Fast-spiking GABAergic Interneurons Research Articles

Cocaine Exposure Modulates Perineuronal Nets and Synaptic Excitability of Fast-Spiking Interneurons in the Medial Prefrontal Cortex.

We previously reported that perineuronal nets (PNNs) are required for cocaine-associated memories. Perineuronal nets are extracellular matrix that primarily surrounds parvalbumin (PV)-containing, GABAergic fast-spiking interneurons (FSIs) in the medial prefrontal cortex (mPFC). Here we measured the impact of acute (1 d) or repeated (5 d) cocaine exposure on PNNs and PV cells within the prelimbic and infralimbic regions of the mPFC. Adult rats were exposed to 1 or 5 d of cocaine and stained for PNNs (using Wisteria floribunda agglutinin) and PV intensity 2 or 24 h later. In the prelimbic and infralimbic PFC, PNN staining intensity decreased 2 h after 1 d of cocaine exposure but increased after 5 d of cocaine exposure. Cocaine also produced changes in PV intensity, which generally lagged behind that of PNNs. In the prelimbic PFC, both 1 and 5 d of cocaine exposure increased GAD65/67 puncta near PNN-surrounded PV cells, with an increase in the GAD65/67-to-VGluT1 puncta ratio after 5 d of cocaine exposure. In the prelimbic PFC, slice electrophysiology studies in FSIs surrounded by PNNs revealed that both 1 and 5 d of cocaine exposure reduced the number of action potentials 2 h later. Synaptic changes demonstrated that 5 d of cocaine exposure increased the inhibition of FSIs, potentially reducing the inhibition of pyramidal neurons and contributing to their hyperexcitability during relapse behavior. These early and rapid responses to cocaine may alter the network stability of PV FSIs that partially mediate the persistent and chronic nature of drug addiction.

Substantia nigra dopaminergic neurons and striatal interneurons are engaged in three parallel but interdependent postnatal neurotrophic circuits.

The striatum integrates motor behavior using a well‐defined microcircuit whose individual components are independently affected in several neurological diseases. The glial cell line‐derived neurotrophic factor (GDNF), synthesized by striatal interneurons, and Sonic hedgehog (Shh), produced by the dopaminergic neurons of the substantia nigra (DA SNpc), are both involved in the nigrostriatal maintenance but the reciprocal neurotrophic relationships among these neurons are only partially understood. To define the postnatal neurotrophic connections among fast‐spiking GABAergic interneurons (FS), cholinergic interneurons (ACh), and DA SNpc, we used a genetically induced mouse model of postnatal DA SNpc neurodegeneration and separately eliminated Smoothened (Smo), the obligatory transducer of Shh signaling, in striatal interneurons. We show that FS postnatal survival relies on DA SNpc and is independent of Shh signaling. On the contrary, Shh signaling but not dopaminergic striatal innervation is required to maintain ACh in the postnatal striatum. ACh are required for DA SNpc survival in a GDNF‐independent manner. These data demonstrate the existence of three parallel but interdependent neurotrophic relationships between SN and striatal interneurons, partially defined by Shh and GDNF. The definition of these new neurotrophic interactions opens the search for new molecules involved in the striatal modulatory circuit maintenance with potential therapeutic value.

Pharmacological rescue of mutated Kv3.1 ion-channel linked to progressive myoclonus epilepsies

Progressive myoclonus epilepsies (PMEs) constitute a cluster of inherent, genetically diverse, rare seizure disorders characterized by ataxia, tonic-clonic seizures, and action myoclonus. Recently, a mutation in the KCNC1 gene (Arg320His) was described in a group of PME patients. The KCNC1 gene encodes the Kv3.1 potassium ion channel responsible for the rapid repolarization of the membrane potential following action potential firing in fast spiking GABAergic interneurons (FSI), thereby enabling high firing frequency. In the present study, we demonstrate that the Arg320His mutation cause a reduction in the Kv3.1 current amplitude and acts in a dominantly negative fashion. The mutation profoundly affects channel activation and deactivation kinetics, and we further find that it impairs recruitment of the Kv3.1 channel to the plasma membrane. The Kv3 activating compound, RE01, partly rescues the electrophysiological deficit, suggesting that pharmacological activation of Kv3.1 activity might be a feasible approach for treatment of this cohort of PME patients.

Expression and localization of CB1R, NAPE-PLD, and FAAH in the vervet monkey nucleus accumbens

Extensive rodent literature suggests that the endocannabinoid (eCB) system present in the nucleus accumbens (NAc) modulates dopamine (DA) release in this area. However, expression patterns of the cannabinoid receptor type 1 (CB1R), the synthesizing enzyme N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD), and the degradation enzyme fatty acid amide hydrolase (FAAH) in the NAc have not yet been described in non-human primates. The goal of this study is therefore to characterize the expression and localization of the eCB system within the NAc of vervet monkeys (Chlorocebus sabaeus) using Western blots and immunohistochemistry. Results show that CB1R, NAPE-PLD, and FAAH are expressed across the NAc rostrocaudal axis, both in the core and shell. CB1R, NAPE-PLD, and FAAH are localized in medium spiny neurons (MSNs) and fast-spiking GABAergic interneurons (FSIs). Dopaminergic projections and astrocytes did not express CB1R, NAPE-PLD, or FAAH. These data show that the eCB system is present in the vervet monkey NAc and supports its role in the primate brain reward circuit.

Multiple long-range inputs evoke NMDA currents in prefrontal cortex fast-spiking interneurons.

Several aspects of schizophrenia can be mimicked acutely in healthy human volunteers via administration of NMDA glutamate receptor (NMDAR) antagonists. As these agents decrease firing rates in prefrontal cortical (PFC) GABAergic fast-spiking interneurons (FSI) in animal studies, a leading hypothesis on schizophrenia pathophysiology is that NMDAR in FSI are impaired. However, whole-cell recordings of FSI in slices of adult mouse PFC revealed limited amounts of NMDAR-mediated current. Since those studies used local electrical stimulation to activate a heterogeneous set of synaptic inputs to the recorded cell, it is unclear whether specific afferent inputs may preferentially drive NMDAR responses in FSI. Here, we expressed opsins in discrete brain regions projecting to the PFC in adult male mice, enabling light-activation of defined, homogenous sets of long-range inputs to FSI and pyramidal neurons recorded in slices containing medial PFC (mPFC). Stimulation of axons originating from either the contralateral mPFC, ventral hippocampus, or mediodorsal thalamus evoked NMDAR-mediated currents in the vast majority of FSI and in all pyramidal neurons recorded. The observation that multiple long-range inputs to mPFC FSI elicit NMDAR currents suggests that the NMDAR-hypofunction model of schizophrenia may still imply a loss of interneuron inputs, but the sources of reduced excitation may originate from sites upstream of the PFC.

Clenbuterol reduces GABAergic transmission in prefrontal cortex layer 5/6 pyramidal neurons of juvenile rat via reducing action potentials firing frequency of GABAergic interneurons.

Beta-adrenoceptors (β2 -ARs) have beneficial effects on prefrontal cortex (PFC) working memory, however, the cellular and molecular mechanisms are unclear yet. In this study, we probed the effect of β2 -AR-selective agonist clenbuterol (Clen) on synaptic transmission in layer 5/6 pyramidal neurons of PFC. Bath application of Clen reduced spontaneous IPSC (sIPSC) frequency without effects on sEPSCs. Clen did not alter the frequency and amplitude of miniature IPSCs (mIPSCs), but exerted heterogeneous effects on evoked IPSCs (eIPSCs) recorded from PFC layer 5/6 pyramidal neurons. Clen decreased the firing rate of action potentials of fast-spiking GABAergic interneurons. Clen-induced hyperpolarization of fast-spiking GABAergic interneurons required potentiation of an inward rectifier K+ channels. Clen-induced hyperpolarization of fast-spiking interneurons was dependent on Gs protein rather than cAMP and protein kinase A. Our findings demonstrate that Clen (10μM) enhances inward rectifier K+ channels via Gs protein to cause membrane hyperpolarization of fast-spiking GABAergic interneurons resulting in reduction of action potentials firing rate to reduce GABAergic transmission.

Delayed Maturation of Fast-Spiking Interneurons Is Rectified by Activation of the TrkB Receptor in the Mouse Model of Fragile X Syndrome.

Fragile X syndrome (FXS) is a neurodevelopmental disorder that is a leading cause of inherited intellectual disability, and the most common known cause of autism spectrum disorder. FXS is broadly characterized by sensory hypersensitivity and several developmental alterations in synaptic and circuit function have been uncovered in the sensory cortex of the mouse model of FXS (Fmr1 KO). GABA-mediated neurotransmission and fast-spiking (FS) GABAergic interneurons are central to cortical circuit development in the neonate. Here we demonstrate that there is a delay in the maturation of the intrinsic properties of FS interneurons in the sensory cortex, and a deficit in the formation of excitatory synaptic inputs on to these neurons in neonatal Fmr1 KO mice. Both these delays in neuronal and synaptic maturation were rectified by chronic administration of a TrkB receptor agonist. These results demonstrate that the maturation of the GABAergic circuit in the sensory cortex is altered during a critical developmental period due in part to a perturbation in BDNF-TrkB signaling, and could contribute to the alterations in cortical development underlying the sensory pathophysiology of FXS.SIGNIFICANCE STATEMENT Fragile X (FXS) individuals have a range of sensory related phenotypes, and there is growing evidence of alterations in neuronal circuits in the sensory cortex of the mouse model of FXS (Fmr1 KO). GABAergic interneurons are central to the correct formation of circuits during cortical critical periods. Here we demonstrate a delay in the maturation of the properties and synaptic connectivity of interneurons in Fmr1 KO mice during a critical period of cortical development. The delays both in cellular and synaptic maturation were rectified by administration of a TrkB receptor agonist, suggesting reduced BDNF-TrkB signaling as a contributing factor. These results provide evidence that the function of fast-spiking interneurons is disrupted due to a deficiency in neurotrophin signaling during early development in FXS.

Ablation of C-fibers decreases quantal size of GABAergic synaptic transmission in the insular cortex

The primary sensory cortex exhibits neuroplastic changes responding to sensory disturbances, and GABAergic synaptic transmission plays a critical role in the regulation of plasticity. The insular cortex (IC) integrates orofacial nociceptive signals conveyed via myelinated Aδ- and unmyelinated C-fibers. However, it has been unknown whether a disturbance of nociceptive inputs, such as a deletion of the peripheral nerves, alters GABAergic local circuit in IC. The present study elucidated GABAergic synaptic transmission in the model rat whose C-fibers were ablated by capsaicin injection 1–2 days after birth. In vivo optical imaging revealed that capsaicin-treated rats showed a facilitative excitatory propagation in IC responding to dental pulp stimulation. Whole-cell patch-clamp recording from pyramidal neurons (Pyr) demonstrated that capsaicin-treated rats showed the smaller amplitude of miniature inhibitory postsynaptic currents (IPSCs) than sham-treated rats without changing the frequency. Furthermore, replacement of extracellular Ca2+ to Sr2+, which induces an asynchronous release of neurotransmitters in the quantal size, induced a smaller amplitude of asynchronous unitary IPSCs recorded from fast-spiking GABAergic interneuron to Pyr connections in capsaicin-treated rats than sham-treated rats. These results suggest that capsaicin treatment depresses IPSCs via a postsynaptic mechanism. To confirm this possibility, the variance-mean analysis of unitary IPSCs was employed and we found that quantal size of GABAergic synaptic transmission was smaller in capsaicin-treated rats than in sham-treated rats. These results suggest that ablation of C-fibers induces plastic changes in GABAergic synaptic transmission by decreasing postsynaptic GABAA receptor-mediated conductance, which is a possible mechanism of the facilitative excitation in IC of capsaicin-treated rats.

Kv3 Channels: Enablers of Rapid Firing, Neurotransmitter Release, and Neuronal Endurance.

The intrinsic electrical characteristics of different types of neurons are shaped by the K+ channels they express. From among the more than 70 different K+ channel genes expressed in neurons, Kv3 family voltage-dependent K+ channels are uniquely associated with the ability of certain neurons to fire action potentials and to release neurotransmitter at high rates of up to 1,000 Hz. In general, the four Kv3 channels Kv3.1-Kv3.4 share the property of activating and deactivating rapidly at potentials more positive than other channels. Each Kv3 channel gene can generate multiple protein isoforms, which contribute to the high-frequency firing of neurons such as auditory brain stem neurons, fast-spiking GABAergic interneurons, and Purkinje cells of the cerebellum, and to regulation of neurotransmitter release at the terminals of many neurons. The different Kv3 channels have unique expression patterns and biophysical properties and are regulated in different ways by protein kinases. In this review, we cover the function, localization, and modulation of Kv3 channels and describe how levels and properties of the channels are altered by changes in ongoing neuronal activity. We also cover how the protein-protein interaction of these channels with other proteins affects neuronal functions, and how mutations or abnormal regulation of Kv3 channels are associated with neurological disorders such as ataxias, epilepsies, schizophrenia, and Alzheimer's disease.

P 76 Maturation deficits in striatal interneurons in a phenotypic model of dystonia

GABAergic disinhibition has been suggested to play a critical role in the pathophysiology of several basal ganglia disorders, including dystonia, a common incurable movement disorder characterized by involuntary muscle contractions resulting in abnormal movements and/or postures. Previous studies have shown a deficit of striatal GABAergic interneurons (IN) in the dtsz mutant hamster, one of the few phenotypic animal models of dystonia. The resulting lack of inhibition of striatal projection neurons is likely involved in development of dystonic episodes which is corroborated by antidystonic effects of GABA potentiating drugs in the animal model and in dystonia patients. However, mechanisms underlying this deficit in striatal IN are largely unknown. In the present study, we investigated the migration and maturation of striatal IN during postnatal development (18 days of age) and at age of highest severity of dystonia (33 days of age) in the dtsz hamster by immunohistochemistry and real time PCR. In line with previous findings, the density of GAD67 positive IN and the level of parvalbumin mRNA, a marker for fast spiking GABAergic IN, were lower in mutant hamsters than in controls. However, an unaltered density of Nkx2.1 labeled cells and Nkx2.1 mRNA level indicated that the migration of GABAergic IN into the striatum was not retarded. KCC2 cation/chloride transporter and cytosolic carboanhydrase VII mRNA expression, used as markers for maturation of striatal GABAergic signaling, as well as expression of BDNF were unaltered. However, we found a reduced number of IN expressing the alpha1 subunit of the GABAA-receptor (37.5%) in dtsz hamsters at an age of 33 days, but not after spontaneous remission of dystonia at an age of 90 days. Since IN shift expression from alpha2 to alpha1 subunits during postnatal maturation, this result together with a decreased parvalbumin mRNA expression suggest a delayed maturation of striatal GABAergic IN in this animal model, which might underlie abnormal neuronal activity and striatal plasticity. The potential link between alterations in GABAA receptor subunits and GABAergic disinhibition in dystonia deserves further attention in research on the pathophysiology and therapeutic targets.

Generation of low-gamma oscillations in a GABAergic network model of the striatum

Striatal oscillations in the low-gamma frequency range have been consistently recorded in a number of experimental studies. However, whether these rhythms are locally generated in the striatum circuit, which is mainly composed of GABAergic neurons, remains an open question. GABAergic medium spiny projection neurons represent the great majority of striatal neurons, but they fire at very low rates. GABAergic fast-spiking interneurons typically show firing rates that are approximately 10 times higher than those of principal neurons, but they are a very small minority of the total neuronal population. In this study, based on physiological constraints we developed a computational network model of these neurons and dissected the oscillations. Simulations showed that the population of medium spiny projection neurons, and not the GABAergic fast-spiking interneurons, determines the frequency range of the oscillations. D2-type dopamine receptor-expressing neurons dominate the generation of low-gamma rhythms. Feedforward inputs from GABAergic fast-spiking interneurons promote the oscillations by strengthening the inhibitory interactions between medium spiny projection neurons. The promotion effect is independent of the degree of synchronization in the fast-spiking interneuron population but affected by the strength of their feedforward inputs to medium spiny projection neurons. Our results provide a theoretical explanation for how firing properties and connections of the three types of GABAergic neurons, which are susceptible to on-going behaviors, experience, and dopamine disruptions, sculpt striatal oscillations.

Structural alterations in fast-spiking GABAergic interneurons in a model of posttraumatic neocortical epileptogenesis

Electrophysiological experiments in the partial cortical isolation (“undercut” or “UC”) model of injury-induced neocortical epileptogenesis have shown alterations in GABAergic synaptic transmission attributable to abnormalities in presynaptic terminals. To determine whether the decreased inhibition was associated with structural abnormalities in GABAergic interneurons, we used immunocytochemical techniques, confocal microscopy and EM in UC and control sensorimotor rat cortex to analyze structural alterations in fast-spiking parvalbumin-containing interneurons and pyramidal (Pyr) cells of layer V. Principle findings were: 1) there were no decreases in counts of parvalbumin (PV)- or GABA-immunoreactive interneurons in UC cortex, however there were significant reductions in expression of VGAT and GAD-65 and -67 in halos of GABAergic terminals around Pyr somata in layer V. 2) Consistent with previous results, somatic size and density of Pyr cells was decreased in infragranular layers of UC cortex. 3) Dendrites of biocytin-filled FS interneurons were significantly decreased in volume. 4) There were decreases in the size and VGAT content of GABAergic boutons in axons of biocytin-filled FS cells in the UC, together with a decrease in colocalization with postsynaptic gephyrin, suggesting a reduction in GABAergic synapses. Quantitative EM of layer V Pyr somata confirmed the reduction in inhibitory synapses. 5) There were marked and lasting reductions in brain derived neurotrophic factor (BDNF)-IR and -mRNA in Pyr cells and decreased TrkB-IR on PV cells in UC cortex. 6) Results lead to the hypothesis that reduction in trophic support by BDNF derived from Pyr cells may contribute to the regressive changes in axonal terminals and dendrites of FS cells in the UC cortex and decreased GABAergic inhibition. SignificanceInjury to cortical structures is a major cause of epilepsy, accounting for about 20% of cases in the general population, with an incidence as high as ~50% among brain-injured personnel in wartime. Loss of GABAergic inhibitory interneurons is a significant pathophysiological factor associated with epileptogenesis following brain trauma and other etiologies. Results of these experiments show that the largest population of cortical interneurons, the parvalbumin-containing fast-spiking (FS) interneurons, are preserved in the partial neocortical isolation model of partial epilepsy. However, axonal terminals of these cells are structurally abnormal, have decreased content of GABA synthetic enzymes and vesicular GABA transporter and make fewer synapses onto pyramidal neurons. These structural abnormalities underlie defects in GABAergic neurotransmission that are a key pathophysiological factor in epileptogenesis found in electrophysiological experiments. BDNF, and its TrkB receptor, key factors for maintenance of interneurons and pyramidal neurons, are decreased in the injured cortex. Results suggest that supplying BDNF to the injured epileptogenic brain may reverse the structural and functional abnormalities in the parvalbumin FS interneurons and provide an antiepileptogenic therapy.

A small molecule activator of Nav 1.1 channels increases fast-spiking interneuron excitability and GABAergic transmission invitro and has anti-convulsive effects invivo.

Nav 1.1 (SCN1A) channels primarily located in gamma-aminobutyric acid (GABA)ergic fast-spiking interneurons are pivotal for action potential generation and propagation in these neurons. Inappropriate function of fast-spiking interneurons, leading to disinhibition of pyramidal cells and network desynchronization, correlates with decreased cognitive capability. Further, reduced functionality of Nav 1.1 channels is linked to various diseases in the central nervous system. There is, at present, however no subtype selective pharmacological activators of Nav 1.1 channels available for studying pharmacological modulation of interneuron function. In the current study, we identified a small molecule Nav 1.1 activator, 3-amino-5-(4-methoxyphenyl)thiophene-2-carboxamide, named AA43279, and provided an invitro to invivo characterization of the compound. In HEK-293 cells expressing human Nav 1.1 channels, AA43279 increased the Nav 1.1-mediated current in a concentration-dependent manner mainly by impairing the fast inactivation kinetics of the channels. In rat hippocampal brain slices, AA43279 increased the firing activity of parvalbumin-expressing, fast-spiking GABAergic interneurons and increased the spontaneous inhibitory post-synaptic currents (sIPSCs) recorded from pyramidal neurons. When tested invivo, AA43279 had anti-convulsive properties in the maximal electroshockseizurethreshold test. AA43279 was tested for off-target effects on 72 different proteins, including Nav 1.2, Nav 1.4, Nav 1.5, Nav 1.6 and Nav 1.7 and exhibited reasonable selectivity. Taken together, AA43279 might constitute a valuable tool compound for revealing biological functions of Nav 1.1 channels.

Kv3.1/Kv3.2 channel positive modulators enable faster activating kinetics and increase firing frequency in fast-spiking GABAergic interneurons

Due to their fast kinetic properties, Kv3.1 voltage gated potassium channels are important in setting and controlling firing frequency in neurons and pivotal in generating high frequency firing of interneurons. Pharmacological activation of Kv3.1 channels may possess therapeutic potential for treatment of epilepsy, hearing disorders, schizophrenia and cognitive impairments.Here we thoroughly investigate the selectivity and positive modulation of the two small molecules, EX15 and RE01, on Kv3 channels. Selectivity studies, conducted in Xenopus laevis oocytes confirmed a positive modulatory effect of the two compounds on Kv3.1 and to a minor extent on Kv3.2 channels. RE01 had no effect on the Kv3.3 and Kv3.4 channels, whereas EX15 had an inhibitory impact on the Kv3.4 mediated current.Voltage-clamp experiments in monoclonal hKv3.1b/HEK293 cells (34 °C) revealed that the two compounds indeed induced larger currents and faster activation kinetics. They also decrease the speed of deactivation and shifted the voltage dependence of activation, to a more negative activation threshold. Application of action potential clamping and repetitive stimulation protocols of hKv3.1b expressing HEK293 cells revealed that EX15 and RE01 significantly increased peak amplitude, half width and decay time of Kv3.1 mediated currents, even during high-frequency action potential clamping (250 Hz).In rat hippocampal slices, EX15 and RE01 increased neuronal excitability in fast-spiking interneurons in dentate gyrus. Action potential frequency was prominently increased at minor depolarizing steps, whereas more marginal effects of EX15 and RE01 were observed after stronger depolarizations.In conclusion, our results suggest that EX15 and RE01 positive modulation of Kv3.1 and Kv3.2 currents facilitate increased firing frequency in fast-spiking GABAergic interneurons.

Plasticity in Single Axon Glutamatergic Connection to GABAergic Interneurons Regulates Complex Events in the Human Neocortex.

In the human neocortex, single excitatory pyramidal cells can elicit very large glutamatergic EPSPs (VLEs) in inhibitory GABAergic interneurons capable of triggering their firing with short (3–5 ms) delay. Similar strong excitatory connections between two individual neurons have not been found in nonhuman cortices, suggesting that these synapses are specific to human interneurons. The VLEs are crucial for generating neocortical complex events, observed as single pyramidal cell spike-evoked discharge of cell assemblies in the frontal and temporal cortices. However, long-term plasticity of the VLE connections and how the plasticity modulates neocortical complex events has not been studied. Using triple and dual whole-cell recordings from synaptically connected human neocortical layers 2–3 neurons, we show that VLEs in fast-spiking GABAergic interneurons exhibit robust activity-induced long-term depression (LTD). The LTD by single pyramidal cell 40 Hz spike bursts is specific to connections with VLEs, requires group I metabotropic glutamate receptors, and has a presynaptic mechanism. The LTD of VLE connections alters suprathreshold activation of interneurons in the complex events suppressing the discharge of fast-spiking GABAergic cells. The VLEs triggering the complex events may contribute to cognitive processes in the human neocortex, and their long-term plasticity can alter the discharging cortical cell assemblies by learning.

Altered postnatal maturation of striatal GABAergic interneurons in a phenotypic animal model of dystonia

GABAergic disinhibition has been suggested to play a critical role in the pathophysiology of several basal ganglia disorders, including dystonia, a common movement disorder. Previous studies have shown a deficit of striatal GABAergic interneurons (IN) in the dtsz mutant hamster, one of the few phenotypic animal models of dystonia. However, mechanisms underlying this deficit are largely unknown. In the present study, we investigated the migration and maturation of striatal IN during postnatal development (18days of age) and at age of highest severity of dystonia (33days of age) in this hamster model. In line with previous findings, the density of GAD67-positive IN and the level of parvalbumin mRNA, a marker for fast spiking GABAergic IN, were lower in the dtsz mutant than in control hamsters. However, an unaltered density of Nkx2.1 labeled cells and Nkx2.1 mRNA level suggested that the migration of GABAergic IN into the striatum was not retarded. Therefore, different factors that indicate maturation of GABAergic IN were determined. While mRNA of the KCC2 cation/chloride transporters and the cytosolic carboanhydrase VII, used as markers for the so called GABA switch, as well as BDNF were unaltered, we found a reduced number of IN expressing the alpha1 subunit of the GABAA-receptor (37.5%) in dtsz hamsters at an age of 33days, but not after spontaneous remission of dystonia at an age of 90days. Since IN shift expression from alpha2 to alpha1 subunits during postnatal maturation, this result together with a decreased parvalbumin mRNA expression suggest a delayed maturation of striatal GABAergic IN in this animal model, which might underlie abnormal neuronal activity and striatal plasticity.

Opioid subtype- and cell-type-dependent regulation of inhibitory synaptic transmission in the rat insular cortex

The insular cortex (IC) plays a principal role in the regulation of pain processing. Although opioidergic agonists depress cortical excitatory synaptic transmission, little is known about opioidergic roles in inhibitory synaptic transmission. In the IC, the opioid receptors differentially regulate the excitatory propagation: agonists of the mu (MOR), delta (DOR), and kappa (KOR) exhibit suppressive, facilitative, and little effects, respectively. Thus, we aimed to examine the effects of opioid receptor agonists on unitary inhibitory postsynaptic currents (uIPSCs) in the IC. Pyramidal and GABAergic neurons in the rat IC were recorded by a multiple whole-cell patch-clamp technique. [D-Ala2,N-Me-Phe4,Gly5-ol]-Enkephalin acetate salt (DAMGO), an MOR agonist, reduced uIPSC amplitude by 74% in fast-spiking GABAergic interneuron (FS)→FS connections without a significant effect on FS→pyramidal cell (Pyr) connections. These effects of DAMGO were also observed in non-FS→FS and non-FS→Pyr connections: DAMGO reduced the uIPSC amplitude in non-FS→FS but not in non-FS→Pyr connections. DAMGO-induced depression of uIPSCs was blocked by the MOR antagonist, D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2. The DOR agonist, [D-Pen2,5]-Enkephalin hydrate (DPDPE), reduced uIPSC amplitude by 39% in FS→FS and by 49% in FS→Pyr connections, which was antagonized by the DOR antagonist, naltrindole. However, DPDPE had little effect on non-FS→FS/Pyr connections. (±)-trans-U-50488 methanesulfonate salt (U50488), a KOR agonist, had little effect on uIPSC in FS→FS/Pyr connections. These results suggest that MOR-induced uIPSC depression in FS→FS and non-FS→FS, but not FS→Pyr and non-FS→Pyr connections, results in the depression of excitatory propagation in the IC, which may be an underlying mechanism of the powerful analgesic effects of MOR agonists.

Prolonged withdrawal from cocaine self-administration affects prefrontal cortex- and basolateral amygdala-nucleus accumbens core circuits but not accumbens GABAergic local interneurons.

Withdrawal from extended-access cocaine self-administration leads to progressive intensification ('incubation') of cocaine craving. After prolonged withdrawal (1-2 months), when craving is high, expression of incubation depends on strengthening of excitatory inputs to medium spiny neurons (MSN) of the nucleus accumbens (NAc). These excitatory inputs interact with the intra-NAc GABAergic 'microcircuit', composed of MSN axon collaterals and GABAergic interneurons. Here, we investigated whether the increased glutamatergic neurotransmission observed after prolonged withdrawal is accompanied by altered GABAergic neurotransmission, focusing on NAc core. Rats self-administered cocaine or saline (6 hours/day) and then underwent >40 days of withdrawal. First, we investigated parvalbumin positive (PV+) interneurons, GABAergic fast-spiking interneurons that regulate MSN activity. Immunohistochemical studies revealed no significant change in PV signal intensity or the number of PV+ cells in cocaine rats versus saline controls. We then screened PV and other interneuron markers using immunoblotting. We detected no changes in levels of PV, calretinin, calbindin or neuronal nitric oxide synthase. Because expression of these markers is activity dependent, our results suggest no marked changes in interneuron activity. Finally, we utilized local field potential recording, which can detect GABA-mediated alterations at the circuit level, to investigate potential changes in two circuits implicated in cocaine craving: prelimbic prefrontal cortex to NAc core and basolateral amygdala to NAc core. We detected differential adaptations in these circuits, some of which may involve GABA. Overall, our results suggest that alterations in GABA transmission may accompany incubation of cocaine craving, but they are circuit specific and less pronounced than alterations in glutamate transmission.

Axon initial segment potassium channel density in cortical neurons

There is a growing interest in estimating actual density ranges of Na+ channels in the very thin axon, especially in the action potential (AP) initiation zone, i.e., the axon initial segment (AIS, 20-50 microns away from the cell body). Both immunostaining studies and patch-clamp recordings indicated a relatively high density of Na+ channels in AIS of either pyramidal regular-spiking (RS) cells [1] or fast-spiking (FS) GABAergic interneurons [2,3]. Here, we investigated potassium channel densities in AISs of both RS and FS cells in same recording conditions. Our axonal recordings directly revealed that there is a very lower potassium density gK = 185.8±19 pS/µm2 N = 16) for the RS AIS while a higher gK (495.7±108 pS/µm2, N = 11) for FS AIS, see Figure ​Figure1A.1A. For both the RS pyramidal cells and FS PV cells, partially blocking K+ channels by applying 4-AP broadened the spike duration and decreased the dV/dt ratio significantly (P < 0.05) (For RS cells: N = 5; For RS cells: N = 4). Interestingly, we observed that the AP dV/dt ratio is an exponentially decaying function of the spike duration for both RS- and FS-spikings (see Figure ​Figure1B),1B), such that y = 0.12+0.16EXP((0.5-x)/0.2), where y represents the dV/dt ratio and × represents the AP duration. These observations suggest strongly that potassium channel density is one of the major intrinsic factors dominating the spike shape properties, especially half-height spike duration and dV/dt ratio. Figure 1 A. The bar graph shows peak K+ conductance density recorded in axon initial segment of cells (16 recording axons for RS pyramidal cell, 11 for FS interneuron) by outside-out axon patch recording technique. B. Summary of the results from partially blocking ... In sum, the significant difference in potassium channel density in axonal initial segment where action potentials are initiated may play a critical role in controlling action potential properties of both RS- and FS-spiking cells in nervous system by the same general biophysical rule. These results may be important for constructing computational models of different types of cortical neurons.

Structured Connectivity in Cerebellar Inhibitory Networks

SummaryDefining the rules governing synaptic connectivity is key to formulating theories of neural circuit function. Interneurons can be connected by both electrical and chemical synapses, but the organization and interaction of these two complementary microcircuits is unknown. By recording from multiple molecular layer interneurons in the cerebellar cortex, we reveal specific, nonrandom connectivity patterns in both GABAergic chemical and electrical interneuron networks. Both networks contain clustered motifs and show specific overlap between them. Chemical connections exhibit a preference for transitive patterns, such as feedforward triplet motifs. This structured connectivity is supported by a characteristic spatial organization: transitivity of chemical connectivity is directed vertically in the sagittal plane, and electrical synapses appear strictly confined to the sagittal plane. The specific, highly structured connectivity rules suggest that these motifs are essential for the function of the cerebellar network.