Golgi Cells Research Articles

Stimulus-Dependent State Transition between Synchronized Oscillation and Randomly Repetitive Burst in a Model Cerebellar Granular Layer

Information processing of the cerebellar granular layer composed of granule and Golgi cells is regarded as an important first step toward the cerebellar computation. Our previous theoretical studies have shown that granule cells can exhibit random alternation between burst and silent modes, which provides a basis of population representation of the passage-of-time (POT) from the onset of external input stimuli. On the other hand, another computational study has reported that granule cells can exhibit synchronized oscillation of activity, as consistent with observed oscillation in local field potential recorded from the granular layer while animals keep still. Here we have a question of whether an identical network model can explain these distinct dynamics. In the present study, we carried out computer simulations based on a spiking network model of the granular layer varying two parameters: the strength of a current injected to granule cells and the concentration of Mg2+ which controls the conductance of NMDA channels assumed on the Golgi cell dendrites. The simulations showed that cells in the granular layer can switch activity states between synchronized oscillation and random burst-silent alternation depending on the two parameters. For higher Mg2+ concentration and a weaker injected current, granule and Golgi cells elicited spikes synchronously (synchronized oscillation state). In contrast, for lower Mg2+ concentration and a stronger injected current, those cells showed the random burst-silent alternation (POT-representing state). It is suggested that NMDA channels on the Golgi cell dendrites play an important role for determining how the granular layer works in response to external input.

Microlesions of the Inferior Olive Reduce Vestibular Modulation of Purkinje Cell Complex and Simple Spikes in Mouse Cerebellum

Cerebellar Purkinje cells have two distinct action potentials: complex spikes (CSs) are evoked by single climbing fibers that originate from the contralateral inferior olive. Simple spikes (SSs) are often ascribed to mossy fiber-granule cell-parallel fiber inputs to Purkinje cells. Although generally accepted, this view lacks experimental support. Vestibular stimulation independently activates primary afferent mossy fibers and tertiary afferent climbing fibers that project to the uvula-nodulus (folia 8-10). CSs and SSs normally discharge antiphasically during sinusoidal roll-tilt. When CSs increase, SSs decrease. We tested the relative independence of these pathways in mice by making electrolytic microlesions of the two inferior olivary nuclei from which vestibular climbing fibers originate; the β-nucleus and dorsomedial cell column. This reduced vestibular climbing fiber signaling to the contralateral folia 8-10, while leaving intact vestibular primary and secondary afferent mossy fibers. We recorded from Purkinje cells and interneurons in folia 8-10, identified by juxtacellular labeling with Neurobiotin. Microlesions of the inferior olive increased the spontaneous discharge of SSs in contralateral folia 8-10, but blocked their modulation during vestibular stimulation. The vestibularly evoked discharge of excitatory cerebellar interneurons (granule cells and unipolar brush cells) was not modified by olivary microlesions. The modulated discharge of stellate cells, but not Golgi cells, was reduced by olivary microlesions. We conclude that vestibular modulation of CSs and SSs depends on intact climbing fibers. The absence of vestibularly modulated SSs following olivary microlesions reflects the loss of climbing fiber-evoked stellate cell discharge.

Further characterization of autoantibodies to GABAergic neurons in the central nervous system produced by a subset of children with autism

BackgroundAutism is a neurodevelopmental disorder characterized by impairments in social interaction and deficits in verbal and nonverbal communication, together with the presence of repetitive behaviors or a limited repertoire of activities and interests. The causes of autism are currently unclear. In a previous study, we determined that 21% of children with autism have plasma autoantibodies that are immunoreactive with a population of neurons in the cerebellum that appear to be Golgi cells, which are GABAergic interneurons.MethodsWe have extended this analysis by examining plasma immunoreactivity in the remainder of the brain. To determine cell specificity, double-labeling studies that included one of the calcium-binding proteins that are commonly colocalized in GABAergic neurons (calbindin, parvalbumin or calretinin) were also carried out to determine which GABAergic neurons are immunoreactive. Coronal sections through the rostrocaudal extent of the macaque monkey brain were reacted with plasma from each of seven individuals with autism who had previously demonstrated positive Golgi cell staining, as well as six negative controls. In addition, brain sections from adult male mice were similarly examined.ResultsIn each case, specific staining was observed for neurons that had the morphological appearance of interneurons. By double-labeling sections with plasma and with antibodies directed against γ-aminobutyric acid (GABA), we determined that all autoantibody-positive neurons were GABAergic. However, not all GABAergic neurons were autoantibody-positive. Calbindin was colabeled in several of the autoantibody-labeled cells, while parvalbumin colabeling was less frequently observed. Autoantibody-positive cells rarely expressed calretinin. Sections from the mouse brain processed similarly to the primate sections also demonstrated immunoreactivity to interneurons distributed throughout the neocortex and many subcortical regions. Some cell populations stained in the primate (such as the Golgi neurons in the cerebellum) were not as robustly immunoreactive in the mouse brain.ConclusionsThese results suggest that the earlier report of autoantibody immunoreactivity to specific cells in the cerebellum extend to other regions of the brain. Further, these findings confirm the autoantibody-targeted cells to be a subpopulation of GABAergic interneurons. The potential impact of these autoantibodies on GABAergic disruption with respect to the etiology of autism is discussed herein.

Long-Term Gene-Silencing Effects of siRNA Introduced by Single-Cell Electroporation into Postmitotic CNS Neurons

To explore how long the gene-silencing effects of siRNA introduced into postmitotic neurons continue, we transferred siRNA against GFP into GFP-expressing Purkinje and Golgi cells in cerebellar cell cultures by single-cell electroporation. The temporal changes in the intensity of GFP fluorescence in the same electroporated cells were monitored in real time using GFP imaging. Under standard conditions, GFP fluorescence was reduced to under one-tenth of the initial levels 4-7days after electroporation. Such effects continued at least up to 14days after electroporation. The effects of siRNAs against endogenous genes also continued for the same period. Thus, this method could be an effective tool for silencing gene expression for a long period in postmitotic neurons.

Distribution of the auxiliary GABAB receptor subunits KCTD8, 12, 12b, and 16 in the mouse brain

AbstractGABAB receptors are the G‐protein‐coupled receptors for γ‐aminobutyric acid (GABA). KCTD8, 12, 12b, and 16 were recently identified as auxiliary GABAB receptor subunits and distinctly influence biophysical and pharmacological properties of the receptor response. Here we examined the expression patterns of the KCTDs in the mouse brain. Using in situ hybridization analysis, we found that most neurons express KCTD transcripts, supporting biochemical data showing that most GABAB receptors in the brain incorporate KCTD proteins. In the adult brain, KCTD12 and 16 have a widespread and KCTD8 and 12b a restricted expression pattern. Individual neurons can coexpress multiple KCTDs, as shown for granule cells and CA1/CA3 pyramidal cells in the hippocampus that coexpress KCTD12 and 16. In contrast, granule, Purkinje, and Golgi cells in the cerebellum selectively express one KCTD at a time. The expression levels of individual KCTD transcripts vary during postnatal brain development. Immunohistochemistry reveals that individual KCTD proteins can exhibit distinct axonal or dendritic localizations in neuronal populations. KCTDs are also detectable in nonneuronal tissues not expected to express GABAB receptors, suggesting that the role of KCTD proteins extends beyond GABAB receptors. In summary, our findings support that most brain GABAB receptors associate with KCTD proteins, but that the repertoire and abundance of KCTDs varies during development, among brain areas, neuronal populations, and at subcellular sites. We propose that the distinct spatial and temporal KCTD distribution patterns underlie functional differences in native GABAB responses. J. Comp. Neurol. 519:1435–1454, 2011. © 2011 Wiley‐Liss, Inc.

Distribution of the auxiliary GABAB receptor subunits KCTD8, 12, 12b, and 16 in the mouse brain

GABA(B) receptors are the G-protein-coupled receptors for γ-aminobutyric acid (GABA). KCTD8, 12, 12b, and 16 were recently identified as auxiliary GABA(B) receptor subunits and distinctly influence biophysical and pharmacological properties of the receptor response. Here we examined the expression patterns of the KCTDs in the mouse brain. Using in situ hybridization analysis, we found that most neurons express KCTD transcripts, supporting biochemical data showing that most GABA(B) receptors in the brain incorporate KCTD proteins. In the adult brain, KCTD12 and 16 have a widespread and KCTD8 and 12b a restricted expression pattern. Individual neurons can coexpress multiple KCTDs, as shown for granule cells and CA1/CA3 pyramidal cells in the hippocampus that coexpress KCTD12 and 16. In contrast, granule, Purkinje, and Golgi cells in the cerebellum selectively express one KCTD at a time. The expression levels of individual KCTD transcripts vary during postnatal brain development. Immunohistochemistry reveals that individual KCTD proteins can exhibit distinct axonal or dendritic localizations in neuronal populations. KCTDs are also detectable in nonneuronal tissues not expected to express GABA(B) receptors, suggesting that the role of KCTD proteins extends beyond GABA(B) receptors. In summary, our findings support that most brain GABA(B) receptors associate with KCTD proteins, but that the repertoire and abundance of KCTDs varies during development, among brain areas, neuronal populations, and at subcellular sites. We propose that the distinct spatial and temporal KCTD distribution patterns underlie functional differences in native GABA(B) responses.

Functional circuitry of a unique cerebellar specialization: the valvula cerebelli of a mormyrid fish

The valvula cerebelli of the mormyrid electric fish is a useful site for the study of cerebellar function. The valvula forms a part of the electrosensory-electromotor system of this fish, a system that offers many possibilities for the study of sensory-motor integration. The valvula also has a number of histological features not present in mammals which facilitate investigation of cerebellar circuitry and its plasticity. This initial study characterizes the basic physiology and pharmacology of cells in the valvula using an in vitro slice preparation. Intrinsic properties and synaptic responses of Purkinje cells and other cell types were examined. We found that Purkinje cells fire a small narrow Na + spike and a large broad Ca 2+ spike, generated in the axon initial segment and dendritic-soma region, respectively. Purkinje cells respond to parallel fiber inputs with graded excitatory postsynaptic potentials (EPSPs) and to climbing fiber inputs with all-or-none EPSPs. Efferent cells, Golgi cells, and deep stellate cells all fire a single type of large narrow spike and respond only to parallel fiber inputs. Both parallel fiber and climbing fiber responses in Purkinje cells appear to be entirely mediated by AMPA-type glutamate receptors, whereas parallel fiber responses in efferent cells and stellate cells include AMPA and NMDA components. In addition, a strong synaptic inhibition was uncovered in both Purkinje cells and efferent cells in response to the focal stimulation of parallel fibers. Dual cell recordings indicate that deep stellate cells contribute at least partially to this inhibition. We conclude that despite its unique histology, the local functional circuitry of the mormyrid valvula cerebelli is largely similar to that of the mammalian cerebellum. Thus, what is learned concerning the functioning of the mormyrid valvula cerebelli may be expected to be informative about cerebellar function in general.

小脳皮質モデル (ゴルジ細胞-顆粒細胞間の返回抑制回路網) の空間周波数特性

小脳皮質モデル (ゴルジ細胞-顆粒細胞間の返回抑制回路網) の空間周波数特性

Robustness effect of gap junctions between Golgi cells on cerebellar cortex oscillations

BackgroundPrevious one-dimensional network modeling of the cerebellar granular layer has been successfully linked with a range of cerebellar cortex oscillations observed in vivo. However, the recent discovery of gap junctions between Golgi cells (GoCs), which may cause oscillations by themselves, has raised the question of how gap-junction coupling affects GoC and granular-layer oscillations. To investigate this question, we developed a novel two-dimensional computational model of the GoC-granule cell (GC) circuit with and without gap junctions between GoCs.ResultsIsolated GoCs coupled by gap junctions had a strong tendency to generate spontaneous oscillations without affecting their mean firing frequencies in response to distributed mossy fiber input. Conversely, when GoCs were synaptically connected in the granular layer, gap junctions increased the power of the oscillations, but the oscillations were primarily driven by the synaptic feedback loop between GoCs and GCs, and the gap junctions did not change oscillation frequency or the mean firing rate of either GoCs or GCs.ConclusionOur modeling results suggest that gap junctions between GoCs increase the robustness of cerebellar cortex oscillations that are primarily driven by the feedback loop between GoCs and GCs. The robustness effect of gap junctions on synaptically driven oscillations observed in our model may be a general mechanism, also present in other regions of the brain.

TrkB (Tropomyosin-Related Kinase B) Controls the Assembly and Maintenance of GABAergic Synapses in the Cerebellar Cortex

Inhibitory interneurons play a critical role in coordinating the activity of neural circuits. To explore the mechanisms that direct the organization of inhibitory circuits, we analyzed the involvement of tropomyosin-related kinase B (TrkB) in the assembly and maintenance of GABAergic inhibitory synapses between Golgi and granule cells in the mouse cerebellar cortex. We show that TrkB acts directly within each cell-type to regulate synaptic differentiation. TrkB is required not only for assembly, but also maintenance of these synapses and acts, primarily, by regulating the localization of synaptic constituents. Postsynaptically, TrkB controls the localization of a scaffolding protein, gephyrin, but acts at a step subsequent to the localization of a cell adhesion molecule, Neuroligin-2. Importantly, TrkB is required for the localization of an Ig superfamily cell adhesion molecule, Contactin-1, in Golgi and granule cells and the absence of Contactin-1 also results in deficits in inhibitory synaptic development. Thus, our findings demonstrate that TrkB controls the assembly and maintenance of GABAergic synapses and suggest that TrkB functions, in part, through promoting synaptic adhesion.

A Subtraction Mechanism of Temporal Coding in Cerebellar Cortex

The temporally specific learning displayed by the cerebellum facilitates mechanistic analysis of neural timing and temporal coding. We report evidence for a subtraction-like mechanism of temporal coding in cerebellar cortex in which activity in a subset of granule cells specifically codes the interval between the offset of two mossy fiber inputs. In a large-scale cerebellar simulation, cessation of one of two ongoing mossy fiber inputs produces a robust temporal code in the population of granule cells. This activity supports simulation learning in response to temporal patterns of stimuli, even when those same stimuli do not support learning when presented individually. Using stimulation of mossy fiber inputs to the cerebellum as training stimuli in rabbits, we confirmed these unusual predictions in a cerebellum-dependent form of learning. Analysis of the simulations reveals a specific working hypothesis for this temporal subtraction process that involves interactions between granule cells and the inhibitory Golgi cells. The results suggest how feedforward inhibition, such as that present in the cerebellar cortex, can contribute to temporal coding.

Current source density correlates of cerebellar Golgi and Purkinje cell responses to tactile input

The overall circuitry of the cerebellar cortex has been known for over a century, but the function of many synaptic connections remains poorly characterized in vivo. We used a one-dimensional multielectrode probe to estimate the current source density (CSD) of Crus IIa in response to perioral tactile stimuli in anesthetized rats and to correlate current sinks and sources to changes in the spike rate of corecorded Golgi and Purkinje cells. The punctate stimuli evoked two distinct early waves of excitation (at <10 and ∼ 20 ms) associated with current sinks in the granular layer. The second wave was putatively of corticopontine origin, and its associated sink was located higher in the granular layer than the first trigeminal sink. The distinctive patterns of granular-layer sinks correlated with the spike responses of corecorded Golgi cells. In general, Golgi cell spike responses could be linearly reconstructed from the CSD profile. A dip in simple-spike activity of coregistered Purkinje cells correlated with a current source deep in the molecular layer, probably generated by basket cell synapses, interspersed between sparse early sinks presumably generated by synapses from granule cells. The late (>30 ms) enhancement of simple-spike activity in Purkinje cells was characterized by the absence of simultaneous sinks in the granular layer and by the suppression of corecorded Golgi cell activity, pointing at inhibition of Golgi cells by Purkinje axon collaterals as a likely mechanism of late Purkinje cell excitation.

Spontaneous Activity Signatures of Morphologically Identified Interneurons in the Vestibulocerebellum

Cerebellar cortical interneurons such as Golgi cells, basket cells, stellate cells, unipolar brush cells, and granule cells play an essential role in the operations of the cerebellum. However, detailed functional studies of the activity of these cells in both anesthetized and behaving animals have been hampered by problems in recognizing their physiological signatures. We have extracellularly recorded the spontaneous activity of vestibulocerebellar interneurons in ketamine/xylazine-anesthetized rats and subsequently labeled them with Neurobiotin using the juxtacellular technique. After recovery and morphological identification of these cells, they were related to statistical measures of their spontaneous activity. Golgi cells display a somewhat irregular firing pattern with relatively low average frequencies. Unipolar brush cells are characterized by more regular firing at higher rates. Basket and stellate cells are alike in their firing characteristics, which mainly stand out by their irregularity; some of them are set apart by their very slow average rate. The spontaneous activity of interneurons examined in the ketamine/xylazine rabbit fit within this general pattern. In the rabbit, granule cells were identified by the spontaneous occurrence of extremely high-frequency bursts of action potentials, which were also recognized in the rat. On the basis of these observations, we devised an algorithm that reliably determined the identity of 75% of the cells with only 2% incorrect classifications. The remaining cells were placed into border categories within which no classification was attempted. We propose that this algorithm can be used to help classify vestibulocerebellar interneurons recorded in awake, behaving animals.

Zac1 plays a key role in the development of specific neuronal subsets in the mouse cerebellum

BackgroundThe cerebellum is composed of a diverse array of neuronal subtypes. Here we have used a candidate approach to identify Zac1, a tumor suppressor gene encoding a zinc finger transcription factor, as a new player in the transcriptional network required for the development of a specific subset of cerebellar nuclei and a population of Golgi cells in the cerebellar cortex.ResultsWe found that Zac1 has a complex expression profile in the developing cerebellum, including in two proliferating progenitor populations; the cerebellar ventricular zone and the external granular layer overlying posterior cerebellar lobules IX and X. Zac1 is also expressed in some postmitotic cerebellar neurons, including a subset of GABAergic interneurons in the medial cerebellar nuclei. Notably, GABAergic interneurons in the cerebellar nuclei are derived from the cerebellar ventricular zone, where Zac1 is also expressed, consistent with a lineage relationship between these two Zac1+ populations. Zac1 is also expressed in a small subset of cells in the posterior vermis, including some neurogranin-immunoreactive (NG+) Golgi cells, which, based on short-term birthdating, are derived from the EGL, where Zac1 is also expressed. However, Zac1+ cells and NG+ Golgi cells in the cerebellar cortex also display unique properties, as they are generated within different, albeit overlapping, time windows. Finally, consistent with the expression profile of Zac1, two conspicuous abnormalities were found in the cerebellum of Zac1 null mice: the medial cerebellar nuclei, and not the others, were significantly reduced in size; and the number of Golgi cells in cerebellar lobule IX was reduced by approximately 60% compared to wild-type littermates.ConclusionsThe data presented here indicate that the tumor suppressor gene Zac1 is expressed in a complex fashion in the developing cerebellum, including in two dividing progenitor populations and in specific subsets of postmitotic neurons, including Golgi cells and GABAergic neurons in the medial nuclei, which require Zac1 for their differentiation. We thus conclude that Zac1 is a critical regulator of normal cerebellar development, adding a new transcriptional regulator to the growing list of factors involved in generating neuronal diversity in the developing cerebellum.

Characterization of electrical coupling between cerebellar Golgi cells

Characterization of electrical coupling between cerebellar Golgi cells

Golgi Cells Operate as State-Specific Temporal Filters at the Input Stage of the Cerebellar Cortex

Cerebellar processing of incoming information begins at the synapse between mossy fibers and granule cells, a synapse that is strongly controlled through Golgi cell inhibition. Thus, Golgi cells are uniquely positioned to control the flow of information into the cerebellar cortex and understanding their responses during behavior is essential to understanding cerebellar function. Here we show, for the first time, that Golgi cells express a unique oculomotor-related signal that can be used to provide state- and time-specific filtering of granule cell activity. We used newly established criteria to identify the unique electrophysiological signature of Golgi cells and recorded these neurons in the squirrel monkey ventral paraflocculus during oculomotor behaviors. We found that they carry eye movement, but not vestibular or visual, information and that this eye movement information is only expressed within a specific range of eye positions for each neuron. In addition, simultaneous recordings of Golgi cells and nearby mossy fibers revealed that Golgi cells have the opposite directional tuning of the mossy fiber(s) that likely drive their responses, and that these responses are more sluggish than their mossy fiber counterparts. Because the mossy fiber inputs appear to convey the activity of burst-tonic neurons in the brainstem, Golgi cell responses reflect a time-filtered negative image of the motor command sent to the extraocular muscles. We suggest a role for Golgi cells in the construction of forward models of movement, commonly hypothesized as a major function of the cerebellar cortex in motor control.

Discovery and rediscoveries of Golgi cells

When Camillo Golgi invented the black reaction in 1873 and first described the fine anatomical structure of the nervous system, he described a ‘big nerve cell’ that later took his name, the Golgi cell of cerebellum (‘Golgi’schen Zellen’, Gustaf Retzius, 1892). The Golgi cell was then proposed as the prototype of type-II interneurons, which form complex connections and exert their actions exclusively within the local network. Santiago Ramón y Cajal (who received the Nobel Prize with Golgi in 1906) proceeded to a detailed description of Golgi cell morphological characteristics, but functional insight remained very limited for many years. The first rediscovery happened in the 1960s, when neurophysiological analysis in vivo revealed that Golgi cells are inhibitory interneurons. This finding promoted the development of two major cerebellar theories, the ‘beam theory’ of John Eccles and the ‘motor learning theory’ of David Marr, in which the Golgi cells regulate the spatial organisation and the gain of input signals to be processed and learned by the cerebellar circuit. However, the matter was not set and a series of pioneering observations using single unit recordings and electronmicroscopy raised new issues that could not be fully explored until the 1990s. Then, the advent of new electrophysiological and imaging techniques in vitro and in vivo demonstrated the cellular and network activities of these neurons. Now we know that Golgi cells, through complex systems of chemical and electrical synapses, effectively control the spatio-temporal organisation of cerebellar responses. The Golgi cells regulate the timing and number of spikes emitted by granule cells and coordinate their coherent activity. Moreover, the Golgi cells regulate the induction of long-term synaptic plasticity along the mossy fibre pathway. Eventually, the Golgi cells transform the granular layer of cerebellum into an adaptable spatio-temporal filter capable of performing several kinds of logical operation. After more than a century, Golgi’s intuition that the Golgi cell had to generate under a new perspective complex ensemble effects at the network level has finally been demonstrated.

Long-Term Depression at Parallel Fiber to Golgi Cell Synapses

Golgi cells (GoCs) are the primary inhibitory interneurons of the granular layer of the cerebellum. Their inhibition of granule cells is central to operate the relay of excitatory inputs to the cerebellar cortex. Parallel fibers (PFs) establish synapses to the GoCs in the molecular layer; these synapses contain AMPA, N-methyl-D-aspartate (NMDA), and mostly group II metabotropic glutamate receptors. Long-term changes in the efficacy of synaptic transmission at the PF-GoC synapse have not been described previously. We used whole cell patch-clamp recordings of GoCs in acute rat cerebellar slices to study synaptic plasticity. We report that high-frequency burst stimulation of PFs, using a current-clamp or voltage-clamp induction protocol, gave rise to long-term depression (LTD) at the PF-GoC synapse. This form of LTD was not associated with persistent changes of paired-pulse ratio, suggesting a postsynaptic origin. Furthermore, LTD induction was not dependent on activation of NMDA receptors. PF-GoC LTD does require activation of specifically group II metabotropic glutamate receptors and of protein kinase A.

Enhanced Functions of Electrical Junctions

Electrical synapses and synchrony are nearly synonymous. In this issue of Neuron, Vervaeke et al. broaden this longstanding association. They found that in the Golgi cell network of the cerebellum, electrical synapses synchronize resting activity, and cause surround inhibition and desynchronization in response to excitatory input.

Rapid Desynchronization of an Electrically Coupled Interneuron Network with Sparse Excitatory Synaptic Input

SummaryElectrical synapses between interneurons contribute to synchronized firing and network oscillations in the brain. However, little is known about how such networks respond to excitatory synaptic input. To investigate this, we studied electrically coupled Golgi cells (GoC) in the cerebellar input layer. We show with immunohistochemistry, electron microscopy, and electrophysiology that Connexin-36 is necessary for functional gap junctions (GJs) between GoC dendrites. In the absence of coincident synaptic input, GoCs synchronize their firing. In contrast, sparse, coincident mossy fiber input triggered a mixture of excitation and inhibition of GoC firing and spike desynchronization. Inhibition is caused by propagation of the spike afterhyperpolarization through GJs. This triggers network desynchronization because heterogeneous coupling to surrounding cells causes spike-phase dispersion. Detailed network models predict that desynchronization is robust, local, and dependent on synaptic input properties. Our results show that GJ coupling can be inhibitory and either promote network synchronization or trigger rapid network desynchronization depending on the synaptic input.