TY - JOUR
T1 - Simultaneous optical recording of membrane potential and intracellular calcium from brain slices
AU - Sinha, Saurabh R.
AU - Saggau, Peter
N1 - Funding Information:
We thank Dr. S. S. Patel for his work in writing most of the software used in this work and for his efforts in developing and maintaining the hardware. This work was supported by an NRSA (MH-10491) from NIMH to S.R.S. and by an NIH grant (NS-33147) to P.S.
Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 1999/6
Y1 - 1999/6
N2 - Optical recording techniques provide a constantly evolving and increasingly powerful set of tools for investigations of cellular physiology. These techniques rely on the use of optical indicators, molecules that change their optical properties depending on the cellular parameter of interest. In this paper we discuss some of the general considerations involved in recording optical signals from multiple indicators. specifically, we describe a technique for simultaneously recording transients of membrane potential and intracellular calcium concentration, two parameters that have a very complex interrelationship in neuronal functioning. This technique relies on the use of two fluorescent indicators (the voltage-sensitive dye RH-414 and the calcium-sensitive dye Calcium Orange) that have overlapping excitation spectra but separable emission spectra. This fact, in combination with the use of fast, spatially resolving photodetectors (10 x 10-element photodiode matrices), allows for truly simultaneous recording of these transients from brain slices with high spatial (~ 200 x 200 μm with a 10 x microscope objective) and temporal (~ 500 μs) resolution. Furthermore, the quality of the signals obtained is sufficient to allow for recording of spontaneous synchronized activity such as epileptiform activity induced by the potassium channel blocker 4-aminopyridine. The nature of the signals obtained by these indicators recorded from guinea pig hippocampal slices and some applications of this technique are discussed.
AB - Optical recording techniques provide a constantly evolving and increasingly powerful set of tools for investigations of cellular physiology. These techniques rely on the use of optical indicators, molecules that change their optical properties depending on the cellular parameter of interest. In this paper we discuss some of the general considerations involved in recording optical signals from multiple indicators. specifically, we describe a technique for simultaneously recording transients of membrane potential and intracellular calcium concentration, two parameters that have a very complex interrelationship in neuronal functioning. This technique relies on the use of two fluorescent indicators (the voltage-sensitive dye RH-414 and the calcium-sensitive dye Calcium Orange) that have overlapping excitation spectra but separable emission spectra. This fact, in combination with the use of fast, spatially resolving photodetectors (10 x 10-element photodiode matrices), allows for truly simultaneous recording of these transients from brain slices with high spatial (~ 200 x 200 μm with a 10 x microscope objective) and temporal (~ 500 μs) resolution. Furthermore, the quality of the signals obtained is sufficient to allow for recording of spontaneous synchronized activity such as epileptiform activity induced by the potassium channel blocker 4-aminopyridine. The nature of the signals obtained by these indicators recorded from guinea pig hippocampal slices and some applications of this technique are discussed.
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U2 - 10.1006/meth.1999.0773
DO - 10.1006/meth.1999.0773
M3 - Article
C2 - 10356352
AN - SCOPUS:0032874263
SN - 1046-2023
VL - 18
SP - 204
EP - 214
JO - Methods: A Companion to Methods in Enzymology
JF - Methods: A Companion to Methods in Enzymology
IS - 2
ER -