optogenetic control of hippocampal network dynamics

Optogenetic Control of Hippocampal Network Dynamics

In this procedure, place an LFP electrode in the CA1 subiculum area, and patch a nearby pyramidal cell in the isolated hippocampus of a mouse expressing the blue light sensitive excitatory opsin ChR2 in PV interneurons.
Place an optic fiber light guide above the hippocampal preparation, and center it on the recorded region. Use blue light from an LED source for up to genetic stimulation, which consists of 10 to 20-millisecond light pulses or sine wave voltage commands delivered at theta frequencies. In current clamp, characterize the activity of the recorded cell during spontaneous theta oscillations. Then, start the stimulation protocol and record the light responses.
Observe that field oscillations and synaptic activity in the recorded neuron become increasingly synchronized during optogenetic stimulation, and that rhythmic pacing of PV cells results in a robust control of both frequency and power of theta oscillations.

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All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee.
<ol>
	<li>Optogenetic control in the isolated hippocampus 
	NOTE: For optogenetic control of hippocampal network activity, a cell-type specific strategy involving rhythmic activation of PV (parvalbumin-positive)-containing GABAergic cells is used here as these cells were shown to play a major role in synchronizing principal cell populations in the isolated hippocampus. Selective expression of the excitatory channelrhodopsin-2 (ChR2) in PV cells is achieved by crossing the PV-Cre mouse line with mice constitutively expressing ChR2 Cre-dependently (Ai32), thereby generating PV-ChY mice. Alternatively, viral vector constructs (e.g., AAVdj-ChETA-eYFP) can be injected into the hippocampus of PV-Cre or PV-TOM mice (P15-16) to drive the expression of the excitatory opsin in CA1/SUB interneurons (Figure 1A).
	<ol>
		<li>Place an LFP electrode in the CA1/SUB (Cornu ammonis-1/subiculum) area and patch a nearby pyramidal cell in the isolated hippocampus of a mouse expressing the blue-light sensitive excitatory opsin ChR2 in PV interneurons.</li>
		<li>Place an optic fiber light guide (0.2 - 1 mm diameter) above the hippocampal preparation and center it on the recorded region. Use blue light (473 nm) from an LED source for optogenetic stimulation, consisting of light pulses (10 - 20 ms, triggered by a transistor-transistor logic signal) or sine wave voltage commands delivered at theta frequencies (4 - 12 Hz).</li>
		<li>Switch to the current clamp and characterize the activity of the pyramidal during spontaneous theta oscillations.</li>
		<li>Start the stimulation protocol and record the light responses. Observe that field oscillations and synaptic activity in the recorded neuron become increasingly synchronized during optogenetic stimulation and that rhythmic pacing of PV cells results in robust control of both frequency and power of theta oscillations (Figure 1).</li>
	</ol>
	</li>
</ol>

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			<td>95% O2/5% CO2 gas mixture (carbogen)</td>
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			<td>perfusion system</td>
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			<td>Glass bottles/flasks (4 x 1 L)</td>
			<td>n.a.</td>
			<td>n.a.</td>
			<td>perfusion system</td>
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			<td>Submerged recording Chamber</td>
			<td>custom design (FM)</td>
			<td>n.a.</td>
			<td>Commercial alternative may be used</td>
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			<td>Multiclamp 700B patch-clamp amplifier</td>
			<td>Molecular devices</td>
			<td>MULTICLAMP</td>
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			<td>Model 3300</td>
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			<td>Custom-built upright fluorescence microscope</td>
			<td>Siskiyou</td>
			<td>n.a.</td>
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			<td>Analogue video camera</td>
			<td>COHU</td>
			<td>4912-2000/0000</td>
			<td>Imaging</td>
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			<td>Digital frame grabber with imaging software</td>
			<td>EPIX, Inc</td>
			<td>PIXCI-SV7</td>
			<td>Imaging</td>
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			<td>Olympus 2.5x objective</td>
			<td>Olympus</td>
			<td>MPLFLN</td>
			<td>Imaging</td>
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			<td>Olympus 40x water immersion objective</td>
			<td>Olympus</td>
			<td>UIS2 LUMPLFLN</td>
			<td>Imaging</td>
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		<tr>
			<td>Custom-made light-emitting diode (LED) system</td>
			<td>custom</td>
			<td>n.a.</td>
			<td>optogenetic stimulation (Amhilon et al., 2015)</td>
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			<td>Animals</td>
			<td></td>
			<td></td>
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			<td>PV::Cre (KI) mice</td>
			<td>Jackson Laboratory</td>
			<td>stock number 008069</td>
			<td>Allow Cre-directed gene expression in PV interneurons</td>
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			<td>Constitutive-conditional Ai9 mice (R26-lox-stop-lox-tdTomato (KI))</td>
			<td>Jackson Laboratory</td>
			<td>stock number 007905</td>
			<td>Express TdTomato following Cre-mediated recombination</td>
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			<td>Ai32 mice (R26-lox-stop-lox-ChR2(H134R)-EYFP</td>
			<td>Jackson Laboratory</td>
			<td>stock number 012569</td>
			<td>Express the improved channelrhodopsin-2/EYFP fusion protein following exposure to Cre recombinase</td>
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			<td>PVChY mice</td>
			<td>In house breeding</td>
			<td>n.a.</td>
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Source: Manseau, F., et al. <a href="https://app.jove.com/v/55851">Tuning in the Hippocampal Theta Band In Vitro: Methodologies for Recording from the Isolated Rodent Septohippocampal Circuit.</a> J. Vis. Exp. (2017)
This video demonstrates the method to use optogenetics to modulate hippocampal network dynamics by activating genetically modified light-sensitive interneurons in a mouse brain slice, enhancing theta oscillation synchronization and studying neural circuit behavior.

<img alt="Figure 1" src="/files/ftp_upload/22951/22951_Figure1.jpg" /> 
Figure 1: Local Field Potential and Simultaneous Patch Clamp Recordings from Single CA1/SUB Pyramidal and PV Neurons during Spontaneous and Optogenetically-driven Theta Oscillations in the Isolated Hippocampus.
(a) Diagram showing the LFP and patch electrode recording sites with the placement of the light-fiber guide above the CA1/SUB region expressing...

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee.

	Optogenetic control in the ...

Begin with a mouse hippocampal slice containing the target pyramidal cells connected to genetically modified light-sensitive interneurons.
Place the slice in a recording chamber.
Position a local field potential or LFP electrode in the desired region to record theta oscillations, a type of rhythmic brain activity.
Using a microelectrode, patch a nearby pyramidal cell to record its activity.
Place an optic fiber light above the hippocampal slice centered on the recording region.
Deliver blue light pulses at a set frequency to the slice to activate the light-sensitive ion channels in the interneuron.
The activation causes an influx of positive ions, leading to depolarization.
This triggers the release of inhibitory neurotransmitters onto the pyramidal neuron, thereby reducing excitability.
The light stimulation enhances the synchronization of the theta oscillations and the synaptic activity of the recorded neuron, suggesting effective modulation of the neural rhythms.

16 vistas. Control optogenético de la dinámica de la red del hipocampo

Video: Control optogenético de la dinámica de la red del hipocampo - Experimento

Automatic Detection of Highly Organized Theta Oscillations in the Murine EEG

Theta activity is generated in the septohippocampal system and can be recorded using deep intrahippocampal electrodes and implantable electroencephalography (EEG) radiotelemetry or tether system approaches. Pharmacologically, hippocampal theta is heterogeneous (see dualistic theory) and can be differentiated into type I and type II theta. These individual EEG subtypes are related to specific cognitive and behavioral states, such as arousal, exploration, learning and memory, higher integrative functions, etc. In neurodegenerative diseases such as Alzheimer&#39;s, structural and functional alterations of the septohippocampal system can result in impaired theta activity/oscillations. A standard quantitative analysis of the hippocampal EEG includes a Fast-Fourier-Transformation (FFT)-based frequency analysis. However, this procedure does not provide details about theta activity in general and highly-organized theta oscillations in particular. In order to obtain detailed information on highly-organized theta oscillations in the hippocampus, we have developed a new analytical approach. This approach allows for time- and cost-effective quantification of the duration of highly-organized theta oscillations and their frequency characteristics.

Theta activity in the hippocampus is related to specific cognitive and behavioral stages. Here, we describe an analytical method to detect highly-organized theta oscillations within the hippocampus using a time-frequency (i.e., wavelet analysis)-based approach.

Detección automática de muy organizada oscilaciones theta en el EEG murino

Theta activity is generated in the septohippocampal system and can be recorded using deep intrahippocampal electrodes and implantable ...

Investigación

JoVE Journal

Comportamiento

Optogenetic Entrainment of Hippocampal Theta Oscillations in Behaving Mice

Extensive data on relationships of neural network oscillations to behavior and organization of neuronal discharge across brain regions call for new tools to selectively manipulate brain rhythms. Here we describe an approach combining projection-specific optogenetics with extracellular electrophysiology for high-fidelity control of hippocampal theta oscillations (5-10 Hz) in behaving mice. The specificity of the optogenetic entrainment is achieved by targeting channelrhodopsin-2 (ChR2) to the GABAergic population of medial septal cells, crucially involved in the generation of hippocampal theta oscillations, and a local synchronized activation of a subset of inhibitory septal afferents in the hippocampus. The efficacy of the optogenetic rhythm control is verified by a simultaneous monitoring of the local field potential (LFP) across lamina of the CA1 area and/or of neuronal discharge. Using this readily implementable preparation we show efficacy of various optogenetic stimulation protocols for induction of theta oscillations and for the manipulation of their frequency and regularity. Finally, a combination of the theta rhythm control with projection-specific inhibition addresses the readout of particular aspects of the hippocampal synchronization by efferent regions.

We describe the use of optogenetics and electrophysiological recordings for selective manipulations of hippocampal theta oscillations (5-10 Hz) in behaving mice. The efficacy of the rhythm entrainment is monitored using local field potential. A combination of opto- and pharmacogenetic inhibition addresses the efferent readout of hippocampal synchronization.

Optogenetic de arrastre de las oscilaciones Theta Hippocampal en comportarse ratones

Extensive data on relationships of neural network oscillations to behavior and organization of neuronal discharge across brain regions call for new tools ...

Neurociencias

Recording Spatially Restricted Oscillations in the Hippocampus of Behaving Mice

The local field potential (LFP) emerges from ion movements across neural membranes. Since the voltage recorded by LFP electrodes reflects the summed electrical field of a large volume of brain tissue, extracting information about local activity is challenging. Studying neuronal microcircuits, however, requires a reliable distinction between truly local events and volume-conducted signals originating in distant brain areas. Current source density (CSD) analysis offers a solution for this problem by providing information about current sinks and sources in the vicinity of the electrodes. In brain areas with laminar cytoarchitecture such as the hippocampus, one-dimensional CSD can be obtained by estimating the second spatial derivative of the LFP. Here, we describe a method to record multilaminar LFPs using linear silicon probes implanted into the dorsal hippocampus. CSD traces are computed along individual shanks of the probe. This protocol thus describes a procedure to resolve spatially restricted neuronal network oscillations in the hippocampus of freely moving mice.

This protocol describes the recording of local field potentials with multi-shank linear silicon probes. Conversion of the signals using current source density analysis allows the reconstruction of local electrical activity in the mouse hippocampus. With this technique, spatially restricted brain oscillations can be studied in freely moving mice.

Grabación espacial restringido a oscilaciones en el hipocampo de ratones de comportarse

The local field potential (LFP) emerges from ion movements across neural membranes. Since the voltage recorded by LFP electrodes reflects the summed ...

Optogenetic Control of Hippocampal Network Dynamics

Transcripción

Optogenetic Control of Hippocampal Network Dynamics