Chondroitinase ABC promotes selective reactivation of somatosensory cortex in squirrel monkeys after a cervical dorsal column lesion

After large but incomplete lesions of ascending dorsal column afferents in the cervical spinal cord, the hand representation in the contralateral primary somatosensory cortex (area 3b) of monkeys is largely or completely unresponsive to touch on the hand. However, after weeks of spontaneous recovery, considerable reactivation of the hand territory in area 3b can occur. Because the reactivation process likely depends on the sprouting of remaining axons from the hand in the cuneate nucleus of the lower brainstem, we sought to influence cortical reactivation by treating the cuneate nucleus with an enzyme, chondroitinase ABC, that digests perineuronal nets, promoting axon sprouting. Dorsal column lesions were placed at a spinal cord level (C5/C6) that allowed a portion of ascending afferents from digit 1 to survive in squirrel monkeys. After 11-12 wk of recovery, the contralateral forelimb cortex was reactivated by stimulating digit 1 more extensively in treated monkeys than in control monkeys. The results are consistent with the proposal that the treatment enhances the sprouting of digit 1 afferents in the cuneate nucleus and that this sprouting allowed these preserved inputs to activate cortex more effectively.     

Birds have primate-like numbers of neurons in the forebrain

Some birds achieve primate-like levels of cognition, even though their brains tend to be much smaller in absolute size. This poses a fundamental problem in comparative and computational neuroscience, because small brains are expected to have a lower information-processing capacity. Using the isotropic fractionator to determine numbers of neurons in specific brain regions, here we show that the brains of parrots and songbirds contain on average twice as many neurons as primate brains of the same mass, indicating that avian brains have higher neuron packing densities than mammalian brains. Additionally, corvids and parrots have much higher proportions of brain neurons located in the pallial telencephalon compared with primates or other mammals and birds. Thus, large-brained parrots and corvids have forebrain neuron counts equal to or greater than primates with much larger brains. We suggest that the large numbers of neurons concentrated in high densities in the telencephalon substantially contribute to the neural basis of avian intelligence.Many birds have cognitive abilities that match or surpass those of mammals (1). Corvids and parrots appear to be cognitively superior to other birds, rivalling great apes in many psychological domains (1–3). They manufacture and use tools (4, 5), solve problems insightfully (6), make inferences about causal mechanisms (7), recognize themselves in a mirror (8), plan for future needs (9), and use their own experience to anticipate future behavior of conspecifics (10) or even humans (11), to mention just a few striking abilities. In addition, parrots and songbirds (including corvids) share with humans and a few other animal groups a rare capacity for vocal learning (12), and parrots can learn words and use them to communicate with humans (13).Superficially, the architecture of the avian brain appears very different from that of mammals, but recent work demonstrates that, despite a lack of layered neocortex, large areas of the avian forebrain are homologous to mammalian cortex (14–16), conform to the same organizational principles (15, 17, 18), and play similar roles in higher cognitive functions (14, 19), including executive control (20, 21). However, bird brains are small and the computational mechanisms enabling corvids and parrots to achieve ape-like intelligence with much smaller brains remain unclear. The notion that higher encephalization (relative brain size deviation from brain–body allometry) endows species with improved cognitive abilities has recently been challenged by data suggesting that intelligence instead depends on the absolute number of cerebral neurons and their connections (22–25). This is in line with recent findings that absolute rather than relative brain size is the best predictor of cognitive capacity (26–28). However, although corvids and parrots feature encephalization comparable to that of monkeys and apes, their absolute brain size remains small (29, 30). The largest average brain size in corvids and parrots does not exceed 15.4 g found in the common raven (29) and 24.7 g found in the hyacinth macaw (30), respectively. Do corvids and parrots provide a strong case for reviving encephalization as a valid measure of brain functional capacity? Not necessarily: it has recently been discovered that the relationship between brain mass and number of brain neurons differs starkly between mammalian clades (31). Avian brains seem to consist of small, tightly packed neurons, and it is thus possible that they can accommodate numbers of neurons that are comparable to those found in the much larger primate brains. However, to date, no quantitative data have been available to test this hypothesis.Here, we analyze how numbers of neurons compare across birds and mammals (32–39) of equivalent brain mass, and determine the cellular scaling rules for brains of songbirds and parrots. Using the isotropic fractionator (40), we estimated the total numbers of neuronal and nonneuronal cells in the cerebral hemispheres, cerebellum, diencephalon, tectum, and brainstem in a sample of 11 parrot species, 13 vocal learning songbird species (including 6 corvids), and 4 additional model species representing other avian clades (Figs. S1 and ​andS2).S2). Because most of the cited mammalian studies analyzed cellular composition of only three brain subdivisions, namely the pallium (referred to as the cerebral cortex in those papers), the cerebellum, and rest of brain, we divided the avian brain identically to ensure an accurate comparison of neuronal numbers, densities, and relative distribution of neurons in birds and mammals. Specifically, the avian pallium (comprising the hyperpallium, mesopallium, nidopallium, arcopallium, and hippocampus) was compared with its homolog—the mammalian pallium (comprising the neocortex, hippocampus, olfactory cortices such as piriform and entorhinal cortex, and pallial amygdala) (14–16, 41). The avian subpallium (formed by the striatum, pallidum, and septum), diencephalon, tectum, and brainstem were pooled and compared with the same regions of mammalian brains that are referred to as “the rest of brain.” The cerebellum is directly compared between the two clades. The results of our study reveal that avian brains contain many more pallial neurons than equivalently sized mammalian brains.Open in a separate windowFig. S1.Phylogenetic relationships among the 28 species examined. The tree was constructed using birdtree.org/; its topology follows recent studies (46–49). Note that songbirds and parrots are sister groups and together with the distantly related barn owl belong to the clade core landbirds (Telluraves); the pigeon represents the Columbea, a basal clade of the Neoaves; the red junglefowl represents the Galloanseres, a sister group of Neoaves and the most basal clade of Neognathae; and the emu represents Paleognathae (tinamous and flightless ostriches), the most basal clade of extant birds (48). Also note that all passerine birds examined were vocal learners belonging to the clade Oscines.Open in a separate windowFig. S2.Brain dissection and labeling of neurons and nonneuronal cells. (A and B) Brain of the raven before and after the dissection. (A) Ventral side of the brain showing approximate lines of dissection of the brainstem and tectum. (B) Brain dissected into parts used for isotropic fractionation. (C) NeuN-immunolabeled transverse section of the zebra finch brain depicting the line of dissection of the tectum from the rest of the mesencephalon. (D–F) Dissection of the telencephalon into pallium and subpallium. NeuN-immunolabeled transverse sections of the zebra finch brain at rostral (D), intermediate (E), and caudal (F) telencephalic levels. Lines of dissection follow the pallial-subpallial lamina and divide the telencephalon into pallium (dorsal part) and subpallium (ventral part). Coordinates anterior to the Y point are indicated in millimeters at Bottom Left (64). (G–I) High-power micrographs showing a sample of homogenate from the telencephalon of the Eurasian jay; dissociated nuclei stained with DAPI (G) and immunolabeled with NeuN antibody (H), dual-fluorescence merge image (I). Note that neurons are double-labeled, whereas the nonneuronal cells are devoid of anti-NeuN immunoreactivity. [Scale bars: 10 mm (A and B); 1 mm (C and F); 50 µm (I).]     

Identification of a population of sleep-active cerebral cortex neurons

The presence of large-amplitude, slow waves in the EEG is a primary characteristic that distinguishes cerebral activity during sleep from that which occurs during wakefulness. Although sleep-active neurons have been identified in other brain areas, neurons that are specifically activated during slow-wave sleep have not previously been described in the cerebral cortex. We have identified a population of cells in the cortex that is activated during sleep in three mammalian species. These cortical neurons are a subset of GABAergic interneurons that express neuronal NOS (nNOS). Because Fos expression in these sleep-active, nNOS-immunoreactive (nNOS-ir) neurons parallels changes in the intensity of slow-wave activity in the EEG, and these neurons are innvervated by neurotransmitter systems previously implicated in sleep/wake control, cortical nNOS-ir neurons may be part of the neurobiological substrate that underlies homeostatic sleep regulation.     

Delayed plasticity of inhibitory neurons in developing visual cortex

During postnatal development, altered sensory experience triggers the rapid reorganization of neuronal responses and connections in sensory neocortex. This experience-dependent plasticity is disrupted by reductions of intracortical inhibition. Little is known about how the responses of inhibitory cells themselves change during plasticity. We investigated the time course of inhibitory cell plasticity in mouse primary visual cortex by using functional two-photon microscopy with single-cell resolution and genetic identification of cell type. Initially, local inhibitory and excitatory cells had similar binocular visual response properties, both favoring the contralateral eye. After 2 days of monocular visual deprivation, excitatory cell responses shifted to favor the open eye, whereas inhibitory cells continued to respond more strongly to the deprived eye. By 4 days of deprivation, inhibitory cell responses shifted to match the faster changes in their excitatory counterparts. These findings reveal a dramatic delay in inhibitory cell plasticity. A minimal linear model reveals that the delay in inhibitory cell plasticity potently accelerates Hebbian plasticity in neighboring excitatory neurons. These findings offer a network-level explanation as to how inhibition regulates the experience-dependent plasticity of neocortex.     

Emergence of functional subnetworks in layer 2/3 cortex induced by sequential spikes in vivo

During cortical circuit development in the mammalian brain, groups of excitatory neurons that receive similar sensory information form microcircuits. However, cellular mechanisms underlying cortical microcircuit development remain poorly understood. Here we implemented combined two-photon imaging and photolysis in vivo to monitor and manipulate neuronal activities to study the processes underlying activity-dependent circuit changes. We found that repeated triggering of spike trains in a randomly chosen group of layer 2/3 pyramidal neurons in the somatosensory cortex triggered long-term plasticity of circuits (LTPc), resulting in the increased probability that the selected neurons would fire when action potentials of individual neurons in the group were evoked. Significant firing pattern changes were observed more frequently in the selected group of neurons than in neighboring control neurons, and the induction was dependent on the time interval between spikes, N-methyl-D-aspartate (NMDA) receptor activation, and Calcium/calmodulin-dependent protein kinase II (CaMKII) activation. In addition, LTPc was associated with an increase of activity from a portion of neighboring neurons with different probabilities. Thus, our results demonstrate that the formation of functional microcircuits requires broad network changes and that its directionality is nonrandom, which may be a general feature of cortical circuit assembly in the mammalian cortex.Layer 2/3 neurons in the barrel cortex play a central role in integrative cortical processing (1–4). Neurons in layer 2/3 are interconnected with each other, and their axons and dendrites traverse adjacent barrel areas (5, 6). Recent calcium (Ca²⁺) imaging studies in awake animals showed that two very closely localized layer 2/3 pyramidal neurons are independently activated by different whiskers (7). In addition, adjacent layer 2/3 neurons have different receptive field properties; signals from different whiskers may emerge on different spines in the same neurons (8, 9). These findings suggest that the organization of functional subnetworks in somatosensory layer 2/3 is heterogeneous at the single-cell level and that microcircuits are assembled at a very fine scale (10). In vivo whole-cell recording experiments have also shown that most, but not all, layer 2/3 pyramidal neurons receive subthreshold depolarization by single-whisker stimulation with much broader receptive fields than neurons in layer 4 (11, 12). These anatomical and functional data suggest that electric signals relayed to the cortex by whisker activation are greatly intermingled within layer 2/3 neurons, and that studying the mechanisms by which these layer 2/3 neurons make connections may be critical for understanding the cortical network organizing principles underlying somatosensation.A previous modeling study suggested that spike timing-dependent plasticity (STDP) can lead to the formation of functional cortical columns and activity-dependent reorganization of neural circuits (13–16). However, how spikes arising in multiple neurons in vivo influence their connectivity is poorly understood. In this study using two-photon glutamate photolysis, which allowed us to control neuronal activity in a spatially and temporally precise manner, we examined activity-dependent cellular mechanisms during network rearrangement generated by repetitive spike trains in a group of neurons. We found that repetitive spikes on a group of neurons induced the probability of the neurons firing together. This circuit plasticity required spiking at short intervals among neurons and is expressed by N-methyl-D-aspartate (NMDA) receptor- and Calcium/calmodulin-dependent protein kinase II (CaMKII)-dependent long-lasting connectivity changes. The probability of firing was differentially affected by the order of the spike sequence but was not dependent on the physical distance between neurons. Thus, our data show that neuronal connectivity within a functional subnetwork is established in not only a preferred but also a directional manner.     

Changing numbers of neuronal and non-neuronal cells underlie postnatal brain growth in the rat

The rat brain increases >6× in mass from birth to adulthood, presumably through the addition of glial cells and increasing neuronal size, without the addition of neurons. To test this hypothesis, here we investigate quantitatively the postnatal changes in the total number of neuronal and non-neuronal cells in the developing rat brain, and examine how these changes correlate with brain growth. Total numbers of cells were determined with the isotropic fractionator in the brains of 53 Wistar rats, from birth to young adulthood. We find that at birth, >90% of the cells in the rat brain are neurons. Following a dormant period of ≈3 days after birth, the net number of neurons in the cerebral cortex, hippocampus, and remaining tissue (excluding cerebellum and olfactory bulb) doubles during the first week, then is reduced by 70% during the second postnatal week, concurrently with net gliogenesis. A second round of net addition of 6 million neurons is observed in the cerebral cortex over the following 2 weeks. During the first postnatal week, brain growth relates mainly to increased numbers of neurons of larger average size. In the second and third weeks, it correlates with increased numbers of non-neuronal cells that are smaller in size than the preexisting neurons. Postnatal rat brain development is thus characterized by dramatic changes in the cellular composition of the brain, whose growth is governed by different combinations of cell addition and loss, and changes in average cell size during the first months after birth.     

Temporally tuned neuronal differentiation supports the functional remodeling of a neuronal network in Drosophila

During insect metamorphosis, neuronal networks undergo extensive remodeling by restructuring their connectivity and recruiting newborn neurons from postembryonic lineages. The neuronal network that directs the essential behavior, ecdysis, generates a distinct behavioral sequence at each developmental transition. Larval ecdysis replaces the cuticle between larval stages, and pupal ecdysis externalizes and expands the head and appendages to their adult position. However, the network changes that support these differences are unknown. Crustacean cardioactive peptide (CCAP) neurons and the peptide hormones they secrete are critical for ecdysis; their targeted ablation alters larval ecdysis progression and results in a failure of pupal ecdysis. In this study, we demonstrate that the CCAP neuron network is remodeled immediately before pupal ecdysis by the emergence of 12 late CCAP neurons. All 12 are CCAP efferents that exit the central nervous system. Importantly, these late CCAP neurons were found to be entirely sufficient for wild-type pupal ecdysis, even after targeted ablation of all other 42 CCAP neurons. Our evidence indicates that late CCAP neurons are derived from early, likely embryonic, lineages. However, they do not differentiate to express their peptide hormone battery, nor do they project an axon via lateral nerve trunks until pupariation, both of which are believed to be critical for the function of CCAP efferent neurons in ecdysis. Further analysis implicated ecdysone signaling via ecdysone receptors A/B1 and the nuclear receptor ftz-f1 as the differentiation trigger. These results demonstrate the utility of temporally tuned neuronal differentiation as a hard-wired developmental mechanism to remodel a neuronal network to generate a scheduled change in behavior.     

Fast-scale network dynamics in human cortex have specific spectral covariance patterns

Whether measured by MRI or direct cortical physiology, infraslow rhythms have defined state invariant cortical networks. The time scales of this functional architecture, however, are unlikely to be able to accommodate the more rapid cortical dynamics necessary for an active cognitive task. Using invasively monitored epileptic patients as a research model, we tested the hypothesis that faster frequencies would spectrally bind regions of cortex as a transient mechanism to enable fast network interactions during the performance of a simple hear-and-repeat speech task. We term these short-lived spectrally covariant networks functional spectral networks (FSNs). We evaluated whether spectrally covariant regions of cortex, which were unique in their spectral signatures, provided a higher degree of task-related information than any single site showing more classic physiologic responses (i.e., single-site amplitude modulation). Taken together, our results showing that FSNs are a more sensitive measure of task-related brain activation and are better able to discern phonemic content strongly support the concept of spectrally encoded interactions in cortex. Moreover, these findings that specific linguistic information is represented in FSNs that have broad anatomic topographies support a more distributed model of cortical processing.The brain''s intrinsic functional architecture of correlated fluctuations in resting state metabolic and electrophysiologic activity has been well established (1, 2). This functional architecture has been shown to be present in the absence of a task, during all stages of sleep, and even under anesthesia (3). How anatomically distributed regions of cortex interact during the performance of a cognitive task is less understood. Due to slower time scales associated with the hemodynamic response of current neuroimaging techniques (4) and their electrophysiologic correlates (1), the more static networks are not adequate to accommodate the more rapid dynamics associated with many behavioral tasks. Given the limitations of the described time scales, we hypothesized that faster frequencies would “spectrally bind” regions of cortex as a transient mechanism to enable fast network interactions that accommodate the flexible use of neuronal resources. Beyond previously described notions that single higher-frequency synchronization enables neuronal interactions (5), we postulated that dynamic networks are represented by a multitude of spectral characteristics. These transient spectrally covariant networks, which we term functional spectral networks (FSNs), would enable a higher level of fidelity in the transmission of cortical-cortical information.Using invasively monitored epileptic patients as a research model, we tested this hypothesis in the setting of a simple hear-and-repeat task. Given that human speech processing involves a widely distributed area located predominantly in perisylvian regions (6), this provided a robust model to evaluate network-derived behavior. We evaluated whether spectrally covariant regions of cortex, which were unique in their spectral signatures, provided a higher degree of task-related information than any single site showing more classic physiologic responses. To minimize the impact of ictal/peri-ictal-induced alterations in cortical coherence (7, 8), all patients had normal speech function, their seizure onset zone was distinct from stimulation-defined speech areas, and testing was done on days without seizures. Taken together, our results showing that FSNs are a more sensitive measure of task-related brain activation and are better able to discern phonemic content strongly support the concept of spectrally encoded interactions in cortex. Moreover, these findings showing that specific linguistic information is represented in FSNs that have broad anatomic topographies supports a more distributed model of cortical processing.     

Interactive effects of age and estrogen on cognition and pyramidal neurons in monkey prefrontal cortex

We previously reported that long-term cyclic estrogen (E) treatment reverses age-related impairment of cognitive function mediated by the dorsolateral prefrontal cortex (dlPFC) in ovariectomized (OVX) female rhesus monkeys, and that E induces a corresponding increase in spine density in layer III dlPFC pyramidal neurons. We have now investigated the effects of the same E treatment in young adult females. In contrast to the results for aged monkeys, E treatment failed to enhance dlPFC-dependent task performance relative to vehicle control values (group young OVX+Veh) but nonetheless led to a robust increase in spine density. This response was accompanied by a decline in dendritic length, however, such that the total number of spines per neuron was equivalent in young OVX+Veh and OVX+E groups. Robust effects of chronological age, independent of ovarian hormone status, were also observed, comprising significant age-related declines in dendritic length and spine density, with a preferential decrease in small spines in the aged groups. Notably, the spine effects were partially reversed by cyclic E administration, although young OVX+Veh monkeys still had a higher complement of small spines than did aged E treated monkeys. In summary, layer III pyramidal neurons in the dlPFC are sensitive to ovarian hormone status in both young and aged monkeys, but these effects are not entirely equivalent across age groups. The results also suggest that the cognitive benefit of E treatment in aged monkeys is mediated by enabling synaptic plasticity through a cyclical increase in small, highly plastic dendritic spines in the primate dlPFC.     

Experience-dependent functional plasticity and visual response selectivity of surviving subplate neurons in the mouse visual cortex

Subplate neurons are early-born cortical neurons that transiently form neural circuits during perinatal development and guide cortical maturation. Thereafter, most subplate neurons undergo cell death, while some survive and renew their target areas for synaptic connections. However, the functional properties of the surviving subplate neurons remain largely unknown. This study aimed to characterize the visual responses and experience-dependent functional plasticity of layer 6b (L6b) neurons, the remnants of subplate neurons, in the primary visual cortex (V1). Two-photon Ca²⁺ imaging was performed in V1 of awake juvenile mice. L6b neurons showed broader tunings for orientation, direction, and spatial frequency than did layer 2/3 (L2/3) and L6a neurons. In addition, L6b neurons showed lower matching of preferred orientation between the left and right eyes compared with other layers. Post hoc 3D immunohistochemistry confirmed that the majority of recorded L6b neurons expressed connective tissue growth factor (CTGF), a subplate neuron marker. Moreover, chronic two-photon imaging showed that L6b neurons exhibited ocular dominance (OD) plasticity by monocular deprivation during critical periods. The OD shift to the open eye depended on the response strength to the stimulation of the eye to be deprived before starting monocular deprivation. There were no significant differences in visual response selectivity prior to monocular deprivation between the OD changed and unchanged neuron groups, suggesting that OD plasticity can occur in L6b neurons showing any response features. In conclusion, our results provide strong evidence that surviving subplate neurons exhibit sensory responses and experience-dependent plasticity at a relatively late stage of cortical development.

The mammalian cerebral cortex consists of six layers, with distinct roles in information processing (1, 2). At the bottom of the neocortex, on the boundary between the gray matter and white matter, there is a thin sheet of neurons called layer 6b (L6b) (3). Layer 6b neurons are thought to be remnants of subplate neurons based on their location and cell-type marker expression (4). During prenatal and early postnatal periods, subplate neurons form transient neuronal circuits that play key roles in cortical maturation (5–7). In the embryonic cortex, subplate neurons form short-lived synapses with early immature neurons to regulate radial migration (8). During perinatal development, subplate neurons transiently receive inputs from ingrowing thalamic axons and innervate layer 4 (L4) to guide thalamic inputs to the eventual target, L4 (5, 6). Thus, the circuits formed by subplate neurons at the perinatal developmental stage are essential to establish basic neuronal circuits before starting experience-dependent refinements (5–7). Subsequently, subplate neurons largely disappear due to programmed cell death, but some survive and reside in L6b (5, 6). In the adult cortex, L6b neurons form neuronal circuits with local and long-distance neurons, which are different from those formed during early development (9–12). Therefore, surviving subplate neurons may acquire a role in information processing after remodeling of neuronal connections. A recent study using three-photon Ca²⁺ imaging demonstrated that L6b neurons show visual responses with broad orientation/direction tuning in the adult mouse primary visual cortex (V1) (13). However, comparable evidence for L6b response properties with other layer neurons in V1 is lacking (14–20). Moreover, L6b neurons have diverse morphology and molecular expression (21–24). Neurons born during subplate neurogenesis show the different expression patterns of subplate markers in postnatal L6b (4). However, the response properties in each subtype of L6b neurons remain unknown.The sensory responsiveness of cortical neurons is considerably refined by sensory experience relatively late in development, referred to as the critical period (25, 26). Previous studies have demonstrated that sensory activities before the onset of the critical period affect the arrangement of subplate neuron neurites in the barrel cortex and local subplate circuits in the auditory cortex (27, 28). However, there is no direct evidence that the sensory responses of surviving subplate neurons are modified by sensory experience during the critical period. If experience-dependent plasticity occurs in subplate neuron responses, they will contribute to the experience-dependent development of sensory functions and possibly to the functions in the mature cortex. Ocular dominance (OD) plasticity in V1 is a canonical model used to examine experience-dependent refinement of sensory responses (25, 26, 29, 30). If one eye is occluded for several days during the critical period, neurons in V1 lose their response to the deprived eye. OD plasticity is robustly preserved across species and cell types. Therefore, OD plasticity is suitable for evaluating experience-dependent plasticity in L6b neurons.This study aimed to characterize the visual responses and OD plasticity of L6b neurons in V1. Toward this goal, two-photon Ca²⁺ imaging was performed in awake juvenile mice, followed by 3D immunohistochemistry with a subplate neuronal marker, connective tissue growth factor (CTGF) (4, 31). L6b neurons showed broader tuning to visual stimuli and lower binocular matching of orientation preference than did layer 2/3 (L2/3) and L6a neurons. Chronic two-photon imaging revealed significant OD plasticity in individual L6b neurons during the critical period. Our results provide strong evidence that L6b neurons, presumed to be subplate neuron remnants, exhibit sensory responses and experience-dependent functional plasticity at a relatively late stage of cortical development.     

Procedure for recording the simultaneous activity of single neurons distributed across cortical areas during sensory discrimination

We report a procedure for recording the simultaneous activity of single neurons distributed across five cortical areas in behaving monkeys. The procedure consists of a commercially available microdrive adapted to a commercially available neural data collection system. The critical advantage of this procedure is that, in each cortical area, a configuration of seven microelectrodes spaced 250-500 mum can be inserted transdurally and each can be moved independently in the z axis. For each microelectrode, the data collection system can record the activity of up to five neurons together with the local field potential (LFP). With this procedure, we normally monitor the simultaneous activity of 70-100 neurons while trained monkeys discriminate the difference in frequency between two vibrotactile stimuli. Approximately 20-60 of these neurons have response properties previously reported in this task. The neuronal recordings show good signal-to-noise ratio, are remarkably stable along a 1-day session, and allow testing several protocols. Microelectrodes are removed from the brain after a 1-day recording session, but are reinserted again the next day by using the same or different x-y microelectrode array configurations. The fact that microelectrodes can be moved in the z axis during the recording session and that the x-y configuration can be changed from day to day maximizes the probability of studying simultaneous interactions, both local and across distant cortical areas, between neurons associated with the different components of this task.     

Spine dynamics of PSD-95-deficient neurons in the visual cortex link silent synapses to structural cortical plasticity

Critical periods (CPs) are time windows of heightened brain plasticity during which experience refines synaptic connections to achieve mature functionality. At glutamatergic synapses on dendritic spines of principal cortical neurons, the maturation is largely governed by postsynaptic density protein-95 (PSD-95)-dependent synaptic incorporation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors into nascent AMPA-receptor silent synapses. Consequently, in mouse primary visual cortex (V1), impaired silent synapse maturation in PSD-95-deficient neurons prevents the closure of the CP for juvenile ocular dominance plasticity (jODP). A structural hallmark of jODP is increased spine elimination, induced by brief monocular deprivation (MD). However, it is unknown whether impaired silent synapse maturation facilitates spine elimination and also preserves juvenile structural plasticity. Using two-photon microscopy, we assessed spine dynamics in apical dendrites of layer 2/3 pyramidal neurons (PNs) in binocular V1 during ODP in awake adult mice. Under basal conditions, spine formation and elimination ratios were similar between PSD-95 knockout (KO) and wild-type (WT) mice. However, a brief MD affected spine dynamics only in KO mice, where MD doubled spine elimination, primarily affecting newly formed spines, and caused a net reduction in spine density similar to what has been observed during jODP in WT mice. A similar increase in spine elimination after MD occurred if PSD-95 was knocked down in single PNs of layer 2/3. Thus, structural plasticity is dictated cell autonomously by PSD-95 in vivo in awake mice. Loss of PSD-95 preserves hallmark features of spine dynamics in jODP into adulthood, revealing a functional link of PSD-95 for experience-dependent synapse maturation and stabilization during CPs.

Early life of an animal is characterized by time windows of functionally and structurally enhanced brain plasticity known as critical periods (CPs), which have been described initially in the primary visual cortex (V1) of kittens (1). During CPs, experience refines the connectivity of principal excitatory neurons to establish the mature functionality of neural networks. This refinement is governed by the constant generation and elimination of nascent synapses on dendritic spines that sample favorable connections to be consolidated and unfavorable ones to be eliminated (2–5). A fraction of nascent synapses is or becomes α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-receptor silent, expressing N-methyl-D-aspartate (NMDA) receptors only (6–8). At eye opening, silent synapses are abundant in the primary visual cortex (V1) (9, 10) and mature during CPs by stable AMPA receptor incorporation (11–14). The pace of silent synapse maturation is governed by the opposing yet cooperative function of postsynaptic density protein of 95 kDa (PSD-95) and its paralog PSD-93, two signaling scaffolds of the postsynaptic density of excitatory synapses (12, 13). However, whether silent synapses are preferential substrates for spine elimination during CPs remains to be investigated.In juvenile mice (postnatal days [P] 20 to 35), a brief monocular deprivation (MD) of the dominant contralateral eye results in a shift of the ocular dominance (OD) of binocular neurons in V1 toward the open eye, mediated by a reduction of responses to visual stimulation of the deprived eye (15–17). Structurally, MD induces an increase in spine elimination in apical dendrites of layer (L) 2/3 and L5 pyramidal neurons (PNs) which is only observed during the CP and constitutes a hallmark of juvenile OD plasticity (jODP) (18–20). After CP closure, cortical plasticity declines progressively, and in standard cage-raised mice beyond P40, a 4-d MD no longer induces the functional nor anatomical changes associated with jODP (21–24).At least three different mechanisms involved in experience-dependent maturation of cortical neural networks have been described, but the molecular and cellular mechanisms that cause CP closure remain highly debated (18, 25, 26). First, plasticity of local inhibitory neurons, such as increased inhibitory tone or a reduction of release probability by experience-dependent endocannabinoid receptor 1 (CB1R) activation was reported to close the critical period in rodent V1 (27–29). Second, the expression of so-called “plasticity brakes,” such as extracellular matrix (ECM), Nogo receptor 1 (NgR1), paired immunoglobulin-like receptor B (PirB), and Lynx1 were correlated with the end of critical periods (30–33). Experimentally decreasing the inhibitory tone or absence of plasticity brakes enhanced ODP expression in various knockout (KO) mouse models (32, 34, 35), among which only Lynx1 KO mice were shown to exhibit functional hallmarks of jODP, such as selective deprived eye depression after a short MD (36). Structurally, Lynx1 KO mice exhibited elevated spine dynamics at baseline; however, MD induced a reduction in spine elimination in apical dendrites of L5 PNs, whereas in L2/3 PNs there was no change (37). Thus, the effects of removing plasticity brakes on structural plasticity are variable, and it remains unclear to what extend manipulating the plasticity brakes can reinstate cellular signatures of CP plasticity in the adult wild-type (WT) brain (38). Third, the progressive maturation of AMPAR-silent synapses was correlated with the closure of the CP for jODP (12, 13). Consequently, in PSD-95 KO mice, the maturation of silent synapses is impaired; their fraction remains at the eye opening level, and jODP is preserved lifelong (13). Furthermore, visual cortex-specific knockdown (KD) of PSD-95 in the adult brain reinstated jODP. In contrast, in PSD-93 KO mice, silent synapses mature precociously and the CP for jODP closes precociously (12), correlating the presence of silent synapses with functional plasticity during CPs.While these three mechanisms of CP closure are not mutually exclusive in regulating cortical plasticity (26), it remains elusive whether CP-like structural plasticity can be expressed in the adult brain and whether silent synapses might be substrates for it. Here, we performed chronic two-photon imaging of dendrites of L2/3 pyramidal neurons in binocular V1 of PSD-95 KO (and KD) and WT mice, tracking the same dendritic spines longitudinally before, during, and after a 4-d period of MD. As previous studies have reported anesthesia effects on spine dynamics (39–41), we performed our experiments in awake mice, thoroughly trained for head fixation under the two-photon microscope. Our chronic spine imaging experiments revealed that in adult PSD-95 KO and KD mice, a brief MD indeed increased spine elimination about twofold, while adult WT mice did not display experience-dependent changes in spine elimination or spine formation. Thus, the loss of PSD-95 led to a high number of AMPAR-silent synapses which were correlated with jODP after MD, and with juvenile-like structural plasticity even in the adult brain, underscoring the importance of silent synapses for CP-timing and network maturation and stabilization.     

Sexual dimorphism in numbers of vasotocin-immunoreactive neurons in brain areas associated with reproductive behaviors in the roughskin newt

Vasotocin (VT) and vasopressin control many endocrine and neuroendocrine functions, including the regulation of reproductive behaviors. In the roughskin newt (Taricha granulosa), VT administration can enhance courtship behaviors in males and egg-laying behaviors in females. This study used immunohistochemistry to investigate whether there are sex differences in VT in specific brain areas, and whether these differences persist in nonbreeding animals. Numbers of VT immunoreactive (ir) cell bodies were counted in males and females collected in February, April, June, and August. Radioimmunoassay of plasma samples confirmed that testosterone and 5alpha-dihydrotestosterone concentrations were higher in males than females, and that 17beta-estradiol concentrations were higher in females than males. In 11 brain areas, no sexual or seasonal differences in the number of VTir cells were found. But in 3 brain regions-the bed nucleus of the stria terminalis (BNST), the nucleus amygdalae dorsolateralis (AMYG), and the anterior preoptic area (aPOA)-there were significantly greater numbers of VTir cells in males than in females, and these differences did not change seasonally. In the aPOA, an area important to male sex behaviors, the sexual dimorphism in VTir was particularly pronounced. In four brain regions, there were significantly greater numbers of VTir cells in females than males, but only in specific seasons. In April-collected (breeding) animals, more VTir cells were found in females than in males in the populations of VT cells within the pars dorsalis hypothalami and ventromedial hypothalamus, brain regions frequently associated with stress responses and female mating behaviors. In August-collected (nonbreeding) animals, more VTir cells were found in females than in males, in the region of the bed nucleus of the decussation of the fasciculus lateralis telencephali and in the nucleus visceralis superior, nucleus isthmi region. Significantly greater numbers of VTir cells were observed in the magnocellular preoptic area of males and females collected in February. These results indicate that the functional interactions between gonadal steroid hormones and VT are complex and appear to involve site-, sex-, and season-specific regulatory mechanisms. Furthermore, it seems likely that populations of VT neurons in the BNST, AMYG, and aPOA are involved in regulating male-specific behaviors, and that the VT neurons in the pars dorsalis hypothalami/ventromedial hypothalamus may be involved in female-specific behaviors.     

Language processing in the occipital cortex of congenitally blind adults

Humans are thought to have evolved brain regions in the left frontal and temporal cortex that are uniquely capable of language processing. However, congenitally blind individuals also activate the visual cortex in some verbal tasks. We provide evidence that this visual cortex activity in fact reflects language processing. We find that in congenitally blind individuals, the left visual cortex behaves similarly to classic language regions: (i) BOLD signal is higher during sentence comprehension than during linguistically degraded control conditions that are more difficult; (ii) BOLD signal is modulated by phonological information, lexical semantic information, and sentence-level combinatorial structure; and (iii) functional connectivity with language regions in the left prefrontal cortex and thalamus are increased relative to sighted individuals. We conclude that brain regions that are thought to have evolved for vision can take on language processing as a result of early experience. Innate microcircuit properties are not necessary for a brain region to become involved in language processing.     

Statistical learning of visual transitions in monkey inferotemporal cortex

One of the most fundamental functions of the brain is to predict upcoming events on the basis of the recent past. A closely related function is to signal when a prediction has been violated. The identity of the brain regions that mediate these functions is not known. We set out to determine whether they are implemented at the level of single neurons in the visual system. We gave monkeys prolonged exposure to pairs of images presented in fixed sequence so that each leading image became a strong predictor for the corresponding trailing image. We then monitored the responses of neurons in the inferotemporal cortex to image sequences that obeyed or violated the transitional rules imposed during training. Inferotemporal neurons exhibited a transitional surprise effect, responding much more strongly to unpredicted transitions than to predicted transitions. Thus, neurons even in the visual system make experience-based predictions and react when they fail.     

Immunolocalization of GLUTX1 in the testis and to specific brain areas and vasopressin-containing neurons.

GLUTX1 or GLUT8 is a newly characterized glucose transporter isoform that is expressed at high levels in the testis and brain and at lower levels in several other tissues. Its expression was mapped in the testis and brain by using specific antibodies. In the testis, immunoreactivity was expressed in differentiating spermatocytes of type 1 stage but undetectable in mature spermatozoa. In the brain, GLUTX1 distribution was selective and localized to a variety of structures, mainly archi- and paleocortex. It was found in hippocampal and dentate gyrus neurons as well as amygdala and primary olfactory cortex. In these neurons, its location was close to the plasma membrane of cell bodies and sometimes in proximal dendrites. High GLUTX1 levels were detected in the hypothalamus, supraoptic nucleus, median eminence, and the posterior pituitary. Neurons of these areas synthesize and secrete vasopressin and oxytocin. As shown by double immunofluorescence microscopy and immunogold labeling, GLUTX1 was expressed only in vasopressin neurons. By immunogold labeling of ultrathin cryosections microscopy, GLUTX1 was identified in dense core vesicles of synaptic nerve endings of the supraoptic nucleus and secretory granules of the vasopressin positive neurons. This localization suggests an involvement of GLUTX1 both in specific neuron function and endocrine mechanisms.     

突触可塑性在血管性认知障碍中的作用

突触损伤在血管性认知障碍(vascular cognitive impairment, VCT)发病的早期即存在,与认知功能障碍关系密切,其具体机制尚不明确.研究突触形态结构的可塑性、突触传递效能的可塑性以及突触蛋白在VCI发病中的作用和机制,有助于进一步阐明VCI的发病机制,从而更有效地防治VCI.     

Predicting visual acuity from the structure of visual cortex

Three decades ago, Rockel et al. proposed that neuronal surface densities (number of neurons under a square millimeter of surface) of primary visual cortices (V1s) in primates is 2.5 times higher than the neuronal density of V1s in nonprimates or many other cortical regions in primates and nonprimates. This claim has remained controversial and much debated. We replicated the study of Rockel et al. with attention to modern stereological precepts and show that indeed primate V1 is 2.5 times denser (number of neurons per square millimeter) than many other cortical regions and nonprimate V1s; we also show that V2 is 1.7 times as dense. As primate V1s are denser, they have more neurons and thus more pinwheels than similar-sized nonprimate V1s, which explains why primates have better visual acuity.Rockel et al. (1), in an influential and controversial article entitled “The basic uniformity in structure of the neocortex,” reported that the number of neurons underneath a square millimeter of neocortical surface is constant for six cortical areas and five species with one exception: primate primary visual cortex (V1) has a surface density of about 250,000 neurons/mm², around two and a half times the usual density for other areas studied.The Rockel et al. paper has, for a third of a century, continued to generate controversy for two reasons. One reason stems from its implications for an equally energetic debate among neuroscientist “lumpers” and “splitters.” Cortical uniformity supports a theory of neocortical processing wherein different cortical areas are subserved by the same canonical circuit, a view favored by lumpers. Splitters, however, believe each cortical area to be different and doubt the paper’s claims (2). The second reason is that studies from various laboratories using different measurement methods over the last three decades have alternately agreed and disagreed with Rockel et al.’s results (2).Notably, however, Rockel et al’s studies have never been directly replicated. We set out to repeat the observations for the same areas and species Rockel et al. used. We used Rockel et al.’s counting techniques but with attention to the precepts of modern stereology (2). Our goal was to simply determine if Rockel et al.’s observations are repeatable rather than address the larger question of numerical uniformity of neocortex across species and areas. In an earlier publication (2), we confirmed Rockel et al.’s conclusions for nonvisual areas. Here we focus on the primary V1 for the same species used in the original report of Rockel et al.V1 is part of the visual circuit from the retina to the cortex, which is retinotopically organized, and the 2D image of the world that is mapped onto the retina is recreated in V1 (3). Cells within the retina capture visual information for each image location or pixel such as color and light intensity and convey it to structures in V1 (4, 5), which perform computations that contribute to visual abilities. An especially well-studied V1 structure is a pinwheel, which comprises orientation columns that extend vertically down from the cortical surface, containing cells with receptive fields responsive to lines or edges at a particular angle. Columns within a pinwheel are organized so that the angle of orientation increases/decreases smoothly as one radially traverses the cortical surface and stays constant as one moves along a spoke of the pinwheel (5, 6). We wondered whether evolution might have designed primate V1 to be denser to increase the number of cells and pinwheels (7) and thus the computational power of V1 and visual abilities (8).We estimated the neuronal surface density (the number of neurons under a square millimeter of neocortical surface) of V1 for mouse, rat, cat, and monkey (rhesus macaque) and confirmed Rockel et al.’s original report: the first three species have a surface density of about 10⁵ neurons/mm² and monkey V1 has about 2.5 × 10⁵ neurons/mm². We also found that monkey V2 has about 1.7 × 10⁵ neurons/mm².     

The basic nonuniformity of the cerebral cortex

Evolutionary changes in the size of the cerebral cortex, a columnar structure, often occur through the addition or subtraction of columnar modules with the same number of neurons underneath a unit area of cortical surface. This view is based on the work of Rockel et al. [Rockel AJ, Hiorns RW, Powell TP (1980) The basic uniformity in structure of the neocortex. Brain 103:221–244], who found a steady number of approximately 110 neurons underneath a surface area of 750 μm² (147,000 underneath 1 mm²) of the cerebral cortex of five species from different mammalian orders. These results have since been either corroborated or disputed by different groups. Here, we show that the number of neurons underneath 1 mm² of the cerebral cortical surface of nine primate species and the closely related Tupaia sp. is not constant and varies by three times across species. We found that cortical thickness is not inversely proportional to neuronal density across species and that total cortical surface area increases more slowly than, rather than linearly with, the number of neurons underneath it. The number of neurons beneath a unit area of cortical surface varies linearly with neuronal density, a parameter that is neither related to cortical size nor total number of neurons. Our finding of a variable number of neurons underneath a unit area of the cerebral cortex across primate species indicates that models of cortical organization cannot assume that cortical columns in different primates consist of invariant numbers of neurons.