Monthly Archives: February 2013

Long-term channelrhodopsin-2 expression can induce abnormal axonal morphology and targeting in cerebral cortex

When using optogenetics to study circuit function or animal behavior, a critical prerequisite is that optogenetic protein expression does not, in itself, perturb the circuit being studied. While short-term expression is very commonly used without observable circuit disruption, whether this is also true for long-term expression is less clear. A recent paper by Miyashita et al. in Frontiers in Neural Circuits shows that long-term, high-level expression of ChR2 can induce abnormal axonal morphology and targeting in cerebral cortex. This underscores the importance of using the lowest expression possible, particularly for long-term studies.

Miyashita et al. expressed a common construct, CAG::hChR2 (H134R)-EYFP-WPRE, in L2/3 pyramidal neurons in rat somatosensory cortex via in utero electroporation (IUE). This same strategy was used in several prior studies of S1 circuit function, with one important difference: Miyashita et al. expressed hChR2 that was codon-optimized for mammalian expression, while prior studies expressed native ChR2 (discussed below). This strategy successfully conferred light-evoked spiking in vivo and in in vitro brain slices. However, long-term expression (> 40 d) also caused major abnormalities in axonal morphology, which included cylinders of axonal membrane that enveloped pyramidal cell proximal dendrites, and spherical, calyx-like axonal swellings that surrounded neuron cell bodies. These … Continue reading

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The optopatcher: an electrode holder allowing the insertion of an optical fiber into a patch pipette

In order to perform simultaneous intracellular recording and light stimulation of a single neuron, two separate positioning systems are often needed (one to position the recording electrode, one to position a waveguide near the recorded neuron). More sophisticated solutions for single neuron photostimulation involve light patterning techniques which are not suited for deep in vivo recordings. Katz et al. came up with a simple and affordable solution for this problem, by designing a patch pipette holder containing an additional port for the insertion of an optical fiber into the pipette.

This device, which they called “OptoPatcher” allows whole cell patch-clamp recording simultaneously with direct projection of light from the recording pipette. The holder spares the use of an additional manipulator and, importantly, enables accurate, stable and reproducible illumination. Moreover, the presence of the bare fiber within an aqueous solution instead of the brain can prevent tissue damage due to heating of the brain. In addition, replacement of standard pipettes is done as easily as with the available commercial holders.

The OptoPatcher was used successfully in vivo for intracellular recordings from different cortical layers in the motor cortex of transgenic mice expressing channelrhodopsin-2 under the Thy1 promoter and it was also … Continue reading

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Micro-electrocorticogram electrodes for simultaneous recording and optical stimulation

Wen Li’s group at Michigan State University recently presented preliminary results on the development of an epidural micro-electrocorticogram (μECoG) array combining microelectrodes and light emitting diodes (LEDs) for optical neural stimulation. This work follows-up on a new line of research aiming at providing μECoG arrays with versatile optical stimulation capabilities (see for example the work of Ledochowitsch et al). The “Opto-μECoG” arrays developped in Wen Li’s group were especially designed to address three major limitations of current designs, in particular the limited cortical area and spatial resolution available for optical stimulation. The key features of these Opto-μECoG arrays are the following:

Untethered system: integration of surface mounted μ-LED light sources (220 × 270 ×50 μm3, wavelength peak at 460nm, Cree® TR2227TM) on the Opto-μECoG array allows the possibility to achieve a truly untethered system. Maximized target cortical area available for optical stimulation: optically transparent indium tin oxide (ITO) [2] epidural electrodes of the Opto-μECoG array allow maximum exposure of the target cortical area for optical stimulation. Maximized spatial resolution of optical stimulation: the embedded light sources, placed on top of the ITO electrodes, were preciously arranged based on a recent study of the optimal spacing of subdural, epidural, and scalp … Continue reading

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Stop lights for temporal lobe seizures

Unlike electrical stimulation, optogenetics allows neuronal manipulation with great cell-type specificity, with light directly affecting only those cells expressing opsins. In a recent report in Nature Communications, Krook-Magnuson et al harnessed this specificity to stop seizures in vivo in a mouse model of temporal lobe epilepsy. Mice were implanted with electrodes to record brain activity and 200µm thick optical fibers to deliver light to the brain. A closed-loop, on-demand responsive system detected seizures in real time, allowing temporal specificity, in addition to the cell-type specificity achieved through selective opsin expression. Specifically, the authors either selectively inhibited excitatory principal cells or, alternatively, excited a subpopulation of GABAergic inhibitory neurons in the hippocampus by delivering light at the time of a seizure. Both approaches proved successful, despite the less than 5% of illuminated neurons expressing opsins in the latter approach. Light arrested ongoing electrical seizure activity and reduced the incidence of events progressing to overt behavioral seizures.

Epilepsy, a condition of recurrent, spontaneous seizures, is a prevalent disorder, with 1 out of 26 people developing epilepsy during their lifetime. Unfortunately, for over 40% of patients, seizures cannot be controlled with current treatment options. Temporal lobe epilepsy, the most common form of epilepsy … Continue reading

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