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Deciphering the Neuronal Circuitry Controlling Local Blood Flow in the Cerebral Cortex with Optogenetics in PV::Cre Transgenic Mice.

Urban A, Rancillac A, Martinez L, Rossier J - Front Pharmacol (2012)

Bottom Line: Recent optogenetic experiments combined with functional magnetic resonance imaging have revealed that light stimulation of neurons expressing the calcium binding protein parvalbumin (PV) is associated with positive blood oxygen level-dependent (BOLD) signal in the corresponding barrel field but also with negative BOLD in the surrounding deeper area.Here, we demonstrate that in acute brain slices, channelrhodopsin-2 (ChR2) based photostimulation of PV containing neurons gives rise to an effective contraction of penetrating arterioles.These results support the neurogenic hypothesis of a complex distributed nervous system controlling the CBF.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Neurobiologie, Equipe Optogenetics and Brain Imaging, CNRS UMR 7637, Ecole Supérieure de Physique et de Chimie Industrielles ParisTech PARIS, France.

ABSTRACT
Although it is know since more than a century that neuronal activity is coupled to blood supply regulation, the underlying pathways remains to be identified. In the brain, neuronal activation triggers a local increase of cerebral blood flow (CBF) that is controlled by the neurogliovascular unit composed of terminals of neurons, astrocytes, and blood vessel muscles. It is generally accepted that the regulation of the neurogliovascular unit is adjusted to local metabolic demand by local circuits. Today experimental data led us to realize that the regulatory mechanisms are more complex and that a neuronal system within the brain is devoted to the control of local brain-blood flow. Recent optogenetic experiments combined with functional magnetic resonance imaging have revealed that light stimulation of neurons expressing the calcium binding protein parvalbumin (PV) is associated with positive blood oxygen level-dependent (BOLD) signal in the corresponding barrel field but also with negative BOLD in the surrounding deeper area. Here, we demonstrate that in acute brain slices, channelrhodopsin-2 (ChR2) based photostimulation of PV containing neurons gives rise to an effective contraction of penetrating arterioles. These results support the neurogenic hypothesis of a complex distributed nervous system controlling the CBF.

No MeSH data available.


Related in: MedlinePlus

Schematic representation of DIO/FLEX system to express ChETA-EYFP in PV::Cre mouse by rAAV infection. This strategy is based on two components: a Cre-dependant virus containing ChETA-EYFP expression cassette under the control of an ubiquitous EF1α promoter and a driver transgenic mouse expressing Cre recombinase in neurons containing parvalbumin. The transgene integrates into the host genome allowing stable expression of a fusion protein of ChETA and yellow fluorescent protein (EYFP).
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Figure 1: Schematic representation of DIO/FLEX system to express ChETA-EYFP in PV::Cre mouse by rAAV infection. This strategy is based on two components: a Cre-dependant virus containing ChETA-EYFP expression cassette under the control of an ubiquitous EF1α promoter and a driver transgenic mouse expressing Cre recombinase in neurons containing parvalbumin. The transgene integrates into the host genome allowing stable expression of a fusion protein of ChETA and yellow fluorescent protein (EYFP).

Mentions: Lee et al. (2010) also studied the BOLD effect after light stimulation of fast-spiking inhibitory interneurons. To overcome the low level of opsin produced when a PV promoter fragment drives its expression, they devised a new strategy combining transgenic mice expressing Cre recombinase under the control of the PV promoter (PV::Cre; Hippenmeyer et al., 2005) and a Cre-dependant AAV. The specificity of this system called flip-excision switch (FLEX) or double inverted open (DIO) reading frame is very high by the introduction of two pairs of heterotypic and antiparallel loxP/lox2272 recombination sites that prevent transcriptional read-through observed with classic lox-STOP-lox cassette (Atasoy et al., 2008; Sohal et al., 2009). Moreover, this system decouples transcriptional strength from the specificity of the PV promoter, allowing expression of ChR2-EYFP by the strong EF1α promoter. Thus, the expression of the Cre recombinase directly under the control of the endogenous PV locus restricts the expression of ChR2-EYFP to PV-expressing cells (Figure 1). Consequently, if neurons are infected by DIO/FLEX ChR2-EYFP rAAV but devoid of Cre recombinase, they cannot process the cassette and will not express any of the genetic tools carried by the viral construct.


Deciphering the Neuronal Circuitry Controlling Local Blood Flow in the Cerebral Cortex with Optogenetics in PV::Cre Transgenic Mice.

Urban A, Rancillac A, Martinez L, Rossier J - Front Pharmacol (2012)

Schematic representation of DIO/FLEX system to express ChETA-EYFP in PV::Cre mouse by rAAV infection. This strategy is based on two components: a Cre-dependant virus containing ChETA-EYFP expression cassette under the control of an ubiquitous EF1α promoter and a driver transgenic mouse expressing Cre recombinase in neurons containing parvalbumin. The transgene integrates into the host genome allowing stable expression of a fusion protein of ChETA and yellow fluorescent protein (EYFP).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3375480&req=5

Figure 1: Schematic representation of DIO/FLEX system to express ChETA-EYFP in PV::Cre mouse by rAAV infection. This strategy is based on two components: a Cre-dependant virus containing ChETA-EYFP expression cassette under the control of an ubiquitous EF1α promoter and a driver transgenic mouse expressing Cre recombinase in neurons containing parvalbumin. The transgene integrates into the host genome allowing stable expression of a fusion protein of ChETA and yellow fluorescent protein (EYFP).
Mentions: Lee et al. (2010) also studied the BOLD effect after light stimulation of fast-spiking inhibitory interneurons. To overcome the low level of opsin produced when a PV promoter fragment drives its expression, they devised a new strategy combining transgenic mice expressing Cre recombinase under the control of the PV promoter (PV::Cre; Hippenmeyer et al., 2005) and a Cre-dependant AAV. The specificity of this system called flip-excision switch (FLEX) or double inverted open (DIO) reading frame is very high by the introduction of two pairs of heterotypic and antiparallel loxP/lox2272 recombination sites that prevent transcriptional read-through observed with classic lox-STOP-lox cassette (Atasoy et al., 2008; Sohal et al., 2009). Moreover, this system decouples transcriptional strength from the specificity of the PV promoter, allowing expression of ChR2-EYFP by the strong EF1α promoter. Thus, the expression of the Cre recombinase directly under the control of the endogenous PV locus restricts the expression of ChR2-EYFP to PV-expressing cells (Figure 1). Consequently, if neurons are infected by DIO/FLEX ChR2-EYFP rAAV but devoid of Cre recombinase, they cannot process the cassette and will not express any of the genetic tools carried by the viral construct.

Bottom Line: Recent optogenetic experiments combined with functional magnetic resonance imaging have revealed that light stimulation of neurons expressing the calcium binding protein parvalbumin (PV) is associated with positive blood oxygen level-dependent (BOLD) signal in the corresponding barrel field but also with negative BOLD in the surrounding deeper area.Here, we demonstrate that in acute brain slices, channelrhodopsin-2 (ChR2) based photostimulation of PV containing neurons gives rise to an effective contraction of penetrating arterioles.These results support the neurogenic hypothesis of a complex distributed nervous system controlling the CBF.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Neurobiologie, Equipe Optogenetics and Brain Imaging, CNRS UMR 7637, Ecole Supérieure de Physique et de Chimie Industrielles ParisTech PARIS, France.

ABSTRACT
Although it is know since more than a century that neuronal activity is coupled to blood supply regulation, the underlying pathways remains to be identified. In the brain, neuronal activation triggers a local increase of cerebral blood flow (CBF) that is controlled by the neurogliovascular unit composed of terminals of neurons, astrocytes, and blood vessel muscles. It is generally accepted that the regulation of the neurogliovascular unit is adjusted to local metabolic demand by local circuits. Today experimental data led us to realize that the regulatory mechanisms are more complex and that a neuronal system within the brain is devoted to the control of local brain-blood flow. Recent optogenetic experiments combined with functional magnetic resonance imaging have revealed that light stimulation of neurons expressing the calcium binding protein parvalbumin (PV) is associated with positive blood oxygen level-dependent (BOLD) signal in the corresponding barrel field but also with negative BOLD in the surrounding deeper area. Here, we demonstrate that in acute brain slices, channelrhodopsin-2 (ChR2) based photostimulation of PV containing neurons gives rise to an effective contraction of penetrating arterioles. These results support the neurogenic hypothesis of a complex distributed nervous system controlling the CBF.

No MeSH data available.


Related in: MedlinePlus