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Chronic cranial window with access port for repeated cellular manipulations, drug application, and electrophysiology.

Roome CJ, Kuhn B - Front Cell Neurosci (2014)

Bottom Line: This chronic cranial window with access port is cheap, easy to manufacture, can be mounted just as the regular chronic cranial window, and is self-sealing after retraction of the pipette or electrode.As an example to the extent of its utility for electrophysiological recording, we describe how simple removal of the silicone seal can permit patch pipette access for whole-cell patch clamp recordings in vivo.During these chronic experiments we do not observe any infections under the window or impairment of animal health.

View Article: PubMed Central - PubMed

Affiliation: Optical Neuroimaging Unit, Okinawa Institute of Science and Technology Graduate University Onna-son, Okinawa, Japan.

ABSTRACT
Chronic cranial windows have been instrumental in advancing optical studies in vivo, permitting long-term, high-resolution imaging in various brain regions. However, once a window is attached it is difficult to regain access to the brain under the window for cellular manipulations. Here we describe a simple device that combines long term in vivo optical imaging with direct brain access via glass or quartz pipettes and metal, glass, or quartz electrodes for cellular manipulations like dye or drug injections and electrophysiological stimulations or recordings while keeping the craniotomy sterile. Our device comprises a regular cranial window glass coverslip with a drilled access hole later sealed with biocompatible silicone. This chronic cranial window with access port is cheap, easy to manufacture, can be mounted just as the regular chronic cranial window, and is self-sealing after retraction of the pipette or electrode. We demonstrate that multiple injections can be performed through the silicone port by repetitively bolus loading calcium sensitive dye into mouse barrel cortex and recording spontaneous cellular activity over a period of weeks. As an example to the extent of its utility for electrophysiological recording, we describe how simple removal of the silicone seal can permit patch pipette access for whole-cell patch clamp recordings in vivo. During these chronic experiments we do not observe any infections under the window or impairment of animal health.

No MeSH data available.


Related in: MedlinePlus

Chronic cranial window with access port for electrophysiology. While metal electrodes and beveled electrodes for single or multi-unit recording can simply penetrate the silicone plug, in our hands, it was necessary to remove the plug for patch clamp experiments to reach a high success rate (A). A GFP-expressing layer 2/3 pyramidal neuron shown in x-z plane projection (B) and x-y plane projection (C) was targeted for whole-cell patch clamp recording under anesthesia (1% isoflurane) (D) and filled with a red fluorescent dye after plug removal. The access port does not affect the imaging quality as shown by in vivo imaging of GFP-expressing neurons in layer 5 of barrel cortex after viral gene transfer (E).
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Figure 4: Chronic cranial window with access port for electrophysiology. While metal electrodes and beveled electrodes for single or multi-unit recording can simply penetrate the silicone plug, in our hands, it was necessary to remove the plug for patch clamp experiments to reach a high success rate (A). A GFP-expressing layer 2/3 pyramidal neuron shown in x-z plane projection (B) and x-y plane projection (C) was targeted for whole-cell patch clamp recording under anesthesia (1% isoflurane) (D) and filled with a red fluorescent dye after plug removal. The access port does not affect the imaging quality as shown by in vivo imaging of GFP-expressing neurons in layer 5 of barrel cortex after viral gene transfer (E).

Mentions: As a second demonstration of how the chronic cranial window with access port may be applied, we conducted targeted whole cell patch clamp recordings (n = 3) on layer 2/3 pyramidal cells previously labeled with GFP (Figure 4).


Chronic cranial window with access port for repeated cellular manipulations, drug application, and electrophysiology.

Roome CJ, Kuhn B - Front Cell Neurosci (2014)

Chronic cranial window with access port for electrophysiology. While metal electrodes and beveled electrodes for single or multi-unit recording can simply penetrate the silicone plug, in our hands, it was necessary to remove the plug for patch clamp experiments to reach a high success rate (A). A GFP-expressing layer 2/3 pyramidal neuron shown in x-z plane projection (B) and x-y plane projection (C) was targeted for whole-cell patch clamp recording under anesthesia (1% isoflurane) (D) and filled with a red fluorescent dye after plug removal. The access port does not affect the imaging quality as shown by in vivo imaging of GFP-expressing neurons in layer 5 of barrel cortex after viral gene transfer (E).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4227473&req=5

Figure 4: Chronic cranial window with access port for electrophysiology. While metal electrodes and beveled electrodes for single or multi-unit recording can simply penetrate the silicone plug, in our hands, it was necessary to remove the plug for patch clamp experiments to reach a high success rate (A). A GFP-expressing layer 2/3 pyramidal neuron shown in x-z plane projection (B) and x-y plane projection (C) was targeted for whole-cell patch clamp recording under anesthesia (1% isoflurane) (D) and filled with a red fluorescent dye after plug removal. The access port does not affect the imaging quality as shown by in vivo imaging of GFP-expressing neurons in layer 5 of barrel cortex after viral gene transfer (E).
Mentions: As a second demonstration of how the chronic cranial window with access port may be applied, we conducted targeted whole cell patch clamp recordings (n = 3) on layer 2/3 pyramidal cells previously labeled with GFP (Figure 4).

Bottom Line: This chronic cranial window with access port is cheap, easy to manufacture, can be mounted just as the regular chronic cranial window, and is self-sealing after retraction of the pipette or electrode.As an example to the extent of its utility for electrophysiological recording, we describe how simple removal of the silicone seal can permit patch pipette access for whole-cell patch clamp recordings in vivo.During these chronic experiments we do not observe any infections under the window or impairment of animal health.

View Article: PubMed Central - PubMed

Affiliation: Optical Neuroimaging Unit, Okinawa Institute of Science and Technology Graduate University Onna-son, Okinawa, Japan.

ABSTRACT
Chronic cranial windows have been instrumental in advancing optical studies in vivo, permitting long-term, high-resolution imaging in various brain regions. However, once a window is attached it is difficult to regain access to the brain under the window for cellular manipulations. Here we describe a simple device that combines long term in vivo optical imaging with direct brain access via glass or quartz pipettes and metal, glass, or quartz electrodes for cellular manipulations like dye or drug injections and electrophysiological stimulations or recordings while keeping the craniotomy sterile. Our device comprises a regular cranial window glass coverslip with a drilled access hole later sealed with biocompatible silicone. This chronic cranial window with access port is cheap, easy to manufacture, can be mounted just as the regular chronic cranial window, and is self-sealing after retraction of the pipette or electrode. We demonstrate that multiple injections can be performed through the silicone port by repetitively bolus loading calcium sensitive dye into mouse barrel cortex and recording spontaneous cellular activity over a period of weeks. As an example to the extent of its utility for electrophysiological recording, we describe how simple removal of the silicone seal can permit patch pipette access for whole-cell patch clamp recordings in vivo. During these chronic experiments we do not observe any infections under the window or impairment of animal health.

No MeSH data available.


Related in: MedlinePlus