Angiotensin-II Modulates GABAergic Neurotransmission in the Mouse Substantia Nigra

Figures

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  Figure 1.. Electrophysiological characteristics of GABAergic and dopaminergic neurons in the mouse substantia nigra. A, SNr GABAergic neurons have a narrower (<1.5 ms) AP width than SNc dopaminergic neurons (>3 ms). B, Representative sustained high-frequency firing of SNr GABAergic neurons recorded in whole-cell (top) and cell-attached configuration (bottom). C, SNr GABAergic neurons, in contrast to SNc dopaminergic neurons, show little to no Ih in response to a series of hyperpolarizing pre pulses (–70 to –140 mV). D, Representative low-frequency, slow, and irregular pacemaker-like firing in SNc dopaminergic neurons.

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  Figure 2.. Ang-II decreases SNr GABAergic projection neuron spike firing. A, Evoked APs in response to 150-pA current injection. B, Individual scatter plot of evoked spike frequency in the SNr GABAergic projection neurons in response to 50-, 100-, and 150-pA current injections. Ang-II significantly decreased evoked firing in all stimulus levels combined, 50–150 pA (***p < 0.001, two-way repeated-measures ANOVA). In 10 out of 17 cells, washout partially reversed the effect of Ang-II. C, Ang-II increased the SNr GABAergic interspike interval (ISI; **p = 0.005, mixed effect analysis) and (D) increased the irregularity of firing as quantified by the ISI SD (*p = 0.01, mixed effect analysis). E, F, The AT1-R-specific blocker losartan (1 μm) abolished the suppression of evoked SNr GABAergic spike firing by Ang-II.

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  Figure 3.. Ang-II slows the evoked AP kinetics of SNr GABAergic neurons. A, Representative AP waveforms of SNr projection neurons showing that Ang-II reversibly slows AP kinetics. B, Ang-II increased the AP half-width in 11 out of 17 SNr GABAergic neurons (*p = 0.010, mixed-effect analysis). C, Ang-II slowed the rise of APs in SNr GABAergic neurons (**p = 0.005, repeated measures one-way ANOVA). D, Ang-II slowed the decay of APs in SNr GABAergic neurons (**p = 0.004, repeated measure one-way ANOVA). Pooled data are a grouped data of all stimulus levels (50, 100, and 150 pA).

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  Figure 4.. Disparate effect of Ang-II on SN GABAergic and dopaminergic neurons. A, Losartan blocks Ang-II-mediated increase in APD in SNr GABAergic neurons; losartan washout with continued Ang-II perfusion prolonged the APD. B, Summary data showing that losartan (1 μm) blocks Ang-II-mediated increase in AP half-width and decay. Data from all stimulus levels (50, 100, and 150 pA) were pooled for each group. C, Ang-II increased the duration of SNr GABAergic neuron spontaneous APs (*p = 0.04, AP half-width; *p = 0.015, AP decay; repeated measures one-way ANOVA). D, Unlike SNr GABAergic neurons, Ang-II had no noticeable effect on the AP kinetics of SNc dopaminergic neurons.

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  Figure 5.. GABAAR blockade prevents Ang-II-mediated suppression of evoked spike firing in the SNr GABAergic projection neurons. A, Overlaid spike waveforms for control conditions, in the presence of the GABAAR antagonist picrotoxin (1 μm), and in the presence of picrotoxin plus Ang-II. B, Summary plot showing evoked spike firing of SNr GABAergic neurons under control conditions, in the presence of picrotoxin (1 μm), and in the presence of picrotoxin plus Ang-II. C, GABAAR blockade with picrotoxin attenuates Ang-II-mediated increases in SNr GABAergic neuron APDs.

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  Figure 6.. Ang-II increases SNr GABAergic neuron spontaneous PSCs. A, At a holding potential of –65 mV (junction potential corrected), spontaneous PSCs were recorded with a high chloride (57.5 nm) internal pipette solution before and after 5 min of Ang-II (500 nm) exposure. B, Ang-II did not markedly change the average PSC amplitude (p = 0.211, paired Student’s t test) and frequency (p = 0.466, paired Student’s t test) of spontaneous postsynaptic Cl– currents. However, Ang-II significantly increased the observed incidence of high-amplitude spontaneous PSCs (right; *p = 0.012; paired Student’s t test). High-amplitude spontaneous PSC, demarcated by the dashed line, were defined a priori as currents with an amplitude more than or equal to three times the mean current amplitude under control conditions.

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  Figure 7.. Ang-II increases picrotoxin-sensitive spontaneous MIPSC in SNr GABAergic neurons. Spontaneous mIPSC recorded from SNr GABAergic and SNc dopaminergic neurons in the presence of excitatory synaptic blockers (DNQX, MK-801; 1 μm) and TTX (500 nm). A, Representative traces showing Ang-II-mediated enhancement of high-amplitude postsynaptic GABAAR currents. High-amplitude spontaneous PSCs, demarcated by the dashed line, were defined a priori as currents with an amplitude more than or equal to three times the mean current amplitude recorded in the presence of excitatory synaptic blockers and TTX. B, Summary data showing Ang-II-mediated ∼2.5-fold increase in high-amplitude mIPSC (*p = 0.019, one-way repeated measures ANOVA), but not in average mIPSC amplitude and frequency. C, In contrast to SNr GABAergic neurons, Ang-II had no detectable effect on spontaneous mIPSC in SNc dopaminergic neurons.

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  Figure 8.. Ang-II modulates SN GABAergic neurotransmission. A, Light-evoked EPSP (bursts) in SNr GABAergic neurons in response to a 100-ms-long light pulse. B, Ang-II decreased light-evoked intraburst spikes in four out of six SNr GABAergic neurons (*p = 0.024, paired Student’s t test); intraburst spikes shown are enlarged from panel A. C, Summary data showing the effect of Ang-II on intraburst spikes (EPSP) in SNr GABAergic neurons in response to 100-ms-long light pulse. In four out of six SNr GABAergic neurons, Ang-II decreased intraburst spikes by ∼25%. D, IPSP recorded from SNc dopaminergic neurons in response to photostimulation of SNr GABAergic neurons, as shown by downward deflection of membrane potential (blue stars). Ang-II enhanced light-evoked inhibitory input onto SNc dopaminergic neurons as shown by an increase in the amplitude of light-evoked IPSP (E) and IPSC (F, G; *p = 0.049, paired Student’s t test).