Engrailed transcription factors direct excitatory cerebellar neuron diversity and survival.

The neurons of the three cerebellar nuclei (CN) are the primary output neurons of the cerebellum. The excitatory neurons (e) of the medial (m) CN (eCNm) were recently divided into molecularly defined subdomains in the adult; however, how they are established during development is not known. We define molecular subdomains of the mouse embryonic eCNm using single-cell RNA-sequencing and spatial expression analysis, showing that they evolve during embryogenesis to prefigure the adult. Furthermore, eCNm are transcriptionally divergent from cells in the other nuclei by embryonic day 14.5. We previously showed that loss of the homeobox genes En1 and En2 leads to loss of approximately half of the embryonic eCNm. We demonstrate that mutation of En1/2 in the embryonic eCNm results in death of specific posterior eCNm molecular subdomains and downregulation of TBR2 (EOMES) in an anterior embryonic subdomain, as well as reduced synaptic gene expression. We further reveal a similar function for EN1/2 in mediating TBR2 expression, neuron differentiation and survival in the other excitatory neurons (granule and unipolar brush cells). Thus, our work defines embryonic eCNm molecular diversity and reveals conserved roles for EN1/2 in the cerebellar excitatory neuron lineage.

Reverse-translational identification of a cerebellar satiation network.

The brain is the seat of body weight homeostasis. However, our inability to control the increasing prevalence of obesity highlights a need to look beyond canonical feeding pathways to broaden our understanding of body weight control1-3. Here we used a reverse-translational approach to identify and anatomically, molecularly and functionally characterize a neural ensemble that promotes satiation. Unbiased, task-based functional magnetic resonance imaging revealed marked differences in cerebellar responses to food in people with a genetic disorder characterized by insatiable appetite. Transcriptomic analyses in mice revealed molecularly and topographically -distinct neurons in the anterior deep cerebellar nuclei (aDCN) that are activated by feeding or nutrient infusion in the gut. Selective activation of aDCN neurons substantially decreased food intake by reducing meal size without compensatory changes to metabolic rate. We found that aDCN activity terminates food intake by increasing striatal dopamine levels and attenuating the phasic dopamine response to subsequent food consumption. Our study defines a conserved satiation centre that may represent a novel therapeutic target for the management of excessive eating, and underscores the utility of a 'bedside-to-bench' approach for the identification of neural circuits that influence behaviour.

A spatial-temporal map of glutamatergic neurogenesis in the murine embryonic cerebellar nuclei uncovers a high degree of cellular heterogeneity.

The nuclei are the main output structures of the cerebellum. Each and every cerebellar cortical computation reaches several areas of the brain by means of cerebellar nuclei processing and integration. Nevertheless, our knowledge of these structures is still limited compared to the cerebellar cortex. Here, we present a mouse genetic inducible fate-mapping study characterizing rhombic lip-derived glutamatergic neurons of the nuclei, the most conspicuous family of long-range cerebellar efferent neurons. Glutamatergic neurons mainly occupy dorsal and lateral territories of the lateral and interposed nuclei, as well as the entire medial nucleus. In mice, they are born starting from about embryonic day 9.5, with a peak between 10.5 and 12.5, and invade the nuclei with a lateral-to-medial progression. While some markers label a heterogeneous population of neurons sharing a common location (BRN2), others appear to be lineage specific (TBR1, LMX1a, and MEIS2). A comparative analysis of TBR1 and LMX1a distributions reveals an incomplete overlap in their expression domains, in keeping with the existence of separate efferent subpopulations. Finally, some tagged glutamatergic progenitors are not labeled by any of the markers used in this study, disclosing further complexity. Taken together, our results obtained in late embryonic nuclei shed light on the heterogeneity of the excitatory neuron pool, underlying the diversity in connectivity and functions of this largely unexplored cerebellar territory. Our findings contribute to laying the groundwork for a comprehensive functional analysis of nuclear neuron subpopulations.

Catecholaminergic Innervation of the Lateral Nucleus of the Cerebellum Modulates Cognitive Behaviors.

The cerebellum processes neural signals related to rewarding and aversive stimuli, suggesting that the cerebellum supports nonmotor functions in cognitive and emotional domains. Catecholamines are a class of neuromodulatory neurotransmitters well known for encoding such salient stimuli. Catecholaminergic modulation of classical cerebellar functions have been demonstrated. However, a role for cerebellar catecholamines in modulating cerebellar nonmotor functions is unknown. Using biochemical methods in male mice, we comprehensively mapped TH+ fibers throughout the entire cerebellum and known precerebellar nuclei. Using electrochemical (fast scan cyclic voltammetry), and viral/genetic methods to selectively delete Th in fibers innervating the lateral cerebellar nucleus (LCN), we interrogated sources and functional roles of catecholamines innervating the LCN, which is known for its role in supporting cognition. The LCN has the most TH+ fibers in cerebellum, as well as the most change in rostrocaudal expression among the cerebellar nuclei. Norepinephrine is the major catecholamine measured in LCN. Distinct catecholaminergic projections to LCN arise only from locus coeruleus, and a subset of Purkinje cells that are positive for staining of TH. LC stimulation was sufficient to produce catecholamine release in LCN. Deletion of Th in fibers innervating LCN (LCN-Th-cKO) resulted in impaired sensorimotor integration, associative fear learning, response inhibition, and working memory in LCN-Th-cKO mice. Strikingly, selective inhibition of excitatory LCN output neurons with inhibitory designer receptor exclusively activated by designer drugs led to facilitation of learning on the same working memory task impaired in LCN-Th-cKO mice. Collectively, these data demonstrate a role for LCN catecholamines in cognitive behaviors.SIGNIFICANCE STATEMENT Here, we report on interrogating sources and functional roles of catecholamines innervating the lateral nucleus of the cerebellum (LCN). We map and quantify expression of TH, the rate-limiting enzyme in catecholamine synthesis, in the entire cerebellar system, including several precerebellar nuclei. We used cyclic voltammetry and pharmacology to demonstrate sufficiency of LC stimulation to produce catecholamine release in LCN. We used advanced viral techniques to map and selectively KO catecholaminergic neurotransmission to the LCN, and characterized significant cognitive deficits related to this manipulation. Finally, we show that inhibition of excitatory LCN neurons with designer receptor exclusively activated by designer drugs, designed to mimic Gi-coupled catecholamine GPCR signaling, results in facilitation of a working memory task impaired in LCN-specific TH KO mice.

Differential Coding Strategies in Glutamatergic and GABAergic Neurons in the Medial Cerebellar Nucleus.

The cerebellum drives motor coordination and sequencing of actions at the millisecond timescale through adaptive control of cerebellar nuclear output. Cerebellar nuclei integrate high-frequency information from both the cerebellar cortex and the two main excitatory inputs of the cerebellum: the mossy fibers and the climbing fiber collaterals. However, how nuclear cells process rate and timing of inputs carried by these inputs is still debated. Here, we investigate the influence of the cerebellar cortical output, the Purkinje cells, on identified cerebellar nuclei neurons in vivo in male mice. Using transgenic mice expressing Channelrhodopsin2 specifically in Purkinje cells and tetrode recordings in the medial nucleus, we identified two main groups of neurons based on the waveform of their action potentials. These two groups of neurons coincide with glutamatergic and GABAergic neurons identified by optotagging after Chrimson expression in VGLUT2-cre and GAD-cre mice, respectively. The glutamatergic-like neurons fire at high rate and respond to both rate and timing of Purkinje cell population inputs, whereas GABAergic-like neurons only respond to the mean population firing rate of Purkinje cells at high frequencies. Moreover, synchronous activation of Purkinje cells can entrain the glutamatergic-like, but not the GABAergic-like, cells over a wide range of frequencies. Our results suggest that the downstream effect of synchronous and rhythmic Purkinje cell discharges depends on the type of cerebellar nuclei neurons targeted.SIGNIFICANCE STATEMENT Motor coordination and skilled movements are driven by the permanent discharge of neurons from the cerebellar nuclei that communicate cerebellar computation to other brain areas. Here, we set out to study how specific subtypes of cerebellar nuclear neurons of the medial nucleus are controlled by Purkinje cells, the sole output of the cerebellar cortex. We could isolate different subtypes of nuclear cell that differentially encode Purkinje cell inhibition. Purkinje cell stimulation entrains glutamatergic projection cells at their firing frequency, whereas GABAergic neurons are only inhibited. These differential coding strategies may favor temporal precision of cerebellar excitatory outputs associated with specific features of movement control while setting the global level of cerebellar activity through inhibition via rate coding mechanisms.

A Slow Short-Term Depression at Purkinje to Deep Cerebellar Nuclear Neuron Synapses Supports Gain-Control and Linear Encoding over Second-Long Time Windows.

Modifications in the sensitivity of neural elements allow the brain to adapt its functions to varying demands. Frequency-dependent short-term synaptic depression (STD) provides a dynamic gain-control mechanism enabling adaptation to different background conditions alongside enhanced sensitivity to input-driven changes in activity. In contrast, synapses displaying frequency-invariant transmission can faithfully transfer ongoing presynaptic rates enabling linear processing, deemed critical for many functions. However, rigid frequency-invariant transmission may lead to runaway dynamics and low sensitivity to changes in rate. Here, I investigated the Purkinje cell to deep cerebellar nuclei neuron synapses (PC_DCNs), which display frequency invariance, and yet, PCs maintain background activity at disparate rates, even at rest. Using protracted PC_DCN activation (120 s) to mimic background activity in cerebellar slices from mature mice of both sexes, I identified a previously unrecognized, frequency-dependent, slow STD (S-STD), adapting IPSC amplitudes in tens of seconds to minutes. However, after changes in activation rates, over a behavior-relevant second-long time window, S-STD enabled scaled linear encoding of PC rates in synaptic charge transfer and DCN spiking activity. Combined electrophysiology, optogenetics, and statistical analysis suggested that S-STD mechanism is input-specific, involving decreased ready-to-release quanta, and distinct from faster short-term plasticity (f-STP). Accordingly, an S-STD component with a scaling effect (i.e., activity-dependent release sites inactivation), extending a model explaining PC_DCN release on shorter timescales using balanced f-STP, reproduced the experimental results. Thus, these results elucidates a novel slow gain-control mechanism able to support linear transfer of behavior-driven/learned PC rates concurrently with background activity adaptation, and furthermore, provides an alternative pathway to refine PC output.SIGNIFICANCE STATEMENT The brain can adapt to varying demands by dynamically changing the gain of its synapses; however, some tasks require ongoing linear transfer of presynaptic rates, seemingly incompatible with nonlinear gain adaptation. Here, I report a novel slow gain-control mechanism enabling scaled linear encoding of presynaptic rates over behavior-relevant time windows, and adaptation to background activity at the Purkinje to deep cerebellar nuclear neurons synapses (PC_DCNs). A previously unrecognized PC_DCNs slow and frequency-dependent short-term synaptic depression (S-STD) mediates this process. Experimental evidence and simulations suggested that scaled linear encoding emerges from the combination of S-STD slow dynamics and frequency-invariant transmission at faster timescales. These results demonstrate a mechanism reconciling rate code with background activity adaptation and suitable for flexibly tuning PCs output via background activity modulation.

State-dependent entrainment of cerebellar nuclear neurons to the local field potential during voluntary movements.

Understanding the relationship between the local field potential (LFP) and single neurons is essential if we are to understand network dynamics and the entrainment of neuronal activity. Here, we investigated the interaction between the LFP and single neurons recorded in the rat cerebellar nuclei (CN), which are part of the sensorimotor network, in freely moving rats. During movement, the LFP displayed persistent oscillations in the theta band frequency, whereas CN neurons displayed intermittent oscillations in the same frequency band contingent on the instantaneous LFP power; the neurons oscillated primarily when the concurrent LFP power was either high or low. Quantification of the relative instantaneous frequency and phase locking showed that CN neurons exhibited phase locked rhythmic activity at a frequency similar to that of the LFP or at a shifted frequency during high and low LFP power, respectively. We suggest that this nonlinear interaction between cerebellar neurons and the LFP power, which occurs solely during movement, contributes to the shaping of cerebellar output patterns.NEW & NOTEWORTHY We studied the interaction between single neurons and the LFP in the cerebellar nuclei of freely moving rats. We show that during movement, the neurons oscillated in the theta frequency band contingent on the concurrent LFP oscillation power in the same band; the neurons oscillated primarily when the LFP power was either high or low. We are the first to demonstrate a nonlinear, state-dependent entrainment of single neurons to the LFP.

Encoding of action by the Purkinje cells of the cerebellum.

Execution of accurate eye movements depends critically on the cerebellum, suggesting that the major output neurons of the cerebellum, Purkinje cells, may predict motion of the eye. However, this encoding of action for rapid eye movements (saccades) has remained unclear: Purkinje cells show little consistent modulation with respect to saccade amplitude or direction, and critically, their discharge lasts longer than the duration of a saccade. Here we analysed Purkinje-cell discharge in the oculomotor vermis of behaving rhesus monkeys (Macaca mulatta) and found neurons that increased or decreased their activity during saccades. We estimated the combined effect of these two populations via their projections to the caudal fastigial nucleus, and uncovered a simple-spike population response that precisely predicted the real-time motion of the eye. When we organized the Purkinje cells according to each cell's complex-spike directional tuning, the simple-spike population response predicted both the real-time speed and direction of saccade multiplicatively via a gain field. This suggests that the cerebellum predicts the real-time motion of the eye during saccades via the combined inputs of Purkinje cells onto individual nucleus neurons. A gain-field encoding of simple spikes emerges if the Purkinje cells that project onto a nucleus neuron are not selected at random but share a common complex-spike property.

New Cerebello-Cortical Pathway Involved in Higher-Order Oculomotor Control.

The cerebellum and the basal ganglia play an important role in the control of voluntary eye movement associated with complex behavior, but little is known about how cerebellar projections project to cortical eye movement areas. Here we used retrograde transneuronal transport of rabies virus to identify neurons in the cerebellar nuclei that project via the thalamus to supplementary eye field (SEF) of the frontal cortex of macaques. After rabies injections into the SEF, many neurons in the restricted region, the ventral aspects of the dentate nucleus (DN), the caudal pole of the DN, and the posterior interpositus nucleus (PIN) were labeled disynaptically via the thalamus, whereas no neuron labeling was found in the anterior interpositus nucleus (AIN). The distribution of the labeled neurons was dorsoventrally different from that of DN and PIN neurons labeled from the motor cortex. In the basal ganglia, a large number of labeled neurons were confined to the dorsomedial portion of the internal segment of the globus pallidus (GPi) as more neurons were labeled in the inner portion of the GPi (GPii) than in the outer portion of the GPi (GPio). This is the first evidence of a projection between cerebellum/basal ganglia and the SEF that could enable the cerebellum to modulate the cognitive control of voluntary eye movement.

Role of Rostral Fastigial Neurons in Encoding a Body-Centered Representation of Translation in Three Dimensions.

Many daily behaviors rely critically on estimates of our body motion. Such estimates must be computed by combining neck proprioceptive signals with vestibular signals that have been transformed from a head- to a body-centered reference frame. Recent studies showed that deep cerebellar neurons in the rostral fastigial nucleus (rFN) reflect these computations, but whether they explicitly encode estimates of body motion remains unclear. A key limitation in addressing this question is that, to date, cell tuning properties have only been characterized for a restricted set of motions across head-re-body orientations in the horizontal plane. Here we examined, for the first time, how 3D spatiotemporal tuning for translational motion varies with head-re-body orientation in both horizontal and vertical planes in the rFN of male macaques. While vestibular coding was profoundly influenced by head-re-body position in both planes, neurons typically reflected at most a partial transformation. However, their tuning shifts were not random but followed the specific spatial trajectories predicted for a 3D transformation. We show that these properties facilitate the linear decoding of fully body-centered motion representations in 3D with a broad range of temporal characteristics from small groups of 5-7 cells. These results demonstrate that the vestibular reference frame transformation required to compute body motion is indeed encoded by cerebellar neurons. We propose that maintaining partially transformed rFN responses with different spatiotemporal properties facilitates the creation of downstream body motion representations with a range of dynamic characteristics, consistent with the functional requirements for tasks such as postural control and reaching.SIGNIFICANCE STATEMENT Estimates of body motion are essential for many daily activities. Vestibular signals are important contributors to such estimates but must be transformed from a head- to a body-centered reference frame. Here, we provide the first direct demonstration that the cerebellum computes this transformation fully in 3D. We show that the output of these computations is reflected in the tuning properties of deep cerebellar rostral fastigial nucleus neurons in a specific distributed fashion that facilitates the efficient creation of body-centered translation estimates with a broad range of temporal properties (i.e., from acceleration to position). These findings support an important role for the rostral fastigial nucleus as a source of body translation estimates functionally relevant for behaviors ranging from postural control to perception.

Developmental Changes in Serotonergic Modulation of GABAergic Synaptic Transmission and Postsynaptic GABAA Receptor Composition in the Cerebellar Nuclei.

Outputs from the cerebellar nuclei (CN) are important for generating and controlling movement. The activity of CN neurons is controlled not only by excitatory inputs from mossy and climbing fibers and by γ-aminobutyric acid (GABA)-based inhibitory transmission from Purkinje cells in the cerebellar cortex but is also modulated by inputs from other brain regions, including serotonergic fibers that originate in the dorsal raphe nuclei. We examined the modulatory effects of serotonin (5-HT) on GABAergic synapses during development, using rat cerebellar slices. As previously reported, 5-HT presynaptically decreased the amplitudes of stimulation-evoked inhibitory postsynaptic currents (IPSCs) in CN neurons, with this effect being stronger in slices from younger animals (postnatal days [P] 11-13) than in slices from older animals (P19-21). GABA release probabilities accordingly exhibited significant decreases from P11-13 to P19-21. Although there was a strong correlation between the GABA release probability and the magnitude of 5-HT-induced inhibition, manipulating the release probability by changing extracellular Ca2+ concentrations failed to control the extent of 5-HT-induced inhibition. We also found that the IPSCs exhibited slower kinetics at P11-13 than at P19-21. Pharmacological and molecular biological tests revealed that IPSC kinetics were largely determined by the prevalence of α1 subunits within GABAA receptors. In summary, pre- and postsynaptic developmental changes in serotonergic modulation and GABAergic synaptic transmission occur during the second to third postnatal weeks and may significantly contribute to the formation of normal adult cerebellar function.

Translaminar neuromorphotopological clustering and classification of dentate nucleus neurons.

Thisstudy aims to determine whether dentate neurons can be translaminarlyneuromorphotopologically classified as ventrolateral or dorsomedial type. Adulthuman dentate interneuron 2D binary images are analyzed. The analysis isperformed on both real and virtual neuron samples and 29 parameters are used.They are divided into the classes: neuron surface, shape, length, branching andcomplexity. Clustering is performed by an algorithm that employs predictor extraction (matrix attractor analysis/non-negative matrix factorization and cluster analysis of predictor factors - separate unifactor analysis/Student's t-test and MANOVA) and multivariate cluster analysis (cluster analysis, principal component analysis, factor analysis with pro/varimax rotation, Fisher's linear discriminant analysis and feed-forward backpropagation artificial neural networks). The separate unifactor analysis extracted as significant the following predictors from the natural cell sample: the Npd (p< 0:05), and from the virtual cell sample: the Adt (p< 0.05),Do (p< 0.001), Ms (p< 0.01), Dwdth (p< 0:001), Npd (p< 0:05), Nsd (p< 0.001), Nt/hod (p< 0.001), Nmax (p< 0.01), Ds (p< 0.001), Cdf (Nt/hod)st (p< 0.05). For the multidimensional analysis, with the exception of the Fisher's linear discriminant analysis which gave a false positive result, all other analyses rejected the translaminar dentate neuron classification. Thus, dentate neurons cannot be classified into ventrolateral/dorsomedial neuromorphotopological subtypes. Although some differences were found to exist, they are not sufficient to carry this classification. The methods of multidimensional statistical analysis are again shown to be the best for such kinds of analysis.

Synaptic excitation by climbing fibre collaterals in the cerebellar nuclei of juvenile and adult mice.

KEY POINTS: The inferior olive sends instructive motor signals to the cerebellum via the climbing fibre projection, which sends collaterals directly to large premotor neurons of the mouse cerebellar nuclei (CbN cells). Optogenetic activation of inferior olivary axons in vitro evokes EPSCs in CbN cells of several hundred pA to more than 1 nA. The inputs are three-fold larger at younger ages, 12 to 14 days old, than at 2 months old, suggesting a strong functional role for this pathway earlier in development. The EPSCs are multipeaked, owing to burst firing in several olivary afferents that fire asynchronously. The convergence of climbing fibre collaterals onto CbN cells decreases from ∼40 to ∼8, which is consistent with the formation of closed-loop circuits in which each CbN neuron receives input from 4-7 collaterals from inferior olivary neurons as well as from all 30-50 Purkinje cells that are innervated by those olivary neurons. ABSTRACT: The inferior olive conveys instructive signals to the cerebellum that drive sensorimotor learning. Inferior olivary neurons transmit their signals via climbing fibres, which powerfully excite Purkinje cells, evoking complex spikes and depressing parallel fibre synapses. Additionally, however, these climbing fibres send collaterals to the cerebellar nuclei (CbN). In vivo and in vitro data suggest that climbing fibre collateral excitation is weak in adult mice, raising the question of whether the primary role of this pathway may be developmental. We therefore examined climbing fibre collateral input to large premotor CbN cells over development by virally expressing channelrhodopsin in the inferior olive. In acute cerebellar slices from postnatal day (P)12-14 mice, light-evoked EPSCs were large (> 1 nA at -70 mV). The amplitude of these EPSCs decreased over development, reaching a plateau of ∼350 pA at P20-60. Trains of EPSCs (5 Hz) depressed strongly throughout development, whereas convergence estimates indicated that the total number of functional afferents decreased with age. EPSC waveforms consisted of multiple peaks, probably resulting from action potential bursts in single collaterals and variable times to spike threshold in converging afferents. Activating climbing fibre collaterals evoked well-timed increases in firing probability in CbN neurons, especially in younger mice. The initially strong input, followed by the decrement in synaptic strength coinciding with the pruning of climbing fibres in the cerebellar cortex, implicates the climbing fibre collateral pathway in early postnatal development. Additionally, the persistence of substantial synaptic input at least to P60 suggests that this pathway may function in cerebellar processing into adulthood.

The control of eye movements by the cerebellar nuclei: polysynaptic projections from the fastigial, interpositus posterior and dentate nuclei to lateral rectus motoneurons in primates.

Premotor circuits driving extraocular motoneurons and downstream motor outputs of cerebellar nuclei are well known. However, there is, as yet, no unequivocal account of cerebellar output pathways controlling eye movements in primates. Using retrograde transneuronal transfer of rabies virus from the lateral rectus (LR) eye muscle, we studied polysynaptic pathways to LR motoneurons in primates. Injections were placed either into the central or distal muscle portion, to identify innervation differences of LR motoneurons supplying singly innervated (SIFs) or multiply innervated muscle fibers (MIFs). We found that SIF motoneurons receive major cerebellar 'output channels' bilaterally, while oligosynaptic cerebellar innervation of MIF motoneurons is negligible and/or more indirect. Inputs originate from the fastigial nuclei di- and trisynaptically, and from a circumscribed rostral portion of the ventrolateral interpositus posterior and from the caudal pole of the dentate nuclei trisynaptically. While disynaptic cerebellar inputs to LR motoneurons stem exclusively from the caudal fastigial region involved in saccades, pursuit and convergence (via its projections to brainstem oculomotor populations), minor trisynaptic inputs from the rostral fastigial nucleus, which contributes to gaze shifts, may reflect access to vestibular and reticular eye-head control pathways. Trisynaptic inputs to LR motoneurons from the rostral ventrolateral interpositus posterior, involved in divergence (far-response), is likely mediated by projections to the supraoculomotor area, contributing to LR motoneuron activation during divergence. Trisynaptic inputs to LR motoneurons from the caudal dentate, which also innervates disynaptically the frontal and parietal eye fields, can be explained by its superior colliculus projections, and likely target saccade-related burst neurons.

Experience-Dependent Regulation of Dentate Gyrus Excitability by Adult-Born Granule Cells.

Behavioral studies have established a role for adult-born dentate granule cells in discriminating between similar memories. However, it is unclear how these cells mediate memory discrimination. Excitability is enhanced in maturing adult-born neurons, spurring the hypothesis that the activity of these cells "directly" encodes and stores memories. An alternative hypothesis posits that maturing neurons "indirectly" contribute to memory encoding by regulating excitation-inhibition balance. We evaluated these alternatives by using dentate-sensitive active place avoidance tasks to assess experience-dependent changes in dentate field potentials in the presence and absence of neurogenesis. Before training, X-ray ablation of adult neurogenesis-reduced dentate responses to perforant-path stimulation and shifted EPSP-spike coupling leftward. These differences were unchanged after place avoidance training with the shock zone in the initial location, which both groups learned to avoid equally well. In contrast, sham-treated mice decreased dentate responses and shifted EPSP-spike coupling leftward after the shock zone was relocated, whereas X-irradiated mice failed to show these changes in dentate function and were impaired on this test of memory discrimination. During place avoidance, excitation-inhibition coupled neural synchrony in dentate local field potentials was reduced in X-irradiated mice, especially in the θ band. The difference was most prominent during conflict learning, which is impaired in the X-irradiated mice. These findings indicate that maturing adult-born neurons regulate both functional network plasticity in response to memory discrimination and dentate excitation-inhibition coordination. The most parsimonious interpretation of these results is that adult neurogenesis indirectly regulates hippocampal information processing. SIGNIFICANCE STATEMENT: Adult-born neurons in the hippocampal dentate gyrus are important for flexibly using memories, but the mechanism is controversial. Using tests of hippocampus-dependent place avoidance learning and dentate electrophysiology in mice with normal or ablated neurogenesis, we find that maturing adult-born neurons are crucial only when memory must be used flexibly, and that these neurons regulate dentate gyrus synaptic and spiking responses to neocortical input rather than directly storing information, as has been proposed. A day after learning to avoid the initial or changed locations of shock, the dentate synaptic responses are enhanced or suppressed, respectively, unlike mice lacking adult neurogenesis, which did not change. The contribution of adult neurogenesis to memory is indirect, by regulating dentate excitation-inhibition coupling.

Synchrony is Key: Complex Spike Inhibition of the Deep Cerebellar Nuclei.

The control of deep cerebellar nuclear (DCN) neuronal firing is central to cerebellar function but is not well understood. The large majority of synapses onto DCN neurons derive from Purkinje cells (PCs), suggesting that PC activity is an important determinant of DCN firing; however, PCs fire both simple and complex spikes (CSs), and little is known about how the latter's action affects DCN activity. Thus, here, we explored the effects of CSs on DCN activity. CSs were recorded from PC arrays along with individual DCN neurons. Presumed synaptically connected PC-DCN cell pairs were identified using CS-triggered correlograms of DCN activity, which also showed that CS activity was associated with a predominantly inhibitory effect on DCN activity. The strength of the CS effect varied as a function of synchrony, such that isolated CSs produced only weak inhibition of DCN activity, whereas highly synchronous CSs caused a larger drop in firing levels. Although the present findings were obtained in anesthetized animals, similar CS synchrony levels exist in awake animals, and changes in synchrony level have been observed in association with movements in awake animals. Thus, the present data suggest that synchronous CS activity may be a mechanism for shaping DCN output related to motor commands.

Mathematical modeling of the neuron morphology using two dimensional images.

In this study mathematical analyses such as the analysis of area and length, fractal analysis and modified Sholl analysis were applied on two dimensional (2D) images of neurons from adult human dentate nucleus (DN). Using mathematical analyses main morphological properties were obtained including the size of neuron and soma, the length of all dendrites, the density of dendritic arborization, the position of the maximum density and the irregularity of dendrites. Response surface methodology (RSM) was used for modeling the size of neurons and the length of all dendrites. However, the RSM model based on the second-order polynomial equation was only possible to apply to correlate changes in the size of the neuron with other properties of its morphology. Modeling data provided evidence that the size of DN neurons statistically depended on the size of the soma, the density of dendritic arborization and the irregularity of dendrites. The low value of mean relative percent deviation (MRPD) between the experimental data and the predicted neuron size obtained by RSM model showed that model was suitable for modeling the size of DN neurons. Therefore, RSM can be generally used for modeling neuron size from 2D images.

Classification of adult human dentate nucleus border neurons: Artificial neural networks and multidimensional approach.

AIMS: Primary aim in this study is to investigate whether external and internal border neurons of adult human dentate nucleus express the same neuromorphological features or belong to a different morphological types i.e. whether can be classified not only by way of their topology as external and internal, but also based on their morphological features or in addition to their topology also by way of their morphology. Secondary aim is to determine and compare various methodologies in order to perform the first aim in a more accurate and efficient manner. MATERIAL AND METHODS: Blocks of tissue were cut out from the adult human cerebellum and stained according to the Kopsch-Bubenaite method. Border neurons of the dentate nucleus were investigated and digitized under the light microscope and processed thereafter. Seventeen parameters quantifying various aspects of neuron morphology are then measured. They can be categorized as shape, magnitude, complexity, length and branching parameters. Analyzes used are neural networks, separate unifactor, cluster, principal component, discriminant and correlation-comparison analysis. RESULTS: The external and internal border neurons differ significantly in six of the seventeen parameters investigated, mainly concerning dendritic ramification patterns, overall shape of dendritic tree and dendritic length. All six methodological approaches are in accordance showing slight clustering of data. Classification is based on six parameters: neuron (field) area, dendritic (field) area, total dendrite length, and position of maximal dendritic arborization density. Cluster analysis shows two data clusters. Separate unifactor analysis demonstrates inter-cluster differences with statistical significance (p < 0.05) for all six parameters separately. Principal component, discriminant and correlation-comparison analysis further prove the result on a more factor integrate manner and explain it, respectively. Thus, these neurons can be classified, not only according to their location but also according to some morphological features. Also, the group if internal border neurons is more homogeneous in itself than the other group of external border neurons. CONCLUSION: Border neurons from adult human dentate nucleus can be divided to external and internal according to its topology and based on neuromorphological computational parameters. This has potentially significant neurofunctional implications but further studies are needed to elucidate that. Multimethodological approach is shown as the best for finding the solution closest to reality. The possible functional meaning of these morphological differences for cerebellar network structure and function are discussed.

Cerebellar Nuclear Neurons Use Time and Rate Coding to Transmit Purkinje Neuron Pauses.

Neurons of the cerebellar nuclei convey the final output of the cerebellum to their targets in various parts of the brain. Within the cerebellum their direct upstream connections originate from inhibitory Purkinje neurons. Purkinje neurons have a complex firing pattern of regular spikes interrupted by intermittent pauses of variable length. How can the cerebellar nucleus process this complex input pattern? In this modeling study, we investigate different forms of Purkinje neuron simple spike pause synchrony and its influence on candidate coding strategies in the cerebellar nuclei. That is, we investigate how different alignments of synchronous pauses in synthetic Purkinje neuron spike trains affect either time-locking or rate-changes in the downstream nuclei. We find that Purkinje neuron synchrony is mainly represented by changes in the firing rate of cerebellar nuclei neurons. Pause beginning synchronization produced a unique effect on nuclei neuron firing, while the effect of pause ending and pause overlapping synchronization could not be distinguished from each other. Pause beginning synchronization produced better time-locking of nuclear neurons for short length pauses. We also characterize the effect of pause length and spike jitter on the nuclear neuron firing. Additionally, we find that the rate of rebound responses in nuclear neurons after a synchronous pause is controlled by the firing rate of Purkinje neurons preceding it.

PECULIARITIES OF THE CEREBELLUM NUCLEI IN AGED PERSONS.

The study of the clinical anatomy and functional features of the cortex, subcortical and conductive pathways of the cerebellum is necessary for clinicians for elaboration rational surgical approaches to these formations, for determination the localization of pathological processes associated with these formations. Cerebellar nucleus neurons are crucial to the olivo-cerebellar circuit as they provide the sole output of the entire cerebellum. The relationship between mobility and cognition in aging is well established, but the relationship between mobility and the structure and function of the aging brain is relatively unknown. In connection with the above, the purpose of our study was detection of the morphological characteristics of the cerebellum nuclei in aged persons. Study was performed on 48 specimens of the cerebellum from people (24 male and 24 female), who died at the age from 75 to 99 years due to diseases, which were not related to the central nervous system damaging. Formalin-fixed human hemispheres were dissected with the Ludwig and Klingler fiber dissection technique under x6 to x40 magnifications of binocular microscope Olympus BX41 (Japan). The morphological features of the human cerebellar nuclei were established. Namely, on the series of sections of the cerebellum in the horizontal, frontal and sagittal planes, as well as on the macro-microscopic preparations of the cerebellar nuclei location, their relative position, shape, linear dimensions, weight and volume were described. The features of macro-microscopic and histological structure of the nuclei of the cerebellum were made own classification of the gyri and teeth of the dentate nucleus of the cerebellum was offered. Macro-microscopic dissection of persons died after 75 years old show no significant variability of linear dimensions of cerebellar nuclei with their specific location and options. Simultaneously, reliable reducing of cellular density was detected for Purkinje, granule and basket neurons more pronounced in male for Purkinje cells.