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Presentation Abstract

Program#/Poster#: 590.12/XX18

Presentation Title: In vivo profiling of astrocytic calcium excitability and downstream vascular effects:

Fluorescence intensity and lifetime measurements

Location: Hall F-J

Presentation time: Tuesday, Oct 16, 2012, 11:00 AM -12:00 PM

Authors: *H. UHLIROVA1, P. A. SAISAN2, E. S. NORHEIM2,4, Q. CHENG2, K. L.

WELDY2, S. L. GRATIY1, G. T. EINEVOLL4, A. M. DALE3, A. DEVOR3,5; 1Radiology, 2Neurosciences, 3Neurosciences & Radiology, Univ. of California San

Diego, La Jolla, CA; 4Dept. of Mathematical Sci. and Technol. and Ctr. for

Integrative Genet. (CIGENE), Norwegian Univ. of Life Sci., Oslo, Norway; 5MGH,

Harvard Med. Sch., Charlestown, MA

Abstract: Astrocytes express a number of metabotropic receptors for messenger molecules,including glutamate and ATP, and can exhibit elevations of intracellular calcium upon

their activation. An increase in astrocytic calcium, in turn, can lead to release ofgliotransmitters implicated in neuroglial and gliovascular interactions. In this study, we

quantitatively assessed in vivo astrocytic calcium dynamics and their downstream

vascular effects in response to local micropharmacologocal stimulations. Time-

resolved astrocytic calcium concentration (in nM) was measured through

fluorescence lifetime imaging microscopy (FLIM) of a popular calcium indicator,

Oregon Green BAPTA-1 AM (1-3). To achieve a comparable temporal resolution

to the widely used fluorescence intensity calcium imaging (~10 Hz), we implemented

user-defined scanning trajectories aiming to collect sufficient numbers of photons

from a region of interest (e.g., a single astrocytic cell body) per time-point (Figure

1A). Our data show large increases in astrocytic intracellular calcium (>150 nM at

the peak) in the absence of arteriolar dilation, and arteriolar dilation in the absence ofthe astrocytic calcium increase. In addition, an increase in astrocytic intracellular

calcium concentration measured by FLIM exceeded the ΔF/F obtained by

fluorescence intensity imaging (Figure 1B). This effect might result from the presence

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of indicator molecules in intracellular calcium stores isolated from the cytosolic

calcium changes. Figure 1. Astrocytic dynamics of the absolute calcium concentration

in vivo. A. Field of view with 3 astrocytes (yellow). Scan trajectory for FLIM

measurement is shown on the right. B-C. FLIM (B) and intensity (C) single-trial

time-courses in response to a micropuff of 1 mM ATP. Red vertical lines indicate the

timing of the puff.1. Wilms CD, et al. (2006) Cell Calcium 40(1):73-79.

2. Lattarulo C, et al.(2011) Methods Mol Biol 793:377-389.

3. Gersbach M, et al. (2009) Opt Lett 34(3):362-364.

Disclosures: H. Uhlirova: None. P.A. Saisan: None. E.S. Norheim: None. Q. Cheng:None. K.L. Weldy: None. S.L. Gratiy: None. G.T. Einevoll: None. A.M. Dale:

None. A. Devor: None.

Keyword(s): 2-photon microscopy

ASTROCYTE

BLOOD FLOW

Support: NIH Grant NS051188

NIH Grant NS057198

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NIH Grant EB009118

NIH Grant EB00790

European Regional Development Fund CEITEC

[Authors]. [Abstract Title]. Program No. XXX.XX. 2012 Neuroscience MeetingPlanner. New Orleans, LA: Society for Neuroscience, 2012. Online.

2012 Copyright by the Society for Neuroscience all rights reserved. Permission to

republish any abstract or part of any abstract in any form must be obtained in writingby SfN office prior to publication.