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Fluorescent protein voltage probes derived from ArcLight that respond to membrane voltage changes with fast kinetics.

Han Z, Jin L, Platisa J, Cohen LB, Baker BJ, Pieribone VA - PLoS ONE (2013)

Bottom Line: ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites.Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human.We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ) less than 6 ms for a 100 mV depolarization.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut, United States of America ; The John B. Pierce Laboratory, Inc., New Haven, Connecticut, United States of America.

ABSTRACT
We previously reported the discovery of a fluorescent protein voltage probe, ArcLight, and its derivatives that exhibit large changes in fluorescence intensity in response to changes of plasma membrane voltage. ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites. The response kinetics of ArcLight (τ1-on ~10 ms, τ2-on ~ 50 ms) are comparable with most published genetically-encoded voltage probes. However, probes using voltage-sensing domains other than that from the Ciona intestinalis voltage sensitive phosphatase exhibit faster kinetics. Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human. We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ) less than 6 ms for a 100 mV depolarization. Although the response amplitude of these two probes (8-9%) is not as large as the Ciona-based ArcLight (~35%), they are better at reporting action potentials from cultured neurons at higher frequency. In contrast, probes based on frog, mouse and human voltage sensing domains were either slower than the Ciona-based ArcLight or had very small signals.

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Related in: MedlinePlus

Chicken Arclight-A173 resolves 100 Hz, 2 ms depolarization pulses.A). Two HEK 293 cells transfected with chicken ArcLight-A173 imaged with the fast CCD camera. The patched cell is in the top-right of this image. The ROI is shown in red. B). The red trace is –ΔF/F of the optical recording. The blue trace is the command voltage pulses. Overlapping the traces shows the temporal relationship between the optical recording and the command voltage pulses. It is most likely that the membrane was not fully charged during the pulse. We estimated that the actual stepped potential to be on the order of 70mV [17].
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pone-0081295-g005: Chicken Arclight-A173 resolves 100 Hz, 2 ms depolarization pulses.A). Two HEK 293 cells transfected with chicken ArcLight-A173 imaged with the fast CCD camera. The patched cell is in the top-right of this image. The ROI is shown in red. B). The red trace is –ΔF/F of the optical recording. The blue trace is the command voltage pulses. Overlapping the traces shows the temporal relationship between the optical recording and the command voltage pulses. It is most likely that the membrane was not fully charged during the pulse. We estimated that the actual stepped potential to be on the order of 70mV [17].

Mentions: Expression of chicken ArcLight-A173 and Q175 in cultured mouse cortical neurons produced brightly fluorescent cells with membrane targeted expression in the soma and dendrites (Figure 4A and B). They appear largely membrane localized, similar to the Ciona-based ArcLight. In spite of the relatively small response amplitude of the chicken ArcLight in HEK 293 cells (~9% ΔF/F in response to a 100mV depolarization), we could detect evoked action potentials in single trials in neurons expressing this probe (Figure 4). The mean soma response was -3.2 ± 2.2% change in the fluorescence intensity (n=20 cells). We also tested whether chicken ArcLight is able to resolve voltage signals at a high frequency. We applied 100Hz depolarizations of 2 ms duration to HEK293 cells transfected with chicken ArcLight-A173 and these signals were resolved in single trials (Figure 5).


Fluorescent protein voltage probes derived from ArcLight that respond to membrane voltage changes with fast kinetics.

Han Z, Jin L, Platisa J, Cohen LB, Baker BJ, Pieribone VA - PLoS ONE (2013)

Chicken Arclight-A173 resolves 100 Hz, 2 ms depolarization pulses.A). Two HEK 293 cells transfected with chicken ArcLight-A173 imaged with the fast CCD camera. The patched cell is in the top-right of this image. The ROI is shown in red. B). The red trace is –ΔF/F of the optical recording. The blue trace is the command voltage pulses. Overlapping the traces shows the temporal relationship between the optical recording and the command voltage pulses. It is most likely that the membrane was not fully charged during the pulse. We estimated that the actual stepped potential to be on the order of 70mV [17].
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3842285&req=5

pone-0081295-g005: Chicken Arclight-A173 resolves 100 Hz, 2 ms depolarization pulses.A). Two HEK 293 cells transfected with chicken ArcLight-A173 imaged with the fast CCD camera. The patched cell is in the top-right of this image. The ROI is shown in red. B). The red trace is –ΔF/F of the optical recording. The blue trace is the command voltage pulses. Overlapping the traces shows the temporal relationship between the optical recording and the command voltage pulses. It is most likely that the membrane was not fully charged during the pulse. We estimated that the actual stepped potential to be on the order of 70mV [17].
Mentions: Expression of chicken ArcLight-A173 and Q175 in cultured mouse cortical neurons produced brightly fluorescent cells with membrane targeted expression in the soma and dendrites (Figure 4A and B). They appear largely membrane localized, similar to the Ciona-based ArcLight. In spite of the relatively small response amplitude of the chicken ArcLight in HEK 293 cells (~9% ΔF/F in response to a 100mV depolarization), we could detect evoked action potentials in single trials in neurons expressing this probe (Figure 4). The mean soma response was -3.2 ± 2.2% change in the fluorescence intensity (n=20 cells). We also tested whether chicken ArcLight is able to resolve voltage signals at a high frequency. We applied 100Hz depolarizations of 2 ms duration to HEK293 cells transfected with chicken ArcLight-A173 and these signals were resolved in single trials (Figure 5).

Bottom Line: ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites.Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human.We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ) less than 6 ms for a 100 mV depolarization.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut, United States of America ; The John B. Pierce Laboratory, Inc., New Haven, Connecticut, United States of America.

ABSTRACT
We previously reported the discovery of a fluorescent protein voltage probe, ArcLight, and its derivatives that exhibit large changes in fluorescence intensity in response to changes of plasma membrane voltage. ArcLight allows the reliable detection of single action potentials and sub-threshold activities in individual neurons and dendrites. The response kinetics of ArcLight (τ1-on ~10 ms, τ2-on ~ 50 ms) are comparable with most published genetically-encoded voltage probes. However, probes using voltage-sensing domains other than that from the Ciona intestinalis voltage sensitive phosphatase exhibit faster kinetics. Here we report new versions of ArcLight, in which the Ciona voltage-sensing domain was replaced with those from chicken, zebrafish, frog, mouse or human. We found that the chicken and zebrafish-based ArcLight exhibit faster kinetics, with a time constant (τ) less than 6 ms for a 100 mV depolarization. Although the response amplitude of these two probes (8-9%) is not as large as the Ciona-based ArcLight (~35%), they are better at reporting action potentials from cultured neurons at higher frequency. In contrast, probes based on frog, mouse and human voltage sensing domains were either slower than the Ciona-based ArcLight or had very small signals.

Show MeSH
Related in: MedlinePlus