Striatal microcircuitry and movement disorders.

<p>The basal ganglia network serves to integrate information about context, actions, and outcomes to shape the behavior of an animal based on its past experience. Clinically, the basal ganglia receive the most attention for their role in movement disorders. Recent advances in technology have opened new avenues of research into the structure and function of basal ganglia circuits. One emerging theme is the importance of GABAergic interneurons in coordinating and regulating network function. Here, we discuss evidence that changes in striatal GABAergic microcircuits contribute to basal ganglia dysfunction in several movement disorders. Because interneurons are genetically and neurochemically unique from striatal projection neurons, they may provide promising therapeutic targets for the treatment of a variety of striatal-based disorders.</p

Distinct roles of GABAergic interneurons in the regulation of striatal output pathways.

Striatal GABAergic microcircuits are critical for motor function, yet their properties remain enigmatic due to difficulties in targeting striatal interneurons for electrophysiological analysis. Here, we use Lhx6-GFP transgenic mice to identify GABAergic interneurons and investigate their regulation of striatal direct- and indirect-pathway medium spiny neurons (MSNs). We find that the two major interneuron populations, persistent low-threshold spiking (PLTS) and fast spiking (FS) interneurons, differ substantially in their excitatory inputs and inhibitory outputs. Excitatory synaptic currents recorded from PLTS interneurons are characterized by a small, nonrectifying AMPA receptor-mediated component and a NMDA receptor-mediated component. In contrast, glutamatergic synaptic currents in FS interneurons have a large, strongly rectifying AMPA receptor-mediated component, but no detectable NMDA receptor-mediated responses. Consistent with their axonal morphology, the output of individual PLTS interneurons is relatively weak and sparse, whereas FS interneurons are robustly connected to MSNs and other FS interneurons and appear to mediate the bulk of feedforward inhibition. Synaptic depression of FS outputs is relatively insensitive to firing frequency, and dynamic-clamp experiments reveal that these short-term dynamics enable feedforward inhibition to remain efficacious across a broad frequency range. Surprisingly, we find that FS interneurons preferentially target direct-pathway MSNs over indirect-pathway MSNs, suggesting a potential mechanism for rapid pathway-specific regulation of striatal output pathways.</p

Selective inhibition of striatal fast-spiking interneurons causes dyskinesias.

Fast-spiking interneurons (FSIs) can exert powerful control over striatal output, and deficits in this cell population have been observed in human patients with Tourette syndrome and rodent models of dystonia. However, a direct experimental test of striatal FSI involvement in motor control has never been performed. We applied a novel pharmacological approach to examine the behavioral consequences of selective FSI suppression in mouse striatum. IEM-1460, an inhibitor of GluA2-lacking AMPARs, selectively blocked synaptic excitation of FSIs but not striatal projection neurons. Infusion of IEM-1460 into the sensorimotor striatum reduced the firing rate of FSIs but not other cell populations, and elicited robust dystonia-like impairments. These results provide direct evidence that hypofunction of striatal FSIs can produce movement abnormalities, and suggest that they may represent a novel therapeutic target for the treatment of hyperkinetic movement disorders.</p

Rapid target-specific remodeling of fast-spiking inhibitory circuits after loss of dopamine.

In Parkinson's disease (PD), dopamine depletion alters neuronal activity in the direct and indirect pathways and leads to increased synchrony in the basal ganglia network. However, the origins of these changes remain elusive. Because GABAergic interneurons regulate activity of projection neurons and promote neuronal synchrony, we recorded from pairs of striatal fast-spiking (FS) interneurons and direct- or indirect-pathway MSNs after dopamine depletion with 6-OHDA. Synaptic properties of FS-MSN connections remained similar, yet within 3 days of dopamine depletion, individual FS cells doubled their connectivity to indirect-pathway MSNs, whereas connections to direct-pathway MSNs remained unchanged. A model of the striatal microcircuit revealed that such increases in FS innervation were effective at enhancing synchrony within targeted cell populations. These data suggest that after dopamine depletion, rapid target-specific microcircuit organization in the striatum may lead to increased synchrony of indirect-pathway MSNs that contributes to pathological network oscillations and motor symptoms of PD.</p

Cholinergic interneurons mediate fast VGluT3-dependent glutamatergic transmission in the striatum.

The neurotransmitter glutamate is released by excitatory projection neurons throughout the brain. However, non-glutamatergic cells, including cholinergic and monoaminergic neurons, express markers that suggest that they are also capable of vesicular glutamate release. Striatal cholinergic interneurons (CINs) express the Type-3 vesicular glutamate transporter (VGluT3), although whether they form functional glutamatergic synapses is unclear. To examine this possibility, we utilized mice expressing Cre-recombinase under control of the endogenous choline acetyltransferase locus and conditionally expressed light-activated Channelrhodopsin2 in CINs. Optical stimulation evoked action potentials in CINs and produced postsynaptic responses in medium spiny neurons that were blocked by glutamate receptor antagonists. CIN-mediated glutamatergic responses exhibited a large contribution of NMDA-type glutamate receptors, distinguishing them from corticostriatal inputs. CIN-mediated glutamatergic responses were insensitive to antagonists of acetylcholine receptors and were not seen in mice lacking VGluT3. Our results indicate that CINs are capable of mediating fast glutamatergic transmission, suggesting a new role for these cells in regulating striatal activity.</p

Light-evoked CIN action potentials evoke glutamatergic responses in MSNs.

<p>(A) <i>inset,</i> Schematic diagram of the recording conditions: whole-cell recordings were made from MSNs that neighbored ChR2-expressing CINs (red). The blue circle indicates the region stimulated by blue light. <i>main panel,</i> Example EPSP recorded in an MSN in response to a 4 ms light pulse (blue bar). (B) Amplitudes of light-evoked EPSPs in the presence of antagonists of GABA_A (Picrotoxin, Ptx), muscarinic (Scopolamine, Scop), and nicotinic (Mecamylamine, Mec) receptors and following application of the AMPA/kainate glutamate receptor antagonist NBQX. (C) Example of light evoked (blue bar) CIN-mediated EPSCs in a voltage-clamped MSN at holding potentials of −70 and +40 mV demonstrating the large current that is visible more than 100 ms after the light pulse at +40 mV. The amplitudes of the rapid −70 and prolonged +40 mV EPSC components were measured in the periods indicated by the gray bars. (D) Confocal image of mCherry-positive ChR2-expressing fibers in the motor cortex, white matter (WM) and underlying striatum (Str). Large bundles of corticofugal fibers (white arrowheads) and diffuse small axonal collaterals are visible throughout the dorsolateral striatum. Scale bar 200 µm. (E) <i>inset,</i> Schematic diagram of the recording conditions: whole-cell recordings were made from MSNs neighboring ChR2-expressing corticostriatal fibers (red), and blue light was delivered to the region indicated by the blue circle. <i>main panel,</i> Example EPSCs recorded in an MSN in response to a blue light pulse at the indicated holding potentials. (F) <i>left</i>, Average amplitudes of light-evoked EPSCs measured in MSNs held at either −70 or +40 mV in response to ChR2-mediated activation of either CINs or corticostriatal fibers (Cx). <i>right</i>, Average ratio of +40/−70 mV current amplitudes measured following ChR2-mediated activation of either CINs or corticostriatal fibers.</p

ChR2-mediated activation of striatal cholinergic interneurons.

<p>(Ai) Confocal image of mCherry-positive ChR2-expressing neurons in the dorsal striatum of a ChAT-IRES-Cre mouse injected with AAV encoding DFI-ChR2-mCherry. (Aii) Fluorescence immunohistochemical staining for ChAT reveals cholinergic neurons. (Aiii) Merged image. (B) <i>inset,</i> Schematic diagram of the recording conditions: cell-attached recordings were made from ChR2-expressing CINs (red) in the dorsal striatum and blue light was delivered to the surrounding area (blue circle) through the microscope objective. <i>main panel,</i> Example action potential recorded in cell-attached mode from a ChR2-expressing CIN evoked by a 4 ms pulse of 473 nm light.</p

CIN-mediated glutamatergic currents in MSNs require VGluT3 expression.

<p>(A) Example light-evoked EPSCs at 32–34°C in an MSN held at −70 or +40 mV in an acute striatal slice of a DFI-ChR2-mCherry AAV injected ChAT-IRES-Cre mouse. (B) As in Panel A, showing recordings at 32–34°C obtained from an MSN in an acute slice of a DFI-ChR2-mCherry AAV injected GM60 mouse that expresses Cre under control of a BAC spanning the ChAT genomic locus. (C) As in Panel B, showing failures to evoke EPSCs in an MSN of a DFI-ChR2-mCherry AAV injected GM60; VGluT3^−/− mouse. (D) Average light-evoked EPSC amplitudes measured in MSNs at holding potentials of −70 and +40 mV in acute slices prepared from mice of the indicated genotypes.</p

Light-elicited inhibition of hilar GABAergic interneurons and activation of dentate granule neurons.

<p>(A, B) Example trace (A) and summary graph (B, n = 5) of eNpHR3.0-mediated membrane hyperpolarization of hilar GABAergic interneurons in brain slices. In this and subsequent panels, yellow bars indicate illumination time. (C) eNpHR3.0-mediated inhibition of spiking of hilar GABAergic interneurons in brain slices. (D) Schematic of <i>in vivo</i> optical stimulation and recording in the dentate gyrus (DG) of I12b-Cre mice injected with AAV1-DIO-eNpHR3.0-eYFP virus. (E) Example of a granule neuron recorded from the DG of an anaesthetized I12b-Cre mouse injected with AAV1-DIO-eNpHR3.0-eYFP that showed increased firing in response to yellow laser illumination. Inset shows spike waveform with laser illumination. (F) Average change in dentate granule neuron firing rates in response to yellow laser illumination in I12b-Cre mice injected with AAV1-DIO-eNpHR3.0-eYFP virus. Values are mean ± SEM (n = 7, *p<0.05, two-tailed and unpaired <i>t</i>-test).</p

Inhibition of hilar GABAergic interneuron activity impaired spatial learning and memory.

<p>(A) Coronal schematic of cannula placement and bilateral fiber-optic stimulation. (B) Protocols of mice used and laser illumination during hidden platform (H1–5) and probe (P-24 h and P-72 h) trials in the Morris water maze (MWM) test. (C–E) Learning curves of I12b-Cre (eNpHR3.0⁺) and wildtype (eNpHR3.0⁻) littermates injected with AAV1-DIO-eNpHR3.0-eYFP virus, with or without laser illumination in MWM tests. Points represent averages of daily trials. H, hidden platform sessions (two trials/session, two sessions/day); H0, first trial on H1; V, visible platform sessions (two trials/session, two sessions/day). Y-axis indicates time to reach the target platform (escape latency). Values are mean ± SEM and statistically evaluated by repeated measures ANOVA. (F) Swim speed did not differ significantly among different groups of mice during the MWM test. (G) Percent time spent in the target quadrant versus the other quadrants in the probe trial performed 24 h (probe 1) after the last hidden platform trial. (H) Percent time spent in the target quadrant versus the other quadrants in the probe trial performed 72 h (probe 2) after the last hidden platform trial. Values are mean ± SEM. n = 7–20 mice/group. *p<0.05 **p<0.01, ***p<0.005 (two tailed and unpaired <i>t</i>-test).</p