Activity Of Neurons In Layers Research Articles

General anesthesia and the cortical stranglehold on consciousness

In this issue of Neuron, Bharioke etal. (2022) demonstrate that diverse general anesthetic regimens all reversibly and selectively synchronize spontaneous activity of pyramidal neurons in layer 5 of mouse cortex. We discuss the implications of these findings for the mechanism of consciousness and anesthetic-induced unconsciousness.

An increase in spontaneous activity mediates visual habituation.

SUMMARYThe cerebral cortex is spontaneously active, but the function of this ongoing activity remains unclear. To test whether spontaneous activity encodes learned experiences, we measured the response of neuronal populations in mouse primary visual cortex with chronic two-photon calcium imaging during visual habituation to a specific oriented stimulus. We find that, during habituation, spontaneous activity increases in neurons across the full range of orientation selectivity, eventually matching that of evoked levels. This increase in spontaneous activity robustly correlates with the degree of habituation. Moreover, boosting spontaneous activity with two-photon optogenetic stimulation to the levels of visually evoked activity accelerates habituation. Our study shows that cortical spontaneous activity is linked to habituation, and we propose that habituation unfolds by minimizing the difference between spontaneous and stimulus-evoked activity levels. We conclude that baseline spontaneous activity could gate incoming sensory information to the cortex based on the learned experience of the animal.

Neuronal activity reorganization in motor cortex for successful locomotion after a lesion in the ventrolateral thalamus.

Thalamic stroke leads to ataxia if the cerebellum-receiving ventrolateral thalamus (VL) is affected. The compensation mechanisms for this deficit are not well understood, particularly the roles that single neurons and specific neuronal subpopulations outside the thalamus play in recovery. The goal of this study was to clarify neuronal mechanisms of the motor cortex involved in mitigation of ataxia during locomotion when part of the VL is inactivated or lesioned. In freely ambulating cats, we recorded the activity of neurons in layer V of the motor cortex as the cats walked on a flat surface and horizontally placed ladder. We first reversibly inactivated ∼10% of the VL unilaterally using glutamatergic transmission antagonist CNQX and analyzed how the activity of motor cortex reorganized to support successful locomotion. We next lesioned 50%-75% of the VL bilaterally using kainic acid and analyzed how the activity of motor cortex reorganized when locomotion recovered. When a small part of the VL was inactivated, the discharge rates of motor cortex neurons decreased, but otherwise the activity was near normal, and the cats walked fairly well. Individual neurons retained their ability to respond to the demand for accuracy during ladder locomotion; however, most changed their response. When the VL was lesioned, the cat walked normally on the flat surface but was ataxic on the ladder for several days after lesion. When ladder locomotion normalized, neuronal discharge rates on the ladder were normal, and the shoulder-related group was preferentially active during the stride's swing phase.NEW & NOTEWORTHY This is the first analysis of reorganization of the activity of single neurons and subpopulations of neurons related to the shoulder, elbow, or wrist, as well as fast- and slow-conducting pyramidal tract neurons in the motor cortex of animals walking before and after inactivation or lesion in the thalamus. The results offer unique insights into the mechanisms of spontaneous recovery after thalamic stroke, potentially providing guidance for new strategies to alleviate locomotor deficits after stroke.

Tracking calcium dynamics from individual neurons in behaving animals.

Measuring the activity of neuronal populations with calcium imaging can capture emergent functional properties of neuronal circuits with single cell resolution. However, the motion of freely behaving animals, together with the intermittent detectability of calcium sensors, can hinder automatic monitoring of neuronal activity and their subsequent functional characterization. We report the development and open-source implementation of a multi-step cellular tracking algorithm (Elastic Motion Correction and Concatenation or EMC2) that compensates for the intermittent disappearance of moving neurons by integrating local deformation information from detectable neurons. We demonstrate the accuracy and versatility of our algorithm using calcium imaging data from two-photon volumetric microscopy in visual cortex of awake mice, and from confocal microscopy in behaving Hydra, which experiences major body deformation during its contractions. We quantify the performance of our algorithm using ground truth manual tracking of neurons, along with synthetic time-lapse sequences, covering a wide range of particle motions and detectability parameters. As a demonstration of the utility of the algorithm, we monitor for several days calcium activity of the same neurons in layer 2/3 of mouse visual cortex in vivo, finding significant turnover within the active neurons across days, with only few neurons that remained active across days. Also, combining automatic tracking of single neuron activity with statistical clustering, we characterize and map neuronal ensembles in behaving Hydra, finding three major non-overlapping ensembles of neurons (CB, RP1 and RP2) whose activity correlates with contractions and elongations. Our results show that the EMC2 algorithm can be used as a robust and versatile platform for neuronal tracking in behaving animals.

Long-term stability of cortical ensembles.

Neuronal ensembles, coactive groups of neurons found in spontaneous and evoked cortical activity, are causally related to memories and perception, but it is still unknown how stable or flexible they are over time. We used two-photon multiplane calcium imaging to track over weeks the activity of the same pyramidal neurons in layer 2/3 of the visual cortex from awake mice and recorded their spontaneous and visually evoked responses. Less than half of the neurons remained active across any two imaging sessions. These stable neurons formed ensembles that lasted weeks, but some ensembles were also transient and appeared only in one single session. Stable ensembles preserved most of their neurons for up to 46 days, our longest imaged period, and these 'core' cells had stronger functional connectivity. Our results demonstrate that neuronal ensembles can last for weeks and could, in principle, serve as a substrate for long-lasting representation of perceptual states or memories.

Experience-dependent spatial expectations in mouse visual cortex.

In generative models of brain function, internal representations are used to generate predictions of sensory input, yet little is known about how internal models influence sensory processing. Here we show that, with experience in a virtual environment, the activity of neurons in layer 2/3 of mouse primary visual cortex (V1) becomes increasingly informative of spatial location. We found that a subset of V1 neurons exhibited responses that were predictive of the upcoming visual stimulus in a spatially dependent manner and that the omission of an expected stimulus drove strong responses in V1. Stimulus-predictive responses also emerged in V1-projecting anterior cingulate cortex axons, suggesting that anterior cingulate cortex serves as a source of predictions of visual input to V1. These findings are consistent with the hypothesis that visual cortex forms an internal representation of the visual scene based on spatial location and compares this representation with feed-forward visual input.

ProBDNF and p75NTR Control Excitability and Persistent Firing of Cortical Pyramidal Neurons.

Persistent firing of entorhinal cortex (EC) pyramidal neurons is required for working memory. We report here that pro-brain-derived neurotrophic factor (proBDNF) activates p75NTR to induce a Rac1-dependent and phosphatidylinositol 4,5-bisphosphate-dependent signaling cascade that suppresses persistent activity. Conversely, using loss-of-function approaches, we find that endogenous proBDNF or p75NTR activation strongly decreases pyramidal neuron excitability and persistent firing, suggesting that a physiological role of this proBDNF-p75NTR cascade may be to regulate working memory in vivo. Consistent with this, mice rendered null for p75NTR during adulthood show improvements in working memory but also display an increased propensity for severe seizures. We propose that by attenuating EC network performance, the proBDNF-p75NTR signaling cascade reduces the probability of epileptogenesis.

The Functional Microarchitecture of the Mouse Barrel Cortex

Cortical maps, consisting of orderly arrangements of functional columns, are a hallmark of the organization of the cerebral cortex. However, the microorganization of cortical maps at the level of single neurons is not known, mainly because of the limitations of available mapping techniques. Here, we used bulk loading of Ca2+ indicators combined with two-photon microscopy to image the activity of multiple single neurons in layer (L) 2/3 of the mouse barrel cortex in vivo. We developed methods that reliably detect single action potentials in approximately half of the imaged neurons in L2/3. This allowed us to measure the spiking probability following whisker deflection and thus map the whisker selectivity for multiple neurons with known spatial relationships. At the level of neuronal populations, the whisker map varied smoothly across the surface of the cortex, within and between the barrels. However, the whisker selectivity of individual neurons recorded simultaneously differed greatly, even for nearest neighbors. Trial-to-trial correlations between pairs of neurons were high over distances spanning multiple cortical columns. Our data suggest that the response properties of individual neurons are shaped by highly specific subcolumnar circuits and the momentary intrinsic state of the neocortex.

The functional microarchitecture of the mouse barrel cortex

Cortical maps, consisting of orderly arrangements of functional columns, are a hallmark of the organization of the cerebral cortex. However, the microorganization of cortical maps at the level of single neurons is not known, mainly because of the limitations of available mapping techniques. Here, we used bulk loading of Ca(2+) indicators combined with two-photon microscopy to image the activity of multiple single neurons in layer (L) 2/3 of the mouse barrel cortex in vivo. We developed methods that reliably detect single action potentials in approximately half of the imaged neurons in L2/3. This allowed us to measure the spiking probability following whisker deflection and thus map the whisker selectivity for multiple neurons with known spatial relationships. At the level of neuronal populations, the whisker map varied smoothly across the surface of the cortex, within and between the barrels. However, the whisker selectivity of individual neurons recorded simultaneously differed greatly, even for nearest neighbors. Trial-to-trial correlations between pairs of neurons were high over distances spanning multiple cortical columns. Our data suggest that the response properties of individual neurons are shaped by highly specific subcolumnar circuits and the momentary intrinsic state of the neocortex.

Layer-Specific Touch-Dependent Facilitation and Depression in the Somatosensory Cortex during Active Whisking

Brains adapt to new situations by retuning their neurons. The most common form of neuronal adaptation, typically observed with repetitive stimulations of passive sensory organs, is depression (responses gradually decrease until stabilized). We studied cortical adaptation when stimuli are acquired by active movements of the sensory organ. In anesthetized rats, artificial whisking was induced at 5 Hz, and activity of individual neurons in layers 2-5 was recorded during whisking in air (Whisking condition) and whisking against an object (Touch condition). Response strengths were assessed by spike counts. Input-layer responses (layers 4 and 5a) usually facilitated during the whisking train, whereas superficial responses (layer 2/3) usually depressed. In layers 2/3 and 4, but not 5a, responses were usually stronger during touch trials than during whisking in air. Facilitations were specific to the protraction phase; during retraction, responses depressed in all layers and conditions. These dynamic processes were accompanied by a slow positive wave of activity progressing from superficial to deeper layers and lasting for approximately 1 s, during the transient phase of response. Our results indicate that, in the cortex, adaptation does not depend only on the level of activity or the frequency of its repetition but rather on the nature of the sensory information that is conveyed by that activity and on the processing layer. The input and laminar specificities observed here are consistent with the hypothesis that the paralemniscal layer 5a is involved in the processing of whisker motion, whereas the lemniscal barrels in layer 4 are involved in the processing of object identity.

Neural basis of localized and delocalized fMRI patterns

What connections can we draw between fMRI and neural firing patterns? The basic requirement of differencing in neuroimaging has limitations because subtraction of baseline activity removes an important part of the total activity 1. Our prior studies have quantitatively revealed that most neurons in the ensemble (in a voxel) contribute differently before, during, and after the stimulation 2. Since the ensemble activity of pyramidal neurons in layer 4 depends on sensory (or localized) input by the thalamus and global (or delocalized) input from other cortical regions, delocalized signals from a global workspace may affect localized stimulus-induced responses. To test the hypothesis that delocalized signals, which have been proposed to include subjective contributions 3, can modulate localized signals we conducted fMRI and electrophysiological studies in rats under varied conditions of anesthesia and forepaw stimulation. The fMRI experiments of -chloralose (40 mg/kg/hr) and halothane (0.7%) anesthetized rats were conducted on 7.0 T or 9.4 T systems using calibrated fMRI 1, 2. The electrical activity was measured by Tungsten microelectodes. The extracellular signals were filtered to separate field and action potentials (FP, AP), where the AP data were binned for spiking frequency. While the fMRI data from both baselines (i.e., halothane and -chloralose) showed sensory-induced changes in the contralateral primary somatosensory and motor areas, there were increased delocalized activities observed with (light) halothane anesthesia. These regions were ipsilateral primary somatosensory, contralateral secondary somatosensory, as well as perirhinal and retrosplenial agranular areas. In contrast signals from delocalized regions were notably absent with (deep) -chloralose anesthesia. These results were supported by correlated changes in AP and FP time courses. The spiking frequency distributions measured from the most activated foci in the contralateral (and ipsilateral) primary somatosensory area(s). The contralateral distributions with (deep) -chloralose anesthesia showed a significant shift from low to high frequency values upon stimulation. The contralateral distribution shift was less noteworthy with (light) halothane anesthesia. The distributions for the baselines with halothane and -chloralose were significantly different, in which the degrees of global inputs assigned to higher frequencies dominated under (light) halothane anesthesia. The contralateral distributions upon stimulation with halothane and -chloralose were also significantly different, suggesting that under the (light) halothane anesthesia level the global inputs dominated whereas local changes were dominated under the (deep) -chloralose anesthesia. Since neuronal populations in different regions do not function as modules, the resultant fMRI or electrophysiological activity patterns arise from interactions of afferent and efferent connections – the latter of which was modulated by anesthesia in this study. These results provide the initial steps to explore the interactions of localized and delocalized underpinnings of the concept of the global workspace model 3 (See Figure 1).

Interaction of sensory responses with spontaneous depolarization in layer 2/3 barrel cortex.

The rodent primary somatosensory cortex is spontaneously active in the form of locally synchronous membrane depolarizations (UP states) separated by quiescent hyperpolarized periods (DOWN states) both under anesthesia and during quiet wakefulness. In vivo whole-cell recordings and tetrode unit recordings were combined with voltage-sensitive dye imaging to analyze the relationship of the activity of individual pyramidal neurons in layer 2/3 to the ensemble spatiotemporal dynamics of the spontaneous depolarizations. These were either brief and localized to an area of a barrel column or occurred as propagating waves dependent on local glutamatergic synaptic transmission in layer 2/3. Spontaneous activity inhibited the sensory responses evoked by whisker deflection, accounting almost entirely for the large trial-to-trial variability of sensory-evoked postsynaptic potentials and action potentials. Subthreshold sensory synaptic responses evoked while a cortical area was spontaneously depolarized were smaller, briefer and spatially more confined. Surprisingly, whisker deflections evoked fewer action potentials during the spontaneous depolarizations despite neurons being closer to threshold. The ongoing spontaneous activity thus regulates the amplitude and the time-dependent spread of the sensory response in layer 2/3 barrel cortex.

Transform-invariant recognition by association in a recurrent network.

Objects can be recognized independently of the view they present, of their position on the retina, or their scale. It has been suggested that one basic mechanism that makes this possible is a memory effect, or a trace, that allows associations to be made between consecutive views of one object. In this work, we explore the possibility that this memory trace is provided by the sustained activity of neurons in layers of the visual pathway produced by an extensive recurrent connectivity. We describe a model that contains this high recurrent connectivity and synaptic efficacies built with contributions from associations between pairs of views that is simple enough to be treated analytically. The main result is that there is a change of behavior as the strength of the association between views of the same object, relative to the association within each view of an object, increases. When its value is small, sustained activity in the network is produced by the views themselves. As it increases above a threshold value, the network always reaches a particular state (which represents the object) independent of the particular view that was seen as a stimulus. In this regime, the network can still store an extensive number of objects, each defined by a finite (although it can be large) number of views.

The fine structural demonstration of monoaminergic synapses in immature rat neocortex

The radial distribution of monoaminergic (MA) synapses has been studied in lateral neocortex (SI) of infant rats using an electron microscopic histochemical technique. Systemically administered 5-hydroxydopamine crosses the blood-brain barrier and produces small granular vesicles (SGV) in 30% of all synaptic terminals in SI. In the primordium of layer IV, at six days postnatal, 70% of the synaptic terminals contain SGVs. Immature cortex thus contains a high proportion of synapses with a vesicular uptake-storage mechanism for catecholamines. It is proposed that in infant rat there is a major projection from brain stem MA nuclei to the neocortex; this projection may exert a potent influence on the activity of neurons in layer IV.

Change of activity in visual cortical units during electrocortical desynchronization in rabbits

1. Most (90%) recorded neurons had background activity of a continuous type and were uniformly distributed in all layers of the cortex, while a minority (10%) discharged in groups of spikes and were located mainly in deep layers of the cortex. 2. Neurons with a predominantly continuous type of background activity, with firing rates of 5–17/sec, showed both excitatory and inhibitory types of response, in the proportion of 2:1 respectively, to desychronization of the ECoG. Neurons of the middle cortical layers responded mainly by excitation, and in the deep layers by inhibition. 3. Spread of ECoG desynchronization was not blocked by undercutting the area of cortex. The undercutting, however, selectively depressed both background and evoked activity of neurons in layers IV and V, producing a relative increase in activity of neurons in the upper layers. 4. Statistical characteristics of the relationship between responses of spike activity of simultaneously recorded neurons showed no significant change after the underoutting operation, indicating the importance of intracortical systems of relations for maintaining functional organization of cortical neurons in working groups.