Aberrant activity of mitochondrial NCLX is linked to impaired synaptic transmission and is associated with mental retardation.

Calcium dynamics control synaptic transmission. Calcium triggers synaptic vesicle fusion, determines release probability, modulates vesicle recycling, participates in long-term plasticity and regulates cellular metabolism. Mitochondria, the main source of cellular energy, serve as calcium signaling hubs. Mitochondrial calcium transients are primarily determined by the balance between calcium influx, mediated by the mitochondrial calcium uniporter (MCU), and calcium efflux through the sodium/lithium/calcium exchanger (NCLX). We identified a human recessive missense SLC8B1 variant that impairs NCLX activity and is associated with severe mental retardation. On this basis, we examined the effect of deleting NCLX in mice on mitochondrial and synaptic calcium homeostasis, synaptic activity, and plasticity. Neuronal mitochondria exhibited basal calcium overload, membrane depolarization, and a reduction in the amplitude and rate of calcium influx and efflux. We observed smaller cytoplasmic calcium transients in the presynaptic terminals of NCLX-KO neurons, leading to a lower probability of release and weaker transmission. In agreement, synaptic facilitation in NCLX-KO hippocampal slices was enhanced. Importantly, deletion of NCLX abolished long term potentiation of Schaffer collateral synapses. Our results show that NCLX controls presynaptic calcium transients that are crucial for defining synaptic strength as well as short- and long-term plasticity, key elements of learning and memory processes.

A Socially Assistive Robot for Stroke Patients: Acceptance, Needs, and Concerns of Patients and Informal Caregivers.

Stroke patients often contend with long-term physical challenges that require treatment and support from both formal and informal caregivers. Socially Assistive Robots (SARs) can assist patients in their physical rehabilitation process and relieve some of the burden on the informal caregivers, such as spouses and family members. We collected and analyzed information from 23 participants (11 stroke patients and 12 informal caregivers) who participated in a total of six focus-group discussions. The participants responded to questions regarding using a SAR to promote physical exercises during the rehabilitation process: (a) the advantages and disadvantages of doing so; (b) specific needs that they wish a SAR would address; (c) patient-specific adaptations they would propose to include; and (d) concerns they had regarding the use of such technology in stroke rehabilitation. We found that the majority of the participants in both groups were interested in experiencing the use of a SAR for rehabilitation, in the clinic and at home. Both groups noted the advantage of having the constant presence of a motivating entity with whom they can practice their rehabilitative exercises. The patients noted how such a device can assist formal caregivers in managing their workload, while the informal caregivers indicated that such a system could ease their own workload and sense of burden. The main disadvantages that participants noted related to the robot not possessing human abilities, such as the ability to hold a conversation, to physically guide the patient's movements, and to express or understand emotions. We anticipate that the data collected in this study-input from the patients and their family members, including the similarities and differences between their points of view-will aid in improving the development of SARs for rehabilitation, so that they can better suit people who have had a stroke, and meet their individual needs.

Auto-correlated directional swimming can enhance settlement success and connectivity in fish larvae.

Larvae of coastal-marine fishes have been shown repeatedly to swim directionally in the pelagic environment. Yet, biophysical models of larval dispersal typically impose a Simple Random Walk (SRW) algorithm to simulate non-directional movement in the open ocean. Here we investigate the use of a Correlated Random Walk (CRW) algorithm; imposing auto-correlated directional swimming onto simulated larvae within a high-resolution 3D biophysical model of the Gulf of Aqaba, the Red Sea. Our findings demonstrate that implementation of auto-correlated directional swimming can result in an increase of up to ×2.7 in the estimated success rate of larval-settlement, as well as an increase in the extent of connectivity. With accumulating empirical support for the capacity for directional-swimming during the pelagic phase, we propose that CRW should be applied in biophysical models of dispersal by coastal marine fish-larvae.

Long-range intralaminar noise correlations in the barrel cortex.

Identifying the properties of correlations in the firing of neocortical neurons is central to our understanding of cortical information processing. It has been generally assumed, by virtue of the columnar organization of the neocortex, that the firing of neurons residing in a certain vertical domain is highly correlated. On the other hand, firing correlations between neurons steeply decline with horizontal distance. Technical difficulties in sampling neurons with sufficient spatial information have precluded the critical evaluation of these notions. We used 128-channel "silicon probes" to examine the spike-count noise correlations during spontaneous activity between multiple neurons with identified laminar position and over large horizontal distances in the anesthetized rat barrel cortex. Eigen decomposition of correlation coefficient matrices revealed that the laminar position of a neuron is a significant determinant of these correlations, such that the fluctuations of layer 5B/6 neurons are in opposite direction to those of layers 5A and 4. Moreover, we found that within each experiment, the distribution of horizontal, intralaminar spike-count correlation coefficients, up to a distance of ∼1.5 mm, is practically identical to the distribution of vertical correlations. Taken together, these data reveal that the neuron's laminar position crucially affects its role in cortical processing. Moreover, our analyses reveal that this laminar effect extends over several functional columns. We propose that within the cortex the influence of the horizontal elements exists in a dynamic balance with the influence of the vertical domain and this balance is modulated with brain states to shape the network's behavior.

Spatiotemporal alterations of cortical network activity by selective loss of NOS-expressing interneurons.

Deciphering the role of GABAergic neurons in large neuronal networks such as the neocortex forms a particularly complex task as they comprise a highly diverse population. The neuronal isoform of the enzyme nitric oxide synthase (nNOS) is expressed in the neocortex by specific subsets of GABAergic neurons. These neurons can be identified in live brain slices by the nitric oxide (NO) fluorescent indicator diaminofluorescein-2 diacetate (DAF-2DA). However, this indicator was found to be highly toxic to the stained neurons. We used this feature to induce acute phototoxic damage to NO-producing neurons in cortical slices, and measured subsequent alterations in parameters of cellular and network activity. Neocortical slices were briefly incubated in DAF-2DA and then illuminated through the 4× objective. Histochemistry for NADPH-diaphorase (NADPH-d), a marker for nNOS activity, revealed elimination of staining in the illuminated areas following treatment. Whole cell recordings from several neuronal types before, during, and after illumination confirmed the selective damage to non-fast-spiking (FS) interneurons. Treated slices displayed mild disinhibition. The reversal potential of compound synaptic events on pyramidal neurons became more positive, and their decay time constant was elongated, substantiating the removal of an inhibitory conductance. The horizontal decay of local field potentials (LFPs) was significantly reduced at distances of 300-400 µm from the stimulation, but not when inhibition was non-selectively weakened with the GABA(A) blocker picrotoxin. Finally, whereas the depression of LFPs along short trains of 40 Hz stimuli was linearly reduced with distance or initial amplitude in control slices, this ordered relationship was disrupted in DAF-treated slices. These results reveal that NO-producing interneurons in the neocortex convey lateral inhibition to neighboring columns, and shape the spatiotemporal dynamics of the network's activity.

Getting drowsy? Alert/nonalert transitions and visual thalamocortical network dynamics.

The effects of different EEG brain states on spontaneous firing of cortical populations are not well understood. Such state shifts may occur frequently under natural conditions, and baseline firing patterns can impact neural coding (e.g., signal-to-noise ratios, sparseness of coding). Here, we examine the effects of spontaneous transitions from alert to nonalert awake EEG states in the rabbit visual cortex (5 s before and after the state-shifts). In layer 4, we examined putative spiny neurons and fast-spike GABAergic interneurons; in layer 5, we examined corticotectal neurons. We also examined the behavior of retinotopically aligned dorsal lateral geniculate nucleus (LGNd) neurons, usually recorded simultaneously with the above cortical populations. Despite markedly reduced firing and sharply increased bursting in the LGNd neurons following the transition to the nonalert state, little change occurred in the spiny neurons of layer 4. However, fast-spike neurons of layer 4 showed a paradoxical increase in firing rates as thalamic drive decreased in the nonalert state, even though some of these cells received potent monosynaptic input from the same LGNd neurons whose rates were reduced. The firing rates of corticotectal neurons of layer 5, similarly to spiny cells of layer 4, were not state-dependent, but these cells did become more bursty in the nonalert state, as did the fast-spike cells. These results show that spontaneous firing rates of midlayer spiny populations are remarkably conserved following the shift from alert to nonalert states, despite marked reductions in excitatory thalamic drive and increased activity in local fast-spike inhibitory interneurons.

Physiologic role for "inducible" nitric oxide synthase: a new form of astrocytic-neuronal interface.

Nitric oxide (NO) has been long recognized as an atypical neuronal messenger affecting excitatory synaptic transmission, but its cellular source has remained unresolved as the neuronal isoform of NO synthase (nNOS) in many brain regions is expressed only by small subsets of inhibitory neurons. It is generally believed that the glial NO-producing isoform (iNOS) is not expressed in the normal brain, but rather it undergoes a transcription-mediated up-regulation following an immunological challenge. Therefore, the involvement of iNOS in modulating normal neuronal functions has been largely ignored. Here I review evidence to the contrary: I summarize data pointing to the existence of a functioning iNOS in normal undisturbed mammalian brains, and experimental results tracing this expression to astrocytes. Finally, I review recent findings asserting that iNOS-dependent NO modulates synaptic release from presynaptic terminals. Based on these data, I propose that astrocytes express basal levels of iNOS. Flanking synaptic elements, astrocytes are perfectly positioned to release NO and affect synaptic transmission.

Astrocytic iNOS-dependent enhancement of synaptic release in mouse neocortex.

Nitric oxide (NO) has been recognized as an atypical neuronal messenger affecting synaptic transmission, but its cellular source has remained unresolved as the neuronal NO synthase isoform (nNOS) in brain areas such as the neocortex is expressed only by a small subset of inhibitory neurons. The involvement of the glial NOS isoform (iNOS) in modulating neuronal activity has been largely ignored because it has been accepted that this enzyme is regulated by gene induction following detrimental stimuli. Using acute brain slices from mouse neocortex and electrophysiology, we found that selective inhibition of iNOS reduced both spontaneous and evoked synaptic release. Moreover, iNOS inhibition partially prevented and reversed the potentiation of excitatory synapses in layer 2/3 pyramidal neurons. NOS enzymatic assay confirmed a small but reliable Ca(2+)-independent activity fraction, consistent with the existence of functioning iNOS in the tissue. Together these data point to astrocytes as a source for the nitrosative regulation of synaptic release in the neocortex.

A dynamic zone defines interneuron remodeling in the adult neocortex.

The contribution of structural remodeling to long-term adult brain plasticity is unclear. Here, we investigate features of GABAergic interneuron dendrite dynamics and extract clues regarding its potential role in cortical function and circuit plasticity. We show that remodeling interneurons are contained within a "dynamic zone" corresponding to a superficial strip of layers 2/3, and remodeling dendrites respect the lower border of this zone. Remodeling occurs primarily at the periphery of dendritic fields with addition and retraction of new branch tips. We further show that dendrite remodeling is not intrinsic to a specific interneuron class. These data suggest that interneuron remodeling is not a feature predetermined by genetic lineage, but rather, it is imposed by cortical laminar circuitry. Our findings are consistent with dynamic GABAergic modulation of feedforward and recurrent connections in response to top-down feedback and suggest a structural component to functional plasticity of supragranular neocortical laminae.

Enhanced astrocytic nitric oxide production and neuronal modifications in the neocortex of a NOS2 mutant mouse.

BACKGROUND: It has been well accepted that glial cells in the central nervous system (CNS) produce nitric oxide (NO) through the induction of a nitric oxide synthase isoform (NOS2) only in response to various insults. Recently we described rapid astroglial, NOS2-dependent, NO production in the neocortex of healthy mice on a time scale relevant to neuronal activity. To explore a possible role for astroglial NOS2 in normal brain function we investigated a NOS2 knockout mouse (B6;129P2-Nos2(tm1Lau)/J, Jackson Laboratory). Previous studies of this mouse strain revealed mainly altered immune responses, but no compensatory pathways and no CNS abnormalities have been reported. METHODOLOGY/PRINCIPAL FINDINGS: To our surprise, using NO imaging in brain slices in combination with biochemical methods we uncovered robust NO production by neocortical astrocytes of the NOS2 mutant. These findings indicate the existence of an alternative pathway that increases basal NOS activity. In addition, the astroglial mutation instigated modifications of neuronal attributes, shown by changes in the membrane properties of pyramidal neurons, and revealed in distinct behavioral abnormalities characterized by an increase in stress-related parameters. CONCLUSIONS/SIGNIFICANCE: The results strongly indicate the involvement of astrocytic-derived NO in modifying the activity of neuronal networks. In addition, the findings corroborate data linking NO signaling with stress-related behavior, and highlight the potential use of this genetic model for studies of stress-susceptibility. Lastly, our results beg re-examination of previous studies that used this mouse strain to examine the pathophysiology of brain insults, assuming lack of astrocytic nitrosative reaction.

Mechanisms of firing patterns in fast-spiking cortical interneurons.

Cortical fast-spiking (FS) interneurons display highly variable electrophysiological properties. Their spike responses to step currents occur almost immediately following the step onset or after a substantial delay, during which subthreshold oscillations are frequently observed. Their firing patterns include high-frequency tonic firing and rhythmic or irregular bursting (stuttering). What is the origin of this variability? In the present paper, we hypothesize that it emerges naturally if one assumes a continuous distribution of properties in a small set of active channels. To test this hypothesis, we construct a minimal, single-compartment conductance-based model of FS cells that includes transient Na(+), delayed-rectifier K(+), and slowly inactivating d-type K(+) conductances. The model is analyzed using nonlinear dynamical system theory. For small Na(+) window current, the neuron exhibits high-frequency tonic firing. At current threshold, the spike response is almost instantaneous for small d-current conductance, gd, and it is delayed for larger gd. As gd further increases, the neuron stutters. Noise substantially reduces the delay duration and induces subthreshold oscillations. In contrast, when the Na(+) window current is large, the neuron always fires tonically. Near threshold, the firing rates are low, and the delay to firing is only weakly sensitive to noise; subthreshold oscillations are not observed. We propose that the variability in the response of cortical FS neurons is a consequence of heterogeneities in their gd and in the strength of their Na(+) window current. We predict the existence of two types of firing patterns in FS neurons, differing in the sensitivity of the delay duration to noise, in the minimal firing rate of the tonic discharge, and in the existence of subthreshold oscillations. We report experimental results from intracellular recordings supporting this prediction.

Dendritic backpropagation and the state of the awake neocortex.

The spread of somatic spikes into dendritic trees has become central to models of dendritic integrative properties and synaptic plasticity. However, backpropagating action potentials (BPAPs) have been studied mainly in slices, in which they are highly sensitive to multiple factors such as firing frequency and membrane conductance, raising doubts about their effectiveness in the awake behaving brain. Here, we examine the spatiotemporal characteristics of BPAPs in layer 5 pyramidal neurons in the visual cortex of adult, awake rabbits, in which EEG-defined brain states ranged from alert vigilance to drowsy/inattention, and, in some cases, to light sleep. To achieve this, we recorded extracellular spikes from layer 5 pyramidal neurons and field potentials above and below these neurons using a 16-channel linear probe, and applied methods of spike-triggered current source-density analysis to these records (Buzsáki and Kandel, 1998; Swadlow et al., 2002). Precise retinotopic alignment of superficial and deep cortical sites was used to optimize alignment of the recording probe with the axis of the apical dendrite. During the above network states, we studied BPAPs generated spontaneously, antidromically (from corticotectal neurons), or via intense synaptic drive caused by natural visual stimulation. Surprisingly, the invasion of BPAPs as far as 800 microm from the soma was little affected by the network state and only mildly attenuated by high firing frequencies. These data reveal that the BPAP is a robust and highly reliable property of neocortical apical dendrites. These events, therefore, are well suited to provide crucial signals for the control of synaptic plasticity during information-processing brain states.

Time-dependent, layer-specific modulation of sensory responses mediated by neocortical layer 1.

An essential component of feedback and top-down information in the cortical column arrives at layer 1 (L1) where it contacts distal dendrites of pyramidal neurons. Although much is known about the anatomical organization of L1 fibers, their contribution to sensory information processing remains to be determined. We assessed the physiological significance of L1 inputs by performing extracellular recordings in vivo from neurons in the primary somatosensory cortex of rodents. We found that blocking activity in L1 increases whisker-evoked response magnitude and variance, suggesting that L1 exerts an inhibitory influence on whisker responses. However, when pairing L1 stimulation with whisker deflection, the interval between the stimuli determined the outcome of the interaction, with facilitation of sensory responses dominating the short intervals (10 ms). These temporal interactions resulted in a time-dependent regulation of direction tuning of cortical neurons. The synaptic mechanisms underlying L1 inputs' influences were examined using whole cell recordings in vitro while pairing L1 and white-matter stimulations. We found time-dependent, layer-specific differences in synaptic summation of the two inputs, with supralinearity at shorter intervals and sublinearity at longer intervals that resulted mainly from shunting inhibition. Taken together, our results demonstrate that L1 inputs impose a time- and layer-specific regulation on sensory-evoked responses. This in turn may lead to a dynamic transmission of sensory information in the somatosensory cortex.

Rapid and reactive nitric oxide production by astrocytes in mouse neocortical slices.

Nitric oxide (NO), a cellular signaling molecule, is produced in the brain by both neurons and astrocytes. While neurons are capable of rapid release of small amounts of NO serving as neurotransmitter, astrocytic NO production has been demonstrated mainly as a slow reaction to various stress stimuli. Little is known about the role of astrocyte-produced NO. Using the NO indicator 4,5-diaminofluorescein-2 diacetate (DAF-2DA) and acute slices from mouse brain, we distinguished neurons from astrocytes based on their different fluorescence kinetics and pattern, cellular morphology, electrophysiology, and responses to selective nitric oxide synthase (NOS) inhibitors. Typically, astrocytic fluorescence followed neuronal fluorescence with a delay of 1-2 min and was dependent on the inducible NOS isoform (iNOS) activity. Western blot analysis established the presence of functional iNOS in the neocortex. An assay for cell death revealed that most DAF-2DA-positive neurons, but not astrocytes, were damaged. Whole cell recordings from astrocytes confirmed that these cells maintained their membrane potential and passive properties during illumination and afterward. Induction of excitotoxicity by brief application of glutamate triggered an immediate and intense astrocytic response, while high-frequency electrical stimulation failed to do so. The present study demonstrates, for the first time, rapid and massive iNOS-dependent NO production by astrocytes in situ, which appears to be triggered by acute neuronal death. These data may bear important implications for our theoretical understanding and practical management of acute brain insults.

Inhibitory effect of mouse neocortex layer I on the underlying cellular network.

The mammalian cortical layer I is a convergence site for axons of sub- and intracortical origin, and the apical dendritic tufts of pyramidal neurons. A prominent feature of layer I is an extensive plexus of inhibitory axons, which originate from stellate cells in all cortical laminae. The role of this inhibitory projection in the activity of cortical networks has yet to be determined. We investigated the degree to which inhibitory inputs within layer I affect the activity of the underlying cellular network. Field potentials (FPs) were recorded in layer II/III. Focal application of the GABAA blocker picrotoxin in layer I above the recording pipette or the removal of layer I resulted in larger FP amplitudes for stimulations at control-equal intensities. When inhibition was partially blocked, the removal of layer I caused a significant reduction in the threshold stimulus intensity required for generating epileptiform events, and a rise in the propagation velocity of these events. Immunocytochemistry for chemical markers of interneurons proved that the inhibitory input to layer I is predominantly somatostatin immunoreactive (SM-ir), such that layer I contains approximately one-third of all SM-ir axons in the cortex. Calretinin-immunoreactive axons were also present in layer I at a lower density. We conclude that the impact of layer I on the cortical cellular network includes a significant inhibitory component. This inhibition confers a moderate restraining influence, and its removal increases the excitability of cortical circuits, but not sufficiently to induce epileptic phenomena.

The spatial dimensions of electrically coupled networks of interneurons in the neocortex.

Inhibitory interneurons of the neocortex are electrically coupled to cells of the same type through gap junctions. We studied the spatial organization of two types of interneurons in the rat somatosensory cortex: fast-spiking (FS) parvalbumin-immunoreactive (PV+) cells, and low threshold-spiking (LTS) somatostatin-immunoreactive (SS+) cells. Paired recordings in layer 4 demonstrated that both the probability of coupling and the coupling coefficient drop steeply with intersomatic distance, reaching zero beyond 200 microm. The dendritic arbors of FS and LTS cells were reconstructed from electrophysiologically characterized, biocytin-filled cells; the two cell types had only minor differences in the number and span of their dendrites. However, there was a markedly higher density of PV+ cells than SS+ cells. PV+ cells were densest in layer 4, while SS+ cell density peaked in the subgranular layers. From these data we estimate that there is measurable electrical coupling (directly or indirectly via intermediary cells) between each interneuron and 20-50 others. The large number of electrical synapses implies that each interneuron participates in a large, continuous syncytium. To evaluate the functional significance of these findings, we examined several simple architectures of coupled networks analytically. We present a mathematical method to estimate the average summated coupling conductance that each cell receives from all of its neighbors, and the average leak conductance of individual cells, and we suggest that these have the same order of magnitude. These quantitative results have important implications for the effects of electrical coupling on the dynamic behavior of interneuron networks.