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Neurochemical measurements in the zebrafish brain.

Jones LJ, McCutcheon JE, Young AM, Norton WH - Front Behav Neurosci (2015)

Bottom Line: In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon.We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain.This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Psychology and Behaviour, University of Leicester Leicester, UK.

ABSTRACT
The zebrafish is an ideal model organism for behavioral genetics and neuroscience. The high conservation of genes and neurotransmitter pathways between zebrafish and other vertebrates permits the translation of research between species. Zebrafish behavior can be studied at both larval and adult stages and recent research has begun to establish zebrafish models for human disease. Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that permits the detection of neurotransmitter release and reuptake. In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon. We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain. This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain.

No MeSH data available.


Comparison of electrically-stimulated release of analytes in the telencephalon. (A) Schematic representation showing lateral view of zebrafish telencephalon. The red crosses show the position of tips of the stimulating electrode and the black cross shows the position of the recording electrode used in these experiments. (B,C) Voltammograms and current vs. time plots from eight repeated stimulations in one slice using optimal parameters [20 pulses (pulse width of 4 ms) and a voltage of 500 μA, 60 Hz]. (D–F) Color plot, voltammogram, and current vs. time plot from a single representative experiment using optimal stimulation. (G–I) Color plot, voltammogram and current vs. time plot from a single representative experiment using low intensity stimulation parameters [20 pulses (pulse width of 4 ms), and a voltage of 300 μA, 60 Hz]. (J–L) Color plot and voltammogram and current vs. time plots from a single representative experiment using high intensity stimulation parameters [60 pulses (pulse width of 4 ms), voltage of 1 mA, 60 Hz]. High intensity stimulation led to the release of analytes at two different points in the voltammogram [black and red lines in (L)]. (D,G,J) Dashed lines show the position at which the voltammograms were extracted and black and red circles depict points at which current vs. time plots were taken.
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Figure 4: Comparison of electrically-stimulated release of analytes in the telencephalon. (A) Schematic representation showing lateral view of zebrafish telencephalon. The red crosses show the position of tips of the stimulating electrode and the black cross shows the position of the recording electrode used in these experiments. (B,C) Voltammograms and current vs. time plots from eight repeated stimulations in one slice using optimal parameters [20 pulses (pulse width of 4 ms) and a voltage of 500 μA, 60 Hz]. (D–F) Color plot, voltammogram, and current vs. time plot from a single representative experiment using optimal stimulation. (G–I) Color plot, voltammogram and current vs. time plot from a single representative experiment using low intensity stimulation parameters [20 pulses (pulse width of 4 ms), and a voltage of 300 μA, 60 Hz]. (J–L) Color plot and voltammogram and current vs. time plots from a single representative experiment using high intensity stimulation parameters [60 pulses (pulse width of 4 ms), voltage of 1 mA, 60 Hz]. High intensity stimulation led to the release of analytes at two different points in the voltammogram [black and red lines in (L)]. (D,G,J) Dashed lines show the position at which the voltammograms were extracted and black and red circles depict points at which current vs. time plots were taken.

Mentions: Bath application of high K+ aCSF produced a current vs. time plot with a prolonged release profile that did not return to pre-stimulation baseline levels. In order to clarify whether this was an artifact caused by electrode drift during the long time necessary for complete washout to occur, we used electrical stimulation to evoke the release of analytes (Figure 4A). Electrical stimulation of local terminals using optimal parameters (20 pulses with a pulse width of 4 ms, 60 Hz, 500 μA) resulted in an increase in current on the forward part of the waveform (Figures 4C,F) that rapidly returned to baseline (n = 8 stimulations from a single sagittal section in Figures 4B,C). It also produced a cyclic voltammogram with a shape similar to that obtained using high K+ aCSF suggesting that both types of stimulation evoke the release of a similar mixture of analytes (Figures 4B,D,E). We tested this possibility using the CV match algorithm in TarHeel. Comparison of an example electrical stimulation with an example K+ stimulation gave an r2-value of 0.876, indicating that both types of stimulation evoke similar neurochemical changes in the tissue. However, the oxidation peak at ~ +0.6 V and reduction peak at ~ −0.2 V were much more prominent when using electrical stimulation than in voltammograms obtained using high K+ aCSF. The lowest intensity stimulation that we could use to trigger analyte release in the telencephalon was 20 pulses with a pulse width of 4 ms, 60 Hz, 300 μA. This produced a voltammogram with an oxidation peak at ~ +0.6 V and a smaller reduction peak at ~0.2 V (Figures 4G–I). In contrast to this, high intensity stimulation (1 mA, 60 Hz, 60 pulses, pulse width 4 ms) produced a cyclic voltammogram with a similar shape to that extracted ~30 s after K+ stimulation (Figures 4J,K) with a small oxidation peak occurring on the reverse scan at ~1.1 V and a large reduction peak at ~ −0.4 V). Furthermore, the change in current ~ +0.6 V showed a large decrease similar to the dip in current observed following K+ stimulation (Figure 4L). The prolonged time-course of alterations in current suggested that an artifact such as a change in pH had occurred. This indicates that stimulation parameters are an important consideration when attempting to obtain reproducible measurements of neurotransmitter release that are not masked by pH shifts or electrode drift.


Neurochemical measurements in the zebrafish brain.

Jones LJ, McCutcheon JE, Young AM, Norton WH - Front Behav Neurosci (2015)

Comparison of electrically-stimulated release of analytes in the telencephalon. (A) Schematic representation showing lateral view of zebrafish telencephalon. The red crosses show the position of tips of the stimulating electrode and the black cross shows the position of the recording electrode used in these experiments. (B,C) Voltammograms and current vs. time plots from eight repeated stimulations in one slice using optimal parameters [20 pulses (pulse width of 4 ms) and a voltage of 500 μA, 60 Hz]. (D–F) Color plot, voltammogram, and current vs. time plot from a single representative experiment using optimal stimulation. (G–I) Color plot, voltammogram and current vs. time plot from a single representative experiment using low intensity stimulation parameters [20 pulses (pulse width of 4 ms), and a voltage of 300 μA, 60 Hz]. (J–L) Color plot and voltammogram and current vs. time plots from a single representative experiment using high intensity stimulation parameters [60 pulses (pulse width of 4 ms), voltage of 1 mA, 60 Hz]. High intensity stimulation led to the release of analytes at two different points in the voltammogram [black and red lines in (L)]. (D,G,J) Dashed lines show the position at which the voltammograms were extracted and black and red circles depict points at which current vs. time plots were taken.
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Related In: Results  -  Collection

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Figure 4: Comparison of electrically-stimulated release of analytes in the telencephalon. (A) Schematic representation showing lateral view of zebrafish telencephalon. The red crosses show the position of tips of the stimulating electrode and the black cross shows the position of the recording electrode used in these experiments. (B,C) Voltammograms and current vs. time plots from eight repeated stimulations in one slice using optimal parameters [20 pulses (pulse width of 4 ms) and a voltage of 500 μA, 60 Hz]. (D–F) Color plot, voltammogram, and current vs. time plot from a single representative experiment using optimal stimulation. (G–I) Color plot, voltammogram and current vs. time plot from a single representative experiment using low intensity stimulation parameters [20 pulses (pulse width of 4 ms), and a voltage of 300 μA, 60 Hz]. (J–L) Color plot and voltammogram and current vs. time plots from a single representative experiment using high intensity stimulation parameters [60 pulses (pulse width of 4 ms), voltage of 1 mA, 60 Hz]. High intensity stimulation led to the release of analytes at two different points in the voltammogram [black and red lines in (L)]. (D,G,J) Dashed lines show the position at which the voltammograms were extracted and black and red circles depict points at which current vs. time plots were taken.
Mentions: Bath application of high K+ aCSF produced a current vs. time plot with a prolonged release profile that did not return to pre-stimulation baseline levels. In order to clarify whether this was an artifact caused by electrode drift during the long time necessary for complete washout to occur, we used electrical stimulation to evoke the release of analytes (Figure 4A). Electrical stimulation of local terminals using optimal parameters (20 pulses with a pulse width of 4 ms, 60 Hz, 500 μA) resulted in an increase in current on the forward part of the waveform (Figures 4C,F) that rapidly returned to baseline (n = 8 stimulations from a single sagittal section in Figures 4B,C). It also produced a cyclic voltammogram with a shape similar to that obtained using high K+ aCSF suggesting that both types of stimulation evoke the release of a similar mixture of analytes (Figures 4B,D,E). We tested this possibility using the CV match algorithm in TarHeel. Comparison of an example electrical stimulation with an example K+ stimulation gave an r2-value of 0.876, indicating that both types of stimulation evoke similar neurochemical changes in the tissue. However, the oxidation peak at ~ +0.6 V and reduction peak at ~ −0.2 V were much more prominent when using electrical stimulation than in voltammograms obtained using high K+ aCSF. The lowest intensity stimulation that we could use to trigger analyte release in the telencephalon was 20 pulses with a pulse width of 4 ms, 60 Hz, 300 μA. This produced a voltammogram with an oxidation peak at ~ +0.6 V and a smaller reduction peak at ~0.2 V (Figures 4G–I). In contrast to this, high intensity stimulation (1 mA, 60 Hz, 60 pulses, pulse width 4 ms) produced a cyclic voltammogram with a similar shape to that extracted ~30 s after K+ stimulation (Figures 4J,K) with a small oxidation peak occurring on the reverse scan at ~1.1 V and a large reduction peak at ~ −0.4 V). Furthermore, the change in current ~ +0.6 V showed a large decrease similar to the dip in current observed following K+ stimulation (Figure 4L). The prolonged time-course of alterations in current suggested that an artifact such as a change in pH had occurred. This indicates that stimulation parameters are an important consideration when attempting to obtain reproducible measurements of neurotransmitter release that are not masked by pH shifts or electrode drift.

Bottom Line: In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon.We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain.This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Psychology and Behaviour, University of Leicester Leicester, UK.

ABSTRACT
The zebrafish is an ideal model organism for behavioral genetics and neuroscience. The high conservation of genes and neurotransmitter pathways between zebrafish and other vertebrates permits the translation of research between species. Zebrafish behavior can be studied at both larval and adult stages and recent research has begun to establish zebrafish models for human disease. Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that permits the detection of neurotransmitter release and reuptake. In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon. We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain. This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain.

No MeSH data available.