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GABAergic and glycinergic inhibitory synaptic transmission in the ventral cochlear nucleus studied in VGAT channelrhodopsin-2 mice.

Xie R, Manis PB - Front Neural Circuits (2014)

Bottom Line: During prolonged stimulation, the ratio of steady state vs. peak IPSC amplitude was significantly lower for glycinergic IPSCs.In the absence of receptor blockers, repetitive light stimulation was only able to effectively evoke IPSCs up to 20 Hz in both bushy and multipolar neurons.We conclude that local GABAergic release within the VCN can differentially influence bushy and multipolar cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology/Head and Neck Surgery, University of North Carolina at Chapel Hill Chapel Hill, NC, USA.

ABSTRACT
Both glycine and GABA mediate inhibitory synaptic transmission in the ventral cochlear nucleus (VCN). In mice, the time course of glycinergic inhibition is slow in bushy cells and fast in multipolar (stellate) cells, and is proposed to contribute to the processing of temporal cues in both cell types. Much less is known about GABAergic synaptic transmission in this circuit. Electrical stimulation of the auditory nerve or the tuberculoventral pathway evokes little GABAergic synaptic current in brain slice preparations, and spontaneous GABAergic miniature synaptic currents occur infrequently. To investigate synaptic currents carried by GABA receptors in bushy and multipolar cells, we used transgenic mice in which channelrhodopsin-2 and EYFP is driven by the vesicular GABA transporter (VGAT-ChR2-EYFP) and is expressed in both GABAergic and glycinergic neurons. Light stimulation evoked action potentials in EYFP-expressing presynaptic cells, and evoked inhibitory postsynaptic potentials (IPSPs) in non-expressing bushy and planar multipolar cells. Less than 10% of the IPSP amplitude in bushy cells arose from GABAergic synapses, whereas 40% of the IPSP in multipolar neurons was GABAergic. In voltage clamp, glycinergic IPSCs were significantly slower in bushy neurons than in multipolar neurons, whereas there was little difference in the kinetics of the GABAergic IPSCs between two cell types. During prolonged stimulation, the ratio of steady state vs. peak IPSC amplitude was significantly lower for glycinergic IPSCs. Surprisingly, the reversal potentials of GABAergic IPSCs were negative to those of glycinergic IPSCs in both bushy and multipolar neurons. In the absence of receptor blockers, repetitive light stimulation was only able to effectively evoke IPSCs up to 20 Hz in both bushy and multipolar neurons. We conclude that local GABAergic release within the VCN can differentially influence bushy and multipolar cells.

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Kinetics of GABAergic and glycinergic IPSCs in bushy and multipolar neurons. (A) A 2 ms light pulse (blue bar on top) evokes IPSCs in a bushy neuron. The IPSC is mostly blocked by strychnine (stry) and is fully blocked by a combination of strychnine and SR95531 (Stry + SR). Inset: magnified IPSC trace after strychnine block. The weighted decay time constants of the IPSCs are obtained by fitting the IPSC decay with double exponential functions (black curves) under both control and stry conditions. (B) 1 ms light pulse evokes IPSCs in a multipolar neuron. Plots are organized the same as in (A). Decay time constants of IPSCs are obtained by fitting the IPSC decay with single exponential functions (black curves). Traces in both (A) and (B) are averages of 10 trials. (C) Comparison of the light evoked IPSC amplitudes between bushy and multipolar neurons including the control IPSC amplitude (ctrl), glycinergic IPSC component (Gly), and GABAergic IPSC component (GABA). Abcissa: B: bushy neurons; M: multipolar neurons. (D) Comparison of the eIPSC decay time constants. * p < 0.05; ** p < 0.01. Data is plotted as mean ± S.D.
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Figure 3: Kinetics of GABAergic and glycinergic IPSCs in bushy and multipolar neurons. (A) A 2 ms light pulse (blue bar on top) evokes IPSCs in a bushy neuron. The IPSC is mostly blocked by strychnine (stry) and is fully blocked by a combination of strychnine and SR95531 (Stry + SR). Inset: magnified IPSC trace after strychnine block. The weighted decay time constants of the IPSCs are obtained by fitting the IPSC decay with double exponential functions (black curves) under both control and stry conditions. (B) 1 ms light pulse evokes IPSCs in a multipolar neuron. Plots are organized the same as in (A). Decay time constants of IPSCs are obtained by fitting the IPSC decay with single exponential functions (black curves). Traces in both (A) and (B) are averages of 10 trials. (C) Comparison of the light evoked IPSC amplitudes between bushy and multipolar neurons including the control IPSC amplitude (ctrl), glycinergic IPSC component (Gly), and GABAergic IPSC component (GABA). Abcissa: B: bushy neurons; M: multipolar neurons. (D) Comparison of the eIPSC decay time constants. * p < 0.05; ** p < 0.01. Data is plotted as mean ± S.D.

Mentions: In a separate population of cells, we investigated the kinetics of the light evoked synaptic currents under voltage clamp (Figure 3). Recordings were made using Cs-based electrode solution (8 mM Cl−) with 3 mM QX-314 to block potassium and sodium channels and improve clamp quality. Cells were held at +42 mV so that the IPSCs were large and outward. Light pulses of 1 or 2 ms were used to evoke repeatable single spikes in presynaptic inputs (Figures 1D,E), to help minimize the possibility that the kinetics of evoked IPSCs were contaminated by multiple synaptic events.


GABAergic and glycinergic inhibitory synaptic transmission in the ventral cochlear nucleus studied in VGAT channelrhodopsin-2 mice.

Xie R, Manis PB - Front Neural Circuits (2014)

Kinetics of GABAergic and glycinergic IPSCs in bushy and multipolar neurons. (A) A 2 ms light pulse (blue bar on top) evokes IPSCs in a bushy neuron. The IPSC is mostly blocked by strychnine (stry) and is fully blocked by a combination of strychnine and SR95531 (Stry + SR). Inset: magnified IPSC trace after strychnine block. The weighted decay time constants of the IPSCs are obtained by fitting the IPSC decay with double exponential functions (black curves) under both control and stry conditions. (B) 1 ms light pulse evokes IPSCs in a multipolar neuron. Plots are organized the same as in (A). Decay time constants of IPSCs are obtained by fitting the IPSC decay with single exponential functions (black curves). Traces in both (A) and (B) are averages of 10 trials. (C) Comparison of the light evoked IPSC amplitudes between bushy and multipolar neurons including the control IPSC amplitude (ctrl), glycinergic IPSC component (Gly), and GABAergic IPSC component (GABA). Abcissa: B: bushy neurons; M: multipolar neurons. (D) Comparison of the eIPSC decay time constants. * p < 0.05; ** p < 0.01. Data is plotted as mean ± S.D.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109614&req=5

Figure 3: Kinetics of GABAergic and glycinergic IPSCs in bushy and multipolar neurons. (A) A 2 ms light pulse (blue bar on top) evokes IPSCs in a bushy neuron. The IPSC is mostly blocked by strychnine (stry) and is fully blocked by a combination of strychnine and SR95531 (Stry + SR). Inset: magnified IPSC trace after strychnine block. The weighted decay time constants of the IPSCs are obtained by fitting the IPSC decay with double exponential functions (black curves) under both control and stry conditions. (B) 1 ms light pulse evokes IPSCs in a multipolar neuron. Plots are organized the same as in (A). Decay time constants of IPSCs are obtained by fitting the IPSC decay with single exponential functions (black curves). Traces in both (A) and (B) are averages of 10 trials. (C) Comparison of the light evoked IPSC amplitudes between bushy and multipolar neurons including the control IPSC amplitude (ctrl), glycinergic IPSC component (Gly), and GABAergic IPSC component (GABA). Abcissa: B: bushy neurons; M: multipolar neurons. (D) Comparison of the eIPSC decay time constants. * p < 0.05; ** p < 0.01. Data is plotted as mean ± S.D.
Mentions: In a separate population of cells, we investigated the kinetics of the light evoked synaptic currents under voltage clamp (Figure 3). Recordings were made using Cs-based electrode solution (8 mM Cl−) with 3 mM QX-314 to block potassium and sodium channels and improve clamp quality. Cells were held at +42 mV so that the IPSCs were large and outward. Light pulses of 1 or 2 ms were used to evoke repeatable single spikes in presynaptic inputs (Figures 1D,E), to help minimize the possibility that the kinetics of evoked IPSCs were contaminated by multiple synaptic events.

Bottom Line: During prolonged stimulation, the ratio of steady state vs. peak IPSC amplitude was significantly lower for glycinergic IPSCs.In the absence of receptor blockers, repetitive light stimulation was only able to effectively evoke IPSCs up to 20 Hz in both bushy and multipolar neurons.We conclude that local GABAergic release within the VCN can differentially influence bushy and multipolar cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology/Head and Neck Surgery, University of North Carolina at Chapel Hill Chapel Hill, NC, USA.

ABSTRACT
Both glycine and GABA mediate inhibitory synaptic transmission in the ventral cochlear nucleus (VCN). In mice, the time course of glycinergic inhibition is slow in bushy cells and fast in multipolar (stellate) cells, and is proposed to contribute to the processing of temporal cues in both cell types. Much less is known about GABAergic synaptic transmission in this circuit. Electrical stimulation of the auditory nerve or the tuberculoventral pathway evokes little GABAergic synaptic current in brain slice preparations, and spontaneous GABAergic miniature synaptic currents occur infrequently. To investigate synaptic currents carried by GABA receptors in bushy and multipolar cells, we used transgenic mice in which channelrhodopsin-2 and EYFP is driven by the vesicular GABA transporter (VGAT-ChR2-EYFP) and is expressed in both GABAergic and glycinergic neurons. Light stimulation evoked action potentials in EYFP-expressing presynaptic cells, and evoked inhibitory postsynaptic potentials (IPSPs) in non-expressing bushy and planar multipolar cells. Less than 10% of the IPSP amplitude in bushy cells arose from GABAergic synapses, whereas 40% of the IPSP in multipolar neurons was GABAergic. In voltage clamp, glycinergic IPSCs were significantly slower in bushy neurons than in multipolar neurons, whereas there was little difference in the kinetics of the GABAergic IPSCs between two cell types. During prolonged stimulation, the ratio of steady state vs. peak IPSC amplitude was significantly lower for glycinergic IPSCs. Surprisingly, the reversal potentials of GABAergic IPSCs were negative to those of glycinergic IPSCs in both bushy and multipolar neurons. In the absence of receptor blockers, repetitive light stimulation was only able to effectively evoke IPSCs up to 20 Hz in both bushy and multipolar neurons. We conclude that local GABAergic release within the VCN can differentially influence bushy and multipolar cells.

Show MeSH
Related in: MedlinePlus