Home LiteratureArticle Details
PMID: 9199810 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

High-speed, random-access fluorescence microscopy: I. High-resolution optical recording with voltage-sensitive dyes and ion indicators.

Biophysical journal ·Vol. 73 ·No. 1 ·1997-07-00 ·Pages 477-91

Bullen A, Patel SS, Saggau P

Abstract

The design and implementation of a high-speed, random-access, laser-scanning fluorescence microscope configured to record fast physiological signals from small neuronal structures with high spatiotemporal resolution is presented. The laser-scanning capability of this nonimaging microscope is provided by two orthogonal acousto-optic deflectors under computer control. Each scanning point can be randomly accessed and has a positioning time of 3-5 microseconds. Sampling time is also computer-controlled and can be varied to maximize the signal-to-noise ratio. Acquisition rates up to 200k samples/s at 16-bit digitizing resolution are possible. The spatial resolution of this instrument is determined by the minimal spot size at the level of the preparation (i.e., 2-7 microns). Scanning points are selected interactively from a reference image collected with differential interference contrast optics and a video camera. Frame rates up to 5 kHz are easily attainable. Intrinsic variations in laser light intensity and scanning spot brightness are overcome by an on-line signal-processing scheme. Representative records obtained with this instrument by using voltage-sensitive dyes and calcium indicators demonstrate the ability to make fast, high-fidelity measurements of membrane potential and intracellular calcium at high spatial resolution (2 microns) without any temporal averaging.

MeSH Terms
Animals Cells, Cultured Equipment Design Fluorescent Dyes Hippocampus/cytology Indicators and Reagents Ions Membrane Potentials Microscopy, Fluorescence/instrumentation,methods Neurons/cytology,physiology Organic Chemicals Sensitivity and Specificity Software Time Factors
Chemicals
Fluorescent Dyes Indicators and Reagents Ions Organic Chemicals calcium orange
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Bullen A
Division of Neuroscience, Baylor College of Medicine, Houston, Texas 77030, USA.
Patel S S
Saggau P
References (28)
28 references, click to expand
  1. [Clinical and laboratory aspects of Kahler's disease. Considerations on 19 cases].
    Med Interna (Bucur). 1966 Sep;18(9):1049-57 PMID: 5975886
  2. An acousto-optically steered laser scanning system for measurement of action potential spread in intact heart.
    Soc Gen Physiol Ser. 1986;40:211-26 PMID: 3487121
  3. Optical recording of neuronal activity in an invertebrate central nervous system: simultaneous monitoring of several neurons.
    J Neurophysiol. 1977 Nov;40(6):1281-91 PMID: 925730
  4. Species-specific effects on the optical signals of voltage-sensitive dyes.
    J Membr Biol. 1979 Aug;48(4):343-56 PMID: 490629
  5. Optical methods for monitoring neuron activity.
    Annu Rev Neurosci. 1978;1:171-82 PMID: 386900
  6. Presynaptic calcium currents in squid giant synapse.
    Biophys J. 1981 Mar;33(3):289-321 PMID: 7225510
  7. Optical recording of calcium action potentials from growth cones of cultured neurons with a laser microbeam.
    Science. 1981 Jun 5;212(4499):1164-7 PMID: 7233210
  8. A new laser scanning system for measuring action potential propagation in the heart.
    Science. 1981 Oct 23;214(4519):453-6 PMID: 6974891
  9. Real-time optical mapping of neuronal activity: from single growth cones to the intact mammalian brain.
    Annu Rev Neurosci. 1985;8:263-305 PMID: 3885828
  10. Optical monitoring of membrane potential: methods of multisite optical measurement.
    Soc Gen Physiol Ser. 1986;40:71-99 PMID: 3520842
  11. Optical recording of synaptic potentials from processes of single neurons using intracellular potentiometric dyes.
    Biophys J. 1987 Apr;51(4):643-51 PMID: 3580490
  12. Optical measurement of action potential activity in invertebrate ganglia.
    Annu Rev Physiol. 1989;51:527-41 PMID: 2653195
  13. Optical imaging of cortical activity: real-time imaging using extrinsic dye-signals and high resolution imaging based on slow intrinsic-signals.
    Annu Rev Physiol. 1989;51:543-59 PMID: 2653196
  14. A confocal video-rate laser-beam scanning reflected-light microscope with no moving parts.
    J Microsc. 1990 Jan;157(Pt 1):29-38 PMID: 2299661
  15. Voltage-sensitive dye recording of action potentials and synaptic potentials from sympathetic microcultures.
    Biophys J. 1991 Sep;60(3):697-711 PMID: 1681956
  16. An apparatus for recording synaptic potentials from neuronal cultures using voltage-sensitive fluorescent dyes.
    J Neurosci Methods. 1991 Jul;38(2-3):93-105 PMID: 1784131
  17. A naphthyl analog of the aminostyryl pyridinium class of potentiometric membrane dyes shows consistent sensitivity in a variety of tissue, cell, and model membrane preparations.
    J Membr Biol. 1992 Oct;130(1):1-10 PMID: 1469705
  18. Optical probing of active membrane potential using lasers and microoptics.
    Jpn J Physiol. 1993;43 Suppl 1:S53-6 PMID: 7505857
  19. Characterization of impulse propagation at the microscopic level across geometrically defined expansions of excitable tissue: multiple site optical recording of transmembrane voltage (MSORTV) in patterned growth heart cell cultures.
    J Gen Physiol. 1994 Aug;104(2):287-309 PMID: 7807050
  20. Multiple site optical recording of transmembrane voltage (MSORTV) in patterned growth heart cell cultures: assessing electrical behavior, with microsecond resolution, on a cellular and subcellular scale.
    Biophys J. 1994 Sep;67(3):1301-15 PMID: 7811945
  21. Spread of epileptiform activity in the immature rat neocortex studied with voltage-sensitive dyes and laser scanning microscopy.
    J Neurophysiol. 1994 Oct;72(4):1756-68 PMID: 7823100
  22. Optical signals from neurons with internally applied voltage-sensitive dyes.
    J Neurosci. 1995 Feb;15(2):1392-405 PMID: 7869106
  23. Use of voltage-sensitive dyes and optical recordings in the central nervous system.
    Prog Neurobiol. 1995 Aug;46(5):463-506 PMID: 8532849
  24. Simultaneous optical recording of evoked and spontaneous transients of membrane potential and intracellular calcium concentration with high spatio-temporal resolution.
    J Neurosci Methods. 1995 Aug;60(1-2):49-60 PMID: 8544487
  25. Near-membrane [Ca2+] transients resolved using the Ca2+ indicator FFP18.
    Proc Natl Acad Sci U S A. 1996 May 28;93(11):5368-73 PMID: 8643581
  26. Multiple spike-initiation zones in single neurons revealed by voltage-sensitive dyes.
    Nature. 1996 May 23;381(6580):322-5 PMID: 8692270
  27. Cable properties of cultured hippocampal neurons determined from sucrose-evoked miniature EPSCs.
    J Neurophysiol. 1996 Mar;75(3):1250-5 PMID: 8867133
  28. Changes in axon light scattering that accompany the action potential: current-dependent components.
    J Physiol. 1972 Aug;224(3):727-52 PMID: 5071935
Article Info
Journal
Biophysical journal
Abbr.
Biophys J
ISSN
0006-3495
DOI
10.1016/S0006-3495(97)78086-X
Published
1997-07-00
Pages
477-91
Language
English
Region
United States
NLM ID
0370626
PMCID
PMC1180947
Subset
IM
Grants
NINDS NIH HHS · NS33147 · United States
External Links
PubMed source DOI: 10.1016/S0006-3495(97)78086-X PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2