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Proceedings Paper

Acousto-optic multiphoton laser scanning microscopy (AO-MPLSM) for structural and functional imaging in living brain slices
Author(s): Vijay Iyer; Tycho Hoogland; Bradley E. Losavio; Rudy Fink; Robert Gaddi; Saumil Patel; Adam Larson; Peter Saggau
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Paper Abstract

The intrinsic optical sectioning, reduced light-scattering, and reduced photodamage of multiphoton laser-scanning microscopy (MPLSM) has generated great interest for this technique in experimental Neuroscience, as it enables to study both structure and function of fine neuronal processes within living brain tissue. At present, virtually all MPLSM systems employ galvanometric beam positioning. Due to this inertia-limited approach, single-dimension line scans are employed to achieve frame rates sufficient for functional imaging. Although such line scans allow adequate sampling rates (≤1kHz), two significant drawbacks remain. First, the majority of scan time is wasted by illuminating regions of no interest, while sacrificing signal integration time at sites-of-interest. Second, the sites from which signals can be recorded are limited to those along a single line. Alternatively, acousto-optic (AO) beam positioning with high-resolution TeO2 deflectors allows inertia-free skipping between arbitrary sites within the field-of-view in <15μs. This achieves high sampling rate recording at multiple, non-adjacent sites quasi-simultaneously (1-5kHz frame rate, 12-60 sites). Such a multi-site optical recording system would greatly advance studying complex neuronal function, by enabling membrane potential or calcium transients to be observed throughout the complex geometry of neuronal dendrites. This paper presents images and functional recordings from living neurons within brain slices, acquired with AO-MPLSM. Our novel imaging system allows a user to collect structural images first and subsequently select sites of interest for fast functional imaging. To demonstrate the system’s power, we present high-speed recordings (1kHz) from >10 sites within the dendrites of pyramidal neurons in acute brain slices, at signal-to-noise ratios comparable to line-scan systems.

Paper Details

Date Published: 30 March 2005
PDF: 12 pages
Proc. SPIE 5700, Multiphoton Microscopy in the Biomedical Sciences V, (30 March 2005); doi: 10.1117/12.587430
Show Author Affiliations
Vijay Iyer, Rice Univ. (United States)
Baylor College of Medicine (United States)
Tycho Hoogland, Baylor College of Medicine (United States)
Bradley E. Losavio, Baylor College of Medicine (United States)
Rudy Fink, Baylor College of Medicine (United States)
Robert Gaddi, Baylor College of Medicine (United States)
Saumil Patel, Baylor College of Medicine (United States)
Adam Larson, Baylor College of Medicine (United States)
Peter Saggau, Rice Univ. (United States)
Baylor College of Medicine (United States)

Published in SPIE Proceedings Vol. 5700:
Multiphoton Microscopy in the Biomedical Sciences V
Ammasi Periasamy; Peter T. C. So, Editor(s)

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