Two-color, one-photon uncaging of glutamate and GABA.

Matthew B Dalva,Stefan Passlick,John T Williams,Graham C R Ellis-Davies,Matthew T Richers,Paul F Kramer

doi:10.1371/journal.pone.0187732

Matthew B Dalva, Stefan Passlick + Show 4 more

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https://doi.org/10.1371/journal.pone.0187732

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Abstract

Neuronal cells receive a variety of excitatory and inhibitory signals which they process to generate an output signal. In order to study the interaction between excitatory and inhibitory receptors with exogenously applied transmitters in the same preparation, two caging chromophores attached to glutamate and GABA were developed that were selectively photolyzed by different wavelengths of light. This technique has the advantage that the biologically inactive caged compound can be applied at equilibrium prior to the near instantaneous release of the transmitters. This method therefore mimics the kinetics of endogenously released transmitters that is otherwise not possible in brain slice preparations. Repeated photolysis with either of the two wavelengths resulted in GABA- or glutamate-induced activation of both ionotropic and metabotropic receptors to evoke reproducible currents. With these compounds, the interaction between inhibitory and excitatory receptors was examined using whole field photolysis.

Highlights

The interaction between excitatory and inhibitory signals is the basis of neuronal computation
The λmax was shifted to 454 nm and, though the λmin was unchanged in position, its value was decreased by 50% compared to PEG-DEAC450-GABA (Fig 1b)
Previous development of optically tuned DEAC photochemical protecting groups resulted in the creation of DEAC450, with a red shift of the λmax and a pronounced λmin at the short wavelength where traditional near-UV caging chromophores absorb [21,22]

Summary

Introduction

The interaction between excitatory and inhibitory signals is the basis of neuronal computation. Ultra-fast application, which is necessary to generate physiologically relevant responses, is difficult to achieve in brain slices. These techniques do not allow the application of two different agonists, i.e. excitatory and inhibitory agonists, to the same preparation at the same time to study their interaction. The use of two caged compounds that allow wavelength-selective activation of different receptors to study their interaction has not been possible. A handful of two-color uncaging studies of neurons in brain slices have appeared recently[4,5,6], none of these use 1P excitation in both optical channels[7].

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