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Stimulus-evoked high frequency oscillations are present in neuronal networks on microelectrode arrays.

Hales CM, Zeller-Townson R, Newman JP, Shoemaker JT, Killian NJ, Potter SM - Front Neural Circuits (2012)

Bottom Line: As with in vivo studies, activity is isolated to a single electrode, however, the MEA provides improved spatial resolution with no spread of the oscillation to adjacent electrodes 200 μm away.Chelating calcium with ethylene glycol tetraacetic acid (EGTA) causes a temporal prolongation of the oscillation.Gap junctions may play a significant role in maintaining the oscillation given the inhibitory effect of carbenoxolone, the propagating effect of reduced calcium conditions and the isolated nature of the activity as demonstrated in previous studies.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Center for Neurodegenerative Diseases, Emory University, Atlanta GA, USA.

ABSTRACT
Pathological high frequency oscillations (250-600 Hz) are present in the brains of epileptic animals and humans. The etiology of these oscillations and how they contribute to the diseased state remains unclear. This work identifies the presence of microstimulation-evoked high frequency oscillations (250-400 Hz) in dissociated neuronal networks cultured on microelectrode arrays (MEAs). Oscillations are more apparent with higher stimulus voltages. As with in vivo studies, activity is isolated to a single electrode, however, the MEA provides improved spatial resolution with no spread of the oscillation to adjacent electrodes 200 μm away. Oscillations develop across four weeks in vitro. Oscillations still occur in the presence of tetrodotoxin and synaptic blockers, and they cause no apparent disruption in the ability of oscillation-presenting electrodes to elicit directly evoked action potentials (dAPs) or promote the spread of synaptic activity throughout the culture. Chelating calcium with ethylene glycol tetraacetic acid (EGTA) causes a temporal prolongation of the oscillation. Finally, carbenoxolone significantly reduces or eliminates the high frequency oscillations. Gap junctions may play a significant role in maintaining the oscillation given the inhibitory effect of carbenoxolone, the propagating effect of reduced calcium conditions and the isolated nature of the activity as demonstrated in previous studies. This is the first demonstration of stimulus-evoked high frequency oscillations in dissociated cultures. Unlike current models that rely on complex in vivo recording conditions, this work presents a simple controllable model in neuronal cultures on MEAs to further investigate how the oscillations occur at the molecular level and how they may contribute to the pathophysiology of disease.

No MeSH data available.


Related in: MedlinePlus

Effect of carbenoxolone (CBX) on the stimulus-evoked oscillation. (A) Three seconds of SALPA filtered trace from two electrodes (hippocampal culture, age six weeks) showing baseline, with 150 μM CBX, and following washout with 0.5 V stimulus (large black arrowhead). TTX 1 μM was present in the medium. The stimulus-evoked oscillation returned within 10–15 min following washout. The oscillation was completely abolished in electrode 2–1 (column-row) and significantly reduced in electrode 3–1. (B) Average power spectra from three cultures (age 5–7 weeks) and 106 electrodes in the presence of TTX 1 μM and (C) following the addition of CBX 150 μM.
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Figure 7: Effect of carbenoxolone (CBX) on the stimulus-evoked oscillation. (A) Three seconds of SALPA filtered trace from two electrodes (hippocampal culture, age six weeks) showing baseline, with 150 μM CBX, and following washout with 0.5 V stimulus (large black arrowhead). TTX 1 μM was present in the medium. The stimulus-evoked oscillation returned within 10–15 min following washout. The oscillation was completely abolished in electrode 2–1 (column-row) and significantly reduced in electrode 3–1. (B) Average power spectra from three cultures (age 5–7 weeks) and 106 electrodes in the presence of TTX 1 μM and (C) following the addition of CBX 150 μM.

Mentions: Previous work has suggested that ripples around 200 Hz may be due to direct electrical coupling (Draguhn et al., 1998; Traub et al., 2002; Roopun et al., 2010). Modeling data by the same group suggested that these types of junctions are theoretically capable of producing 200 Hz oscillations in local neuron populations (Draguhn et al., 1998). The frequency of fast ripples is likely greater than what is physiologically possible for an action potential train from a single neuron. Multi-unit action potentials are possible, however, one would expect spread of the oscillation to other electrodes in the MEA when there is no pharmacological blockade. This spread does was not observed (Figures 4 and 6). Pharmacological blockade of voltage-dependent sodium channels with tetrodotoxin failed to quell the stimulus-evoked high frequency oscillation (Figure 1B). Direct synaptic blockade also had no effect on the stimulus-evoked oscillation (Figure 1C). The addition of carbenoxolone (CBX, 150 μM), a putative gap junction blocker, either reduced the amplitude and duration of the stimulus-evoked oscillation or completely abolished the oscillation (Figure 7A). Average power spectra demonstrated a near complete loss of the oscillation when comparing cultures containing 1 μM TTX (Figure 7B) to cultures containing 1 μM TTX plus 150 μM CBX (Figure 7C). CBX reduced the oscillation amplitude from an average 25.63 ± 7.97 μV to 16.05 ± 4.87 μV with a p-value of 2.26 × 10−23 while the oscillation duration was reduced from 2.89 ± 2.50 s to 0.73 ± 0.87 s with a p-value of 1.97 × 10−16 (Table 1). The oscillation returned to baseline with CBX washout.


Stimulus-evoked high frequency oscillations are present in neuronal networks on microelectrode arrays.

Hales CM, Zeller-Townson R, Newman JP, Shoemaker JT, Killian NJ, Potter SM - Front Neural Circuits (2012)

Effect of carbenoxolone (CBX) on the stimulus-evoked oscillation. (A) Three seconds of SALPA filtered trace from two electrodes (hippocampal culture, age six weeks) showing baseline, with 150 μM CBX, and following washout with 0.5 V stimulus (large black arrowhead). TTX 1 μM was present in the medium. The stimulus-evoked oscillation returned within 10–15 min following washout. The oscillation was completely abolished in electrode 2–1 (column-row) and significantly reduced in electrode 3–1. (B) Average power spectra from three cultures (age 5–7 weeks) and 106 electrodes in the presence of TTX 1 μM and (C) following the addition of CBX 150 μM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3351760&req=5

Figure 7: Effect of carbenoxolone (CBX) on the stimulus-evoked oscillation. (A) Three seconds of SALPA filtered trace from two electrodes (hippocampal culture, age six weeks) showing baseline, with 150 μM CBX, and following washout with 0.5 V stimulus (large black arrowhead). TTX 1 μM was present in the medium. The stimulus-evoked oscillation returned within 10–15 min following washout. The oscillation was completely abolished in electrode 2–1 (column-row) and significantly reduced in electrode 3–1. (B) Average power spectra from three cultures (age 5–7 weeks) and 106 electrodes in the presence of TTX 1 μM and (C) following the addition of CBX 150 μM.
Mentions: Previous work has suggested that ripples around 200 Hz may be due to direct electrical coupling (Draguhn et al., 1998; Traub et al., 2002; Roopun et al., 2010). Modeling data by the same group suggested that these types of junctions are theoretically capable of producing 200 Hz oscillations in local neuron populations (Draguhn et al., 1998). The frequency of fast ripples is likely greater than what is physiologically possible for an action potential train from a single neuron. Multi-unit action potentials are possible, however, one would expect spread of the oscillation to other electrodes in the MEA when there is no pharmacological blockade. This spread does was not observed (Figures 4 and 6). Pharmacological blockade of voltage-dependent sodium channels with tetrodotoxin failed to quell the stimulus-evoked high frequency oscillation (Figure 1B). Direct synaptic blockade also had no effect on the stimulus-evoked oscillation (Figure 1C). The addition of carbenoxolone (CBX, 150 μM), a putative gap junction blocker, either reduced the amplitude and duration of the stimulus-evoked oscillation or completely abolished the oscillation (Figure 7A). Average power spectra demonstrated a near complete loss of the oscillation when comparing cultures containing 1 μM TTX (Figure 7B) to cultures containing 1 μM TTX plus 150 μM CBX (Figure 7C). CBX reduced the oscillation amplitude from an average 25.63 ± 7.97 μV to 16.05 ± 4.87 μV with a p-value of 2.26 × 10−23 while the oscillation duration was reduced from 2.89 ± 2.50 s to 0.73 ± 0.87 s with a p-value of 1.97 × 10−16 (Table 1). The oscillation returned to baseline with CBX washout.

Bottom Line: As with in vivo studies, activity is isolated to a single electrode, however, the MEA provides improved spatial resolution with no spread of the oscillation to adjacent electrodes 200 μm away.Chelating calcium with ethylene glycol tetraacetic acid (EGTA) causes a temporal prolongation of the oscillation.Gap junctions may play a significant role in maintaining the oscillation given the inhibitory effect of carbenoxolone, the propagating effect of reduced calcium conditions and the isolated nature of the activity as demonstrated in previous studies.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, Center for Neurodegenerative Diseases, Emory University, Atlanta GA, USA.

ABSTRACT
Pathological high frequency oscillations (250-600 Hz) are present in the brains of epileptic animals and humans. The etiology of these oscillations and how they contribute to the diseased state remains unclear. This work identifies the presence of microstimulation-evoked high frequency oscillations (250-400 Hz) in dissociated neuronal networks cultured on microelectrode arrays (MEAs). Oscillations are more apparent with higher stimulus voltages. As with in vivo studies, activity is isolated to a single electrode, however, the MEA provides improved spatial resolution with no spread of the oscillation to adjacent electrodes 200 μm away. Oscillations develop across four weeks in vitro. Oscillations still occur in the presence of tetrodotoxin and synaptic blockers, and they cause no apparent disruption in the ability of oscillation-presenting electrodes to elicit directly evoked action potentials (dAPs) or promote the spread of synaptic activity throughout the culture. Chelating calcium with ethylene glycol tetraacetic acid (EGTA) causes a temporal prolongation of the oscillation. Finally, carbenoxolone significantly reduces or eliminates the high frequency oscillations. Gap junctions may play a significant role in maintaining the oscillation given the inhibitory effect of carbenoxolone, the propagating effect of reduced calcium conditions and the isolated nature of the activity as demonstrated in previous studies. This is the first demonstration of stimulus-evoked high frequency oscillations in dissociated cultures. Unlike current models that rely on complex in vivo recording conditions, this work presents a simple controllable model in neuronal cultures on MEAs to further investigate how the oscillations occur at the molecular level and how they may contribute to the pathophysiology of disease.

No MeSH data available.


Related in: MedlinePlus