Abstract
We introduce an optical method to stimulate individual neurons in brain slices in any arbitrary spatiotemporal pattern, using two-photon uncaging of MNI-glutamate with beam multiplexing. This method has single-cell and three-dimensional precision. By sequentially stimulating up to a thousand potential presynaptic neurons, we generated detailed functional maps of inputs to a cell. We combined this approach with two-photon calcium imaging in an all-optical method to image and manipulate circuit activity.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
References
Sterling, P. Retina. in The Synaptic Organization of the Brain (ed. G.M. Shepherd) 170–213 (Oxford University Press, Oxford, 1990).
Crick, F.H. Thinking about the brain. Sci. Am. 241, 219–232 (1979).
Farber, I.C. & Grinvald, A. Identification of presynaptic neurons by laser photostimulation. Science 222, 1025–1027 (1983).
Callaway, E.M. & Katz, L.C. Photostimulation using caged glutamate reveals functional circuitry in living brain slices. Proc. Natl. Acad. Sci. USA 90, 7661–7665 (1993).
Dalva, M.B. & Katz, L.C. Rearrangements of synaptic connections in visual cortex revealed by laser photostimulation. Science 265, 255–258 (1994).
Shepherd, G.M., Pologruto, T.A. & Svoboda, K. Circuit analysis of experience-dependent plasticity in the developing rat barrel cortex. Neuron 38, 277–289 (2003).
Kotter, R., Schubert, D., Dyhrfjeld-Johnsen, J., Luhmann, H.J. & Staiger, J.F. Optical release of caged glutamate for stimulation of neurons in the in vitro slice preparation. J. Biomed. Opt. 10, 11003 (2005).
Dodt, H.U., Schierloh, A., Eder, M. & Zieglgansberger, W. Circuitry of rat barrel cortex investigated by infrared-guided laser stimulation. Neuroreport 14, 623–627 (2003).
Boucsein, C., Nawrot, M., Rotter, S., Aertsen, A. & Heck, D. Controlling synaptic input patterns in vitro by dynamic photo stimulation. J. Neurophysiol. 94, 2948–2958 (2005).
Shoham, S., O'Connor, D.H., Sarkisov, D.V. & Wang, S.S. Rapid neurotransmitter uncaging in spatially defined patterns. Nat. Methods 2, 837–843 (2005).
Denk, W., Strickler, J.H. & Webb, W.W. Two-photon laser scanning fluorescence microscopy. Science 248, 73–76 (1990).
Canepari, M., Nelson, L., Papageorgiou, G., Corrie, J.E. & Ogden, D. Photochemical and pharmacological evaluation of 7-nitroindolinyl-and 4-methoxy-7-nitroindolinyl-amino acids as novel, fast caged neurotransmitters. J. Neurosci. Methods 112, 29–42 (2001).
Ellis-Davies, G.C.R. Development and application of calcium cages. Methods Enymol. 360A, 226–238 (2003).
Yuste, R. & MacLean, J. Loading populations neurons in slices with AM calcium indicators. in Imaging Neurons: a Laboratory Manual (eds. R. Yuste, F. Lanni & A. Konnerth) 351–355 (Cold Spring Harbor Press, Cold Spring Harbor, NY, 2005).
Nimmerjahn, A., Kirchhoff, F., Kerr, J.N. & Helmchen, F. Sulforhodamine 101 as a specific marker of astroglia in the neocortex in vivo. Nat. Methods 1, 31–37 (2004).
Cossart, R., Aronov, D. & Yuste, R. Attractor dynamics of network UP states in neocortex. Nature 423, 283–289 (2003).
Nikolenko, V., Nemet, B. & Yuste, R. A custom two-photon and second harmonic microscope. Methods 30, 3–5 (2003).
Peterlin, Z.A., Kozloski, J., Mao, B., Tsiola, A. & Yuste, R. Optical probing of neuronal circuits with calcium indicators. Proc. Natl. Acad. Sci. USA 97, 3619–3624 (2000).
Sacconi, L.F.E., Antolini, R., Taghizadeh, M.R., Choudhury, A. & Pavone, F.S. Multiphoton multifocal microscopy exploiting a diffractive optical element. Opt. Lett. 28, 1918–1920 (2003).
Yoshimura, Y., Dantzker, J.L. & Callaway, E.M. Excitatory cortical neurons form fine-scale functional networks. Nature 433, 868–873 (2005).
Le Be, J.V. & Markram, H. Spontaneous and evoked synaptic rewiring in the neonatal neocortex. Proc. Natl. Acad. Sci. USA 103, 13214–13219 (2006).
Yuste, R. & Katz, L.C. Control of postsynaptic Ca2+ influx in developing neocortex by excitatory and inhibitory neurotransmitters. Neuron 6, 333–344 (1991).
Kozloski, J., Hamzei-Sichani, F. & Yuste, R. Stereotyped position of local synaptic targets in neocortex. Science 293, 868–872 (2001).
Wang, Y. et al. Anatomical, physiological and molecular properties of Martinotti cells in the somatosensory cortex of the juvenile rat. J. Physiol. (Lond.) 561, 65–90 (2004).
Badea, T., Goldberg, J., Mao, B.Q. & Yuste, R. Calcium imaging of epileptiform events with single-cell resolution. J. Neurobiol. 48, 215–227 (2001).
Trevelyan, A.J., Sussillo, D., Watson, B.O. & Yuste, R. Modular propagation of epileptiform activity: evidence for an inhibitory veto in neocortex. J. Neurosci. 26, 12447–12455 (2006).
Boyden, E.S., Zhang, F., Bamberg, E., Nagel, G. & Deisseroth, K. Millisecond-timescale, genetically targeted optical control of neural activity. Nat. Neurosci. 8, 1263–1268 (2005).
Zhang, F. et al. Multimodal fast optical interrogation of neural circuitry. Nature 446, 633–639 (2007).
Majewska, A., Yiu, G. & Yuste, R. A custom-made two-photon microscope and deconvolution system. Pflügers Arch. 441, 398–409 (2000).
Oliva, A.A., Jr., Jiang, M., Lam, T., Smith, K.L. & Swann, J.W. Novel hippocampal interneuronal subtypes identified using transgenic mice that express green fluorescent protein in GABAergic interneurons. J. Neurosci. 20, 3354–3368 (2000).
Acknowledgements
We thank E. Callaway for advice, A. Packer for online analysis, B. Nemet for help with optics, G. Ellis-Davies for initial samples of MNI-glutamate, D. Aronov for the convex hull algorithm, Y. Duanmu for histology, and L. Abbott, E. Schaffer and members of the laboratory for comments. Supported by the US National Eye Institute, the National Institute of Neurological Disorders and Stroke, the New York State Foundation for Science Technology and Innovation (NYSTAR) program and the Kavli Institute for Brain Studies. K.E.P. is a Patterson Trust postdoctoral fellow. We dedicate this study to the memory of Larry Katz, who pioneered and stimulated these experiments.
Author information
Authors and Affiliations
Corresponding author
Supplementary information
Supplementary Text and Figures
Supplementary figures 1–5, Supplementary Note, Supplementary Methods (PDF 1033 kb)
Rights and permissions
About this article
Cite this article
Nikolenko, V., Poskanzer, K. & Yuste, R. Two-photon photostimulation and imaging of neural circuits. Nat Methods 4, 943–950 (2007). https://doi.org/10.1038/nmeth1105
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nmeth1105
This article is cited by
-
Axon hillock currents enable single-neuron-resolved 3D reconstruction using diamond nitrogen-vacancy magnetometry
Communications Physics (2020)
-
NeuroPath2Path: Classification and elastic morphing between neuronal arbors using path-wise similarity
Neuroinformatics (2020)
-
Closed-loop all-optical interrogation of neural circuits in vivo
Nature Methods (2018)