Tibor Koós

Inhibitory control of neostriatal projection neurons by GABAergic interneurons

The basal ganglia are a highly interconnected network of nuclei essential for the modulation and execution of voluntary behavior. The neostriatum is the principal input and one of the principal controllers of the output of the basal ganglia. Neostriatal projection neurons seem to be dynamically and powerfully controlled by GABAergic inputs, but the source(s) and physiological properties of these inputs remain unclear. Here we use paired whole-cell recordings to show that this inhibition derives from small populations of GABAergic interneurons that are themselves interconnected through functional electrotonic synapses. Inhibitory synaptic potentials generated from single interneurons are sufficiently powerful to delay or entirely block the generation of action potentials in a large number of projection neurons simultaneously.

Heterogeneity and diversity of striatal GABAergic interneurons.

The canonical view of striatal GABAergic interneurons has evolved over several decades of neuroanatomical/neurochemical and electrophysiological studies. From the anatomical studies, three distinct GABAergic interneuronal subtypes are generally recognized. The best-studied subtype expresses the calcium-binding protein, parvalbumin. The second best known interneuron type expresses a number of neuropeptides and enzymes, including neuropeptide Y, somatostatin, and nitric oxide synthase. The last GABAergic interneuron subtype expresses the calcium binding protein, calretinin. There is no overlap or co-localization of these three different sets of markers. The parvalbumin-immunoreactive GABAergic interneurons have been recorded in vitro and shown to exhibit a fast-spiking phenotype characterized by short duration action potentials with large and rapid spike AHPs. They often fire in a stuttering pattern of high frequency firing interrupted by periods of silence. They are capable of sustained firing rates of over 200 Hz. The NPY/SOM/NOS interneurons have been identified as PLTS cells, exhibiting very high input resistances, low threshold spike and prolonged plateau potentials in response to intracellular depolarization or excitatory synaptic stimulation. Thus far, no recordings from identified CR interneurons have been obtained. Recent advances in technological approaches, most notably the generation of several BAC transgenic mouse strains which express a fluorescent marker, enhanced green fluorescent protein, specifically and selectively only in neurons of a certain genetic makeup (e.g., parvalbumin-, neuropeptide Y-, or tyrosine hydroxylase-expressing neurons etc.) have led to the ability of electrophysiologists to visualize and patch specific neuron types in brain slices with epifluorescence illumination. This has led to a rapid expansion of the number of neurochemically and/or electrophysiologically identified interneuronal cell types in the striatum and elsewhere. This article will review the anatomy, neurochemistry, electrophysiology, synaptic connections, and function of the three “classic” striatal GABAergic interneurons as well as more recent data derived from in vitro recordings from BAC transgenic mice as well as recent in vivo data.

Glutamatergic Signaling by Mesolimbic Dopamine Neurons in the Nucleus Accumbens

Recent evidence suggests the intriguing possibility that midbrain dopaminergic (DAergic) neurons may use fast glutamatergic transmission to communicate with their postsynaptic targets. Because of technical limitations, direct demonstration of the existence of this signaling mechanism has been limited to experiments using cell culture preparations that often alter neuronal function including neurotransmitter phenotype. Consequently, it remains uncertain whether glutamatergic signaling between DAergic neurons and their postsynaptic targets exists under physiological conditions. Here, using an optogenetic approach, we provide the first conclusive demonstration that mesolimbic DAergic neurons in mice release glutamate and elicit excitatory postsynaptic responses in projection neurons of the nucleus accumbens. In addition, we describe the properties of the postsynaptic glutamatergic responses of these neurons during experimentally evoked burst firing of DAergic axons that reproduce the reward-related phasic population activity of the mesolimbic projection. These observations indicate that, in addition to DAergic mechanisms, mesolimbic reward signaling may involve glutamatergic transmission.

GABAergic circuits mediate the reinforcement-related signals of striatal cholinergic interneurons

Neostriatal cholinergic interneurons are believed to be important for reinforcement-mediated learning and response selection by signaling the occurrence and motivational value of behaviorally relevant stimuli through precisely timed multiphasic population responses. An important problem is to understand how these signals regulate the functioning of the neostriatum. Here we describe the synaptic organization of a previously unknown circuit that involves direct nicotinic excitation of several classes of GABAergic interneurons, including neuroptide Y–expressing neurogilaform neurons, and enables cholinergic interneurons to exert rapid inhibitory control of the activity of projection neurons. We also found that, in vivo, the dominant effect of an optogenetically reproduced pause-excitation population response of cholinergic interneurons was powerful and rapid inhibition of the firing of projection neurons that is coincident with synchronous cholinergic activation. These results reveal a previously unknown circuit mechanism that transmits reinforcement-related information of ChAT interneurons in the mouse neostriatal network.

Postnatal development of the rat neostriatum: electrophysiological, light- and electron-microscopic studies.

The postnatal development of the electrophysiological properties and morphology of rat neostriatum was studied using in vivo and in vitro intracellular recording and biocytin staining and light and electron microscopy. The principal neurons, the medium spiny neurons, were found to undergo a protracted postnatal development of their electrophysiological and morphological characteristics. Most of the intrinsic membrane properties of medium spiny neurons came to resemble those in the adult by the end of the 3rd postnatal week. Synaptic responses and spontaneous activity patterns in medium spiny neurons were dependent on the arrival and functional maturation of excitatory afferents from cortex and thalamus and did not become adult-like until the end of the 1st postnatal month.

Endogenous Hydrogen Peroxide Regulates the Excitability of Midbrain Dopamine Neurons via ATP-Sensitive Potassium Channels

ATP-sensitive K+ (KATP) channels link metabolic state to cell excitability. Here, we examined regulation of KATP channels in substantia nigra dopamine neurons by hydrogen peroxide (H2O2), which is produced in all cells during aerobic metabolism. Blockade of KATP channels by glibenclamide (100 nm) or depletion of intracellular H2O2 by including catalase, a peroxidase enzyme, in the patch pipette increased the spontaneous firing rate of all dopamine neurons tested in guinea pig midbrain slices. Using fluorescence imaging with dichlorofluorescein to visualize intracellular H2O2, we found that moderate increases in H2O2 during partial inhibition of glutathione (GSH) peroxidase by mercaptosuccinate (0.1-0.3 mm) had no effect on dopamine neuron firing rate. However, with greater GSH inhibition (1 mm mercaptosuccinate) or application of exogenous H2O2, 50% of recorded cells showed KATP channel-dependent hyperpolarization. Responsive cells also hyperpolarized with diazoxide, a selective opener for KATP channels containing sulfonylurea receptor SUR1 subunits, but not with cromakalim, a selective opener for SUR2-based channels, indicating that SUR1-based KATP channels conveyed enhanced sensitivity to elevated H2O2. In contrast, when endogenous H2O2 levels were increased after inhibition of catalase, the predominant peroxidase in the substantia nigra, with 3-amino-1,2,4-triazole (1 mm), all dopamine neurons responded with glibenclamide-reversible hyperpolarization. Fluorescence imaging of H2O2 indicated that catalase inhibition rapidly amplified intracellular H2O2, whereas inhibition of GSH peroxidase, a predominantly glial enzyme, caused a slower, smaller increase, especially in nonresponsive cells. Thus, endogenous H2O2 modulates neuronal activity via KATP channel opening, thereby enhancing the reciprocal relationship between metabolism and excitability.

Diode-probes for spatiotemporal optical control of multiple neurons in freely-moving animals

Neuronal control with high temporal precision is possible with optogenetics, yet currently available methods do not enable to control independently multiple locations in the brains of freely moving animals. Here, we describe a diode-probe system that allows real-time and location-specific control of neuronal activity at multiple sites. Manipulation of neuronal activity in arbitrary spatiotemporal patterns is achieved by means of an optoelectronic array, manufactured by attaching multiple diode-fiber assemblies to high-density silicon probes or wire tetrodes and implanted into the brains of animals that are expressing light-responsive opsins. Each diode can be controlled separately, allowing localized light stimulation of neuronal activators and silencers in any temporal configuration and concurrent recording of the stimulated neurons. Because the only connections to the animals are via a highly flexible wire cable, unimpeded behavior is allowed for circuit monitoring and multisite perturbations in the intact brain. The capacity of the system to generate unique neural activity patterns facilitates multisite manipulation of neural circuits in a closed-loop manner and opens the door to addressing novel questions.

Electrophysiological and Morphological Characteristics and Synaptic Connectivity of Tyrosine Hydroxylase-Expressing Neurons in Adult Mouse Striatum

Whole-cell recordings were obtained from tyrosine hydroxylase-expressing (TH+) neurons in striatal slices from bacterial artificial chromosome transgenic mice that synthesize enhanced green fluorescent protein (EGFP) selectively in neurons expressing TH transcriptional regulatory sequences. Stereological cell counting indicated that there were ∼2700 EGFP–TH+ neurons/striatum. Whole-cell recordings in striatal slices demonstrated that EGFP–TH+ neurons comprise four electrophysiologically distinct neuron types whose electrophysiological properties have not been reported previously in striatum. EGFP–TH+ neurons were identified in retrograde tracing studies as interneurons. Recordings from synaptically connected pairs of EGFP–TH+ interneurons and spiny neurons showed that the interneurons elicited GABAergic IPSPs/IPSCs in spiny neurons powerful enough to significantly delay evoked spiking. EGFP–TH+ interneurons responded to local or cortical stimulation with glutamatergic EPSPs. Local stimulation also elicited GABAA IPSPs, at least some of which arose from identified spiny neurons. Single-cell reverse transcription-PCR showed expression of VMAT1 in EGFP–TH+ interneurons, consistent with previous suggestions that these interneurons may be dopaminergic as well as GABAergic. All four classes of interneurons were medium sized with modestly branching, varicose dendrites, and dense, highly varicose axon collateral fields. These data show for the first time that there exists in the normal rodent striatum a substantial population of TH+/GABAergic interneurons comprising four electrophysiologically distinct subtypes whose electrophysiological properties differ significantly from those of previously described striatal GABAergic interneurons. These interneurons are likely to play an important role in striatal function through fast GABAergic synaptic transmission in addition to, and independent of, their potential role in compensation for dopamine loss in experimental or idiopathic Parkinsons disease.

A Novel Functionally Distinct Subtype of Striatal Neuropeptide Y Interneuron

We investigated the properties of neostriatal neuropeptide Y (NPY)-expressing interneurons in transgenic GFP (green fluorescent protein)-NPY reporter mice. In vitro whole-cell recordings and biocytin staining demonstrated the existence of a novel class of neostriatal NPY-expressing GABAergic interneurons that exhibit electrophysiological, neurochemical, and morphological properties strikingly different from those of previously described NPY-containing, plateau-depolarization low-threshold spike (NPY–PLTS) interneurons. The novel NPY interneuron type (NPY–neurogliaform) differed from previously described NPY–PLTS interneurons by exhibiting a significantly lower input resistance and hyperpolarized membrane potential, regular, nonaccommodating spiking in response to depolarizing current injections, and an absence of plateau depolarizations or low-threshold spikes. NPY–neurogliaform interneurons were also easily distinguished morphologically by their dense, compact, and highly branched dendritic and local axonal arborizations that contrasted sharply with the sparse and extended axonal and dendritic arborizations of NPY–PLTS interneurons. Furthermore, NPY–neurogliaform interneurons did not express immunofluorescence for somatostatin or nitric oxide synthase that was ubiquitous in NPY–PLTS interneurons. IPSP/Cs could only rarely be elicited in spiny projection neurons (SPNs) in paired recordings with NPY–PLTS interneurons. In contrast, the probability of SPN innervation by NPY–neurogliaform interneurons was extremely high, the synapse very reliable (no failures were observed), and the resulting postsynaptic response was a slow, GABAA receptor-mediated IPSC that has not been previously described in striatum but that has been elicited from NPY–GABAergic neurogliaform interneurons in cortex and hippocampus. These properties suggest unique and distinctive roles for NPY–PLTS and NPY–neurogliaform interneurons in the integrative properties of the neostriatum.

Differential Dopaminergic Modulation of Neostriatal Synaptic Connections of Striatopallidal Axon Collaterals

Recent studies have demonstrated that GABAergic synaptic transmission among neostriatal spiny projection neurons (SPNs) is strongly modulated by dopamine with individual connections exhibiting either D1 receptor (D1R)-mediated facilitation or D2 receptor (D2R)-mediated inhibition and, at least in some preparations, a subset of connections exhibiting both of these effects. In light of the cell type-specific expression of D1aR in striatonigral and D2R in striatopallidal neurons and the differential expression of the other D1 and D2 family dopamine receptors, we hypothesize that the nature of the dopaminergic modulation is specific to the types of SPNs that participate in the connection. Here the biophysical properties and dopaminergic modulation of intrastriatal connections formed by striatopallidal neurons were examined. Contrary to previous expectation, synapses formed by striatopallidal neurons were biophysically and pharmacologically heterogeneous. Two distinct types of axon collateral connections could be distinguished among striatopallidal neurons. The more common, small-amplitude connections (80%) exhibited mean IPSC amplitudes several times smaller than their less frequent large-amplitude counterparts, principally because of a smaller number of release sites involved. The two types of connections were also differentially regulated by dopamine. Small-amplitude connections exhibited strong and exclusively D2R-mediated presynaptic inhibition, whereas large-amplitude connections were unresponsive to dopamine. Synaptic connections from striatopallidal to striatonigral neurons exhibited exclusively D2R-mediated presynaptic inhibition that was similar to the regulation of small-amplitude connections between pairs of striatopallidal cells. Together, these findings demonstrate a previously unrecognized complexity in the organization and dopaminergic control of synaptic communication among SPNs.