical scanning experiments through the striatum of adult transgenic mice expressing tdTomato and EGFP proteins under the control of the D1 and D2 dopamine receptor promoters, respectively [6]–[8]. Probes descents were repeated within the following setreotaxic coordinates: 1.5 to 2.5 mm lateromedial, 0.5 to 1.5 anterior to Bregma and 2 to 3.5 mm below brain surface. Figure 2d and 2e shows the distribution of tdTomato and EGFP in the striatum. Cells expressing the D1 and D2 receptors could be clearly identified (Fig. 2d) and very few cells expressed both markers. Identification of D1 and D2 cells with the microprobe yielded a similar distribution of D1-only, D2-only and combined D1/D2 cell detection as that obtained under microscopy analysis (Fig. 2f). Optical detection of EGFP and tdTomato expressing cells respectively is shown in Figure 2g–h along with five superimposed consecutive spikes recorded at fluorescence peaks. The microprobe thus provides a means to identify and record from different labeled populations of cells intermingled within the same deep brain structure.


A multimodal micro-optrode combining field and single unit recording, multispectral detection and photolabeling capabilities.

Dufour S, Lavertu G, Dufour-Beauséjour S, Juneau-Fecteau A, Calakos N, Deschênes M, Vallée R, De Koninck Y - PLoS ONE (2013)

Multispectral detection allows identifying D1 and D2 expressing neurons.A) Schema of the shutters state and incident signals on the detecting PMTs as the probe passes by a green and a red cell in succession. B) (Left) Micrograph of a GFP fluorescent cell and a microprobe (highlighted with gray contour). Scale bar is 10 µm. (Right) Collected fluorescence in the PMT detector 1 (green dots), and 2 (red dots) as the probe is moved transversally in front of the cell shown in left panel. C) Same result is shown as the probe pass by a tdTomato fluorescent cell. Scale bar is 10 µm. Arrows in (B) and (C) represent the probe displacement in front of the cell. D) Striatal section of a transgenic mouse showing cells expressing fluorescent proteins under the control of D1 (red) or D2 (green) receptor (green) promoters. Some cells co-express both fluorescent proteins (see arrow). E) Average density of the different cell types in the striatum (n = 5 striatal sections). Cells were counted and densities were estimated with the help of the software Image J. F) Histogram of the optically detected cells. Fluorescent cell detections were accompanied by a rise (signal-to-noise >2) and a decay of green and/or red fluorescence as described previously. A total of 29 cells were detected and computed in a histogram according to their fluorescence colour. (G-H) Examples of in vivo simultaneous optical and electrical recordings (inset) as the probe pass by a green (G) or a red (H) cell.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585187&req=5

pone-0057703-g002: Multispectral detection allows identifying D1 and D2 expressing neurons.A) Schema of the shutters state and incident signals on the detecting PMTs as the probe passes by a green and a red cell in succession. B) (Left) Micrograph of a GFP fluorescent cell and a microprobe (highlighted with gray contour). Scale bar is 10 µm. (Right) Collected fluorescence in the PMT detector 1 (green dots), and 2 (red dots) as the probe is moved transversally in front of the cell shown in left panel. C) Same result is shown as the probe pass by a tdTomato fluorescent cell. Scale bar is 10 µm. Arrows in (B) and (C) represent the probe displacement in front of the cell. D) Striatal section of a transgenic mouse showing cells expressing fluorescent proteins under the control of D1 (red) or D2 (green) receptor (green) promoters. Some cells co-express both fluorescent proteins (see arrow). E) Average density of the different cell types in the striatum (n = 5 striatal sections). Cells were counted and densities were estimated with the help of the software Image J. F) Histogram of the optically detected cells. Fluorescent cell detections were accompanied by a rise (signal-to-noise >2) and a decay of green and/or red fluorescence as described previously. A total of 29 cells were detected and computed in a histogram according to their fluorescence colour. (G-H) Examples of in vivo simultaneous optical and electrical recordings (inset) as the probe pass by a green (G) or a red (H) cell.
Mentions: Multispectral detection is used in microscopy to discriminate cells labeled with proteins having different emission spectra. To test the compatibility of multispectral detection with our microprobe, we injected light from two different excitation sources in the probe optical channel. The detection system proposed here (see schematics in Fig. 2a) was successfully used to differentiate two cell populations labeled with two different fluorescent markers in vitro and in vivo. Recorded fluorescence signals as a function of the probe position in front of a fixed GFP cell is shown in Figure 2b. The same result is shown for a tdTomato cell (Fig. 2c). Fluorescence increase and decrease only in the corresponding PMT signal as the probe passed the cell. To test the ability of the probe to differentiate between two labeled populations of cells in vivo, we performed vertical scanning experiments through the striatum of adult transgenic mice expressing tdTomato and EGFP proteins under the control of the D1 and D2 dopamine receptor promoters, respectively [6]–[8]. Probes descents were repeated within the following setreotaxic coordinates: 1.5 to 2.5 mm lateromedial, 0.5 to 1.5 anterior to Bregma and 2 to 3.5 mm below brain surface. Figure 2d and 2e shows the distribution of tdTomato and EGFP in the striatum. Cells expressing the D1 and D2 receptors could be clearly identified (Fig. 2d) and very few cells expressed both markers. Identification of D1 and D2 cells with the microprobe yielded a similar distribution of D1-only, D2-only and combined D1/D2 cell detection as that obtained under microscopy analysis (Fig. 2f). Optical detection of EGFP and tdTomato expressing cells respectively is shown in Figure 2g–h along with five superimposed consecutive spikes recorded at fluorescence peaks. The microprobe thus provides a means to identify and record from different labeled populations of cells intermingled within the same deep brain structure.

Bottom Line: Here, we describe a, aluminum-coated, fibre optic-based glass microprobe with multiple electrical and optical detection capabilities while retaining tip dimensions that enable single cell measurements (diameter ≤10 µm).It also enables color conversion of photoswitchable fluorescent proteins, which can be used for post-hoc identification of the recorded cells.The extended range of functionalities provided by the same microprobe thus opens several avenues for multidimensional structural and functional interrogation of single cells and their surrounding deep within the intact nervous system.

View Article: PubMed Central - PubMed

Affiliation: Unité de neurosciences cellulaires et moléculaires, Institut universitaire en santé mentale de Québec, Québec, Québec, Canada.

ABSTRACT
Microelectrodes have been very instrumental and minimally invasive for in vivo functional studies from deep brain structures. However they are limited in the amount of information they provide. Here, we describe a, aluminum-coated, fibre optic-based glass microprobe with multiple electrical and optical detection capabilities while retaining tip dimensions that enable single cell measurements (diameter ≤10 µm). The probe enables optical separation from individual cells in transgenic mice expressing multiple fluorescent proteins in distinct populations of neurons within the same deep brain nucleus. It also enables color conversion of photoswitchable fluorescent proteins, which can be used for post-hoc identification of the recorded cells. While metal coating did not significantly improve the optical separation capabilities of the microprobe, the combination of metal on the outside of the probe and of a hollow core within the fiber yields a microelectrode enabling simultaneous single unit and population field potential recordings. The extended range of functionalities provided by the same microprobe thus opens several avenues for multidimensional structural and functional interrogation of single cells and their surrounding deep within the intact nervous system.

Show MeSH
Related in: MedlinePlus