Article Figures & Data
Figures
- Fig. 1.
Cellular localization of GluR6 mRNA in the dorsal striatum. Emulsion autoradiograms from double in situhybridization experiments showing the colocalization of GluR6 mRNA (radioactive probe, silver grains) with substance P (SP) or enkephalin (ENK) mRNA (digoxigenin-labeled probe, brown labeling) in the dorsal striatum of a wild-type mouse. The majority of substance P- and enkephalin-containing neurons coexpress GluR6 mRNA.
- Fig. 2.
Functional GluR6 KARs in the mouse striatum.A, Inward current evoked by 500 nm domoate in a neuron recorded from the dorsal striatum of a wild-type mouse, in the presence of TTX (500 nm), dl-AP5 (50 μm), and picrotoxin (100 μm). In GluR6-deficient mice (GluR6−/−), higher concentrations of domoate are needed to activate an inward current in striatal neurons.B, Dose–response curves for low concentrations of domoate in the striatum of wild-type and GluR6-deficient mice. The amplitude of the inward current evoked by each concentration of domoate is represented in the y-axis (mean ± SEM,n = 3–11).
- Fig. 3.
Inward currents activated by low concentrations of domoate are mediated by KARs. A, In wild-type mice (left traces), the inward current activated by domoate (500 nm) was unaffected by low concentrations of NBQX (1 μm). In GluR6-deficient mice (right traces), the inward current activated by a larger concentration of domoate was totally blocked by NBQX (1 μm).B, Mean amplitude of inward currents activated by domoate (500 nm, 1, 2, or 5 μm) alone or in the presence of NBQX (1 μm) or NBQX (1 μm) + GYKI 53655 (50 μm) (mean ± SEM,n = 3–11).
- Fig. 4.
Domoate in the presence of NBQX (1 μm) and dl-AP5 (50 μm) causes depolarization and transiently triggers action potential firing in striatal neurons. A1, Cell-attached recording of a medium-sized striatal neuron in the presence of NBQX (1 μm). In control conditions, this neuron did not discharge spontaneous action potentials. Domoate (500 nm, 2 min) triggered action potential firing (up to a frequency of 11 Hz) that stopped before the end of agonist application. A2,Histogram of the average increase in spike frequency induced by domoate (n = 8 cells). B, Whole-cell current-clamp recording of a medium-size striatal neuron. An intracellular solution containing K-Gluconate (120 mm) and KCl (20 mm) was used. In the presence of NBQX (1 μm), domoate depolarizes the neuronal membrane by >30 mV and triggers action potential discharge. Despite sustained depolarization, action potential firing stops before the end of domoate application.
- Fig. 5.
KARs do not participate in EPSCs.A, In wild-type mice, EPSPs evoked by single pulse intrastriatal stimulation in the presence of bicuculline (20 μm) and dl-AP5 (100 μm) were largely blocked by the AMPA receptor antagonist LY 303070 (25 μm) in wild-type mice (left traces) as well as in GluR6 −/− mice (right traces).B, CNQX (50 μm) does not block small LY 303070-resistant synaptic currents. Traces of evoked synaptic currents recorded in the presence of LY 303070 and in the presence of CNQX (50 μm) are superimposed, showing no difference in PSC amplitude or time course in the presence of these two antagonists. In the bottom traces, train stimulation (five pulses, 5 msec interval) evoked slowly decaying postsynaptic currents. No difference in the pharmacology of EPSCs was observed in GluR6-deficient mice (right traces) as compared to wild-type mice. Average of 20 sweeps for each trace in A andB.
- Fig. 6.
Activation of KARs increases the frequency of spontaneous IPSCs of small amplitude in striatal neurons.A, Plot of the frequency of spontaneous IPSCs recorded in the presence of GYKI 53655 (50 μm) anddl-AP5 (50 μm) in control conditions and in the presence of domoate 200 nm (2 min). B,Traces of recording shown at the same time scale as the histogram inA. Top trace, Trace from which plotA was drawn. Bottom trace, Recording in the presence of picrotoxin (PTX, 100 μm), showing the blockade of spontaneous PSCs by this GABAA receptor antagonist. The white box represents the time during which domoate was applied. C, Traces of spontaneous IPSCs shown at a larger time scale before (top trace) and during (bottom trace) application of domoate.D, Amplitude histograms of spontaneous IPSCs recorded during the same time period (50 sec), in control conditions, and in the presence of domoate (200 nm). No major change in the distribution of IPSC amplitudes was observed except for an increase in the number of events in each bin during domoate application.A–D show data from the same neuron.
- Fig. 7.
Domoate depresses evoked IPSCs. A,IPSCs evoked by minimal intrastriatal stimulation were reversibly depressed by bath application of domoate (200 nm, 2 min) in wild-type mice but not in GluR6-deficient mice. For each set of traces, five sweeps are superimposed. Top trace, Example of a striatal neuron recorded in a wild-type mouse for which no failure was observed and where the average amplitude of the evoked IPSCs was decreased by domoate. The dotted line represents the level of the control holding current. Bottom traces,Example of a neuron recorded in a GluR6-deficient mouse; domoate (200 nm) did not depress the amplitude of the IPSC and did not increase the number of failures. B, Time course of the effect of domoate (200 nm for 2 min) on the IPSC amplitude in wild-type mice (n = 13) and in GluR6-deficient mice (n = 7). Each point represents the mean (± SEM) IPSC amplitude expressed as percentage of baseline amplitude calculated, for each cell, before domoate application.
- Fig. 8.
Effect of domoate on mIPSC frequency and amplitude in wild-type mice. Miniature IPSCs were recorded in the presence of TTX (400 nm), NBQX (1 μm), and APV (50 μm). A, Plot of mIPSC frequency versus time. The dotted line represents the average control value. Superimposed on the plot is the inward current evoked by activation of somatodendritic KARs present in the recorded neuron. Concomitant with this inward current, domoate (500 nm for 2 min) causes a moderate decrease in total mIPSC frequency.B–D, In the same neuron, a closer inspection of the effects of domoate on mIPSCs indicates that domoate mainly affects large-amplitude mIPSCs. B, Each mIPSC is represented by a vertical bar proportional to its amplitude to allow visualization of the activity over long recording periods (2 min).C, Amplitude distribution histogram of mIPSCs recorded over the same time period (3 min) in control conditions (open bars) and after application of domoate (filled bars). D, Cumulative amplitude histogram corresponding to the distributions shown in C for control conditions (dashed line) and after application of domoate (solid line).
- Fig. 9.
A2A receptor antagonists block the action of domoate on evoked IPSCs. A, IPSCs were evoked by intrastriatal stimulation (at a rate 0.2 Hz) in the presence of NBQX (1 μm). Top traces, Domoate (500 nm for 2 min) reversibly decreased evoked IPSC amplitude.Bottom traces, In the same cell, perfusion of the slice with the selective A2A antagonist ZM 241385 (1 μm) prevented the action of domoate on evoked IPSC amplitude. The dotted line represents the level of the control inward current. B, For the same experiment, plot of the amplitude of evoked IPSCs as a function of time. Domoate is applied at the time indicated by the open horizontal bars. As indicated by the bottom horizontal bar, ZM 241385 (1 μm) is perfused several minutes before the second application of domoate. C, Histogram of the average (± SEM) inhibition by domoate (500 nm for 2 min) of evoked IPSC amplitude in control conditions (n = 18), in the presence of ZM 241385 (1 μm) (n = 7), and in the presence of SCH-58261 (1 μm) (n = 6). For both antagonists, the difference in inhibition versus domoate alone was significant withp < 0.001.
