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Second-Harmonic Generation Imaging of Membrane Potential with Photon Counting

Published online by Cambridge University Press:  06 November 2008

Jiang Jiang  and
Rafael Yuste
Jiang Jiang*
Affiliation:
HHMI, Department of Biological Sciences, Columbia University, New York, NY 10027, USA
Rafael Yuste
Affiliation:
HHMI, Department of Biological Sciences, Columbia University, New York, NY 10027, USA
*
Corresponding author. E-mail: [email protected]
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Abstract

Second-harmonic generation (SHG) can be used for imaging membrane potential in neurons, but poor signal-to-noise (S/N) limits accurate measurements of small voltage transients. We use photon counting to improve the S/N of weak SHG signal detection. Photon counting generates shot-noise limited and integrable signals, eliminates pulse-to-pulse variation, and built-in discriminators reduces the background to practically zero. In single trials, by using photon counting, we obtain a more than a twofold S/N increase over analog voltage detection. Trial-to-trial variability is also reduced by 50%. Finally, we show that, using photon counting, the kinetics of fast events such as action potentials can be recorded more accurately.

Keywords

second-harmonic generationphoton countingneuronmembrane potential
Type
Multiphoton Microscopy–Special Section
Information
Microscopy and Microanalysis , Volume 14 , Issue 6 , December 2008 , pp. 526 - 531
Copyright
Copyright © Microscopy Society of America 2008

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References

REFERENCES

Dombeck, D.A., Blanchard-Desce, M. & Webb, W.W. (2004). Optical recording of action potentials with second-harmonic generation microscopy. J Neurosci 24(4), 999–1003.CrossRefGoogle ScholarPubMed
Dombeck, D.A., Sacconi, L., Blanchard-Desce, M. & Webb, W.W. (2005). Optical recording of fast neuronal membrane potential transients in acute mammalian brain slices by second-harmonic generation microscopy. J Neurophysiol 94(5), 3628–3636.Google Scholar
Eisenthal, K.B. (1996). Liquid interfaces probed by second-harmonic and sum-frequency spectroscopy. Chem Rev 96(4), 1343–1360.CrossRefGoogle ScholarPubMed
Iyer, V., Hoogland, T.M. & Saggau, P. (2006). Fast functional imaging of single neurons using random-access multiphoton (RAMP) microscopy. J Neurophysiol 95(1), 535–545.CrossRefGoogle ScholarPubMed
Jiang, J., Eisenthal, K.B. & Yuste, R. (2007). Second harmonic generation in neurons: Electro-optic mechanism of membrane potential sensitivity. Biophys J 93(5), L26–28.CrossRefGoogle ScholarPubMed
Millard, A.C., Campagnola, P.J., Mohler, W., Lewis, A. & Loew, L.M. (2003a). Second harmonic imaging microscopy. Meth Enzymol 361, 47–69.Google Scholar
Millard, A.C., Jin, L., Lewis, A. & Loew, L.M. (2003b). Direct measurement of the voltage sensitivity of second-harmonic generation from a membrane dye in patch-clamped cells. Opt Lett 28(14), 1221–1223.CrossRefGoogle ScholarPubMed
Moreaux, L., Sandre, O., Blanchard-Desce, M. & Mertz, J. (2000). Membrane imaging by simultaneous second-harmonic generation and two-photon microscopy. Opt Lett 25(5), 320–322.Google Scholar
Moreaux, L., Sandre, O., Charpak, S., Blanchard-Desce, M. & Mertz, J. (2001). Coherent scattering in multi-harmonic light microscopy. Biophys J 80(3), 1568–1574.CrossRefGoogle ScholarPubMed
Nemet, B.A., Nikolenko, V. & Yuste, R. (2004). Second harmonic imaging of membrane potential of neurons with retinal. J Biomed Opt 9(5), 873–881.Google Scholar
Nikolenko, V., Nemet, B. & Yuste, R. (2003). A two-photon and second-harmonic microscope. Methods 30(1), 3–15.Google Scholar
Nuriya, M., Jiang, J., Nemet, B., Eisenthal, K.B. & Yuste, R. (2006). Imaging membrane potential in dendritic spines. Proc Natl Acad Sci USA 103(3), 786–790.CrossRefGoogle ScholarPubMed
Pons, T., Moreaux, L. & Mertz, J. (2002). Photoinduced flip-flop of amphiphilic molecules in lipid bilayer membranes. Phys Rev Lett 89(28), 288104.CrossRefGoogle ScholarPubMed
Sacconi, L., Dombeck, D.A. & Webb, W.W. (2006). Overcoming photodamage in second-harmonic generation microscopy: Real-time optical recording of neuronal action potentials. Proc Natl Acad Sci USA 103(9), 3124–3129.Google Scholar
Salomé, R., Kremer, Y., Dieudonné, S., Léger, J.F., Krichevsky, O., Wyart, C., Chatenay, D. & Bourdieu, L. (2006). Ultrafast random-access scanning in two-photon microscopy using acousto-optic deflectors. J Neurosci Meth 154(1–2), 161–174.CrossRefGoogle ScholarPubMed
Svoboda, K., Tank, D.W. & Denk, W. (1996). Direct measurement of coupling between dendritic spines and shafts. Science 272(5262), 716–719.CrossRefGoogle ScholarPubMed
Yuste, R., Nemet, B., Jiang, J., Nuriya, M. & Eisenthal, K. (2005). Second harmonic generation imaging of membrane potential. In Imaging Neurons and Neural Activity: New Methods, New Results, pp. 16–17. Cold Spring Harbor, NY: Cold Spring Harbor Press.Google Scholar