Plasticity of recurring spatiotemporal activity patterns in cortical networks

How do neurons encode and store information for long periods of time? Recurring patterns of activity have been reported in various cortical structures and were suggested to play a role in information processing and memory. To study the potential role of bursts of action potentials in memory mechanisms, we investigated patterns of spontaneous multi-single-unit activity in dissociated rat cortical cultures in vitro. Spontaneous spikes were recorded from networks of approximately 50 000 neurons and glia cultured on a grid of 60 extracellular substrate- embedded electrodes (multi-electrode arrays). These networks expressed spontaneous culture- wide bursting from approximately one week in vitro. During bursts, a large portion of the active electrodes showed elevated levels of firing. Spatiotemporal activity patterns within spontaneous bursts were clustered using a correlation-based clustering algorithm, and the occurrences of these burst clusters were tracked over several hours. This analysis revealed spatiotemporally diverse bursts occurring in well-defined patterns, which remained stable for several hours. Activity evoked by strong local tetanic stimulation resulted in significant changes in the occurrences of spontaneous bursts belonging to different clusters, indicating that the dynamical flow of information in the neuronal network had been altered. The diversity of spatiotemporal structure and long-term stability of spontaneous bursts together with their plastic nature strongly suggests that such network patterns could be used as codes for information transfer and the expression of memories stored in cortical networks.     

Facilitating action of medial prefrontal cortex upon the noxious thermally-evoked responses in thalamic centralis lateralis nucleus.

This paper shows a medial prefrontal cortex (CxAP9) facilitating influence upon the unit activity of the centralis lateralis (Cl) nucleus of the thalamus, in rats anesthetized with urethane. Cortical influences were studied using both cortical cooling and cortical spreading depression (CSD) procedures. Both spontaneous and noxious thermally evoked activities were considered. When CSD was propagated and affected the CxAP9, as well as during the cooling of this area, both spontaneous activity and the responses evoked in Cl cells by noxious stimulation were blocked. This effect was interpreted as a cortical disfacilitation upon Cl cells. During the cortical silent period we tested the excitability of a few Cl cells, provoking their activation by passing electrical current across the same Cl recording electrode. No changes were observed in their excitable response threshold during CSD or cortical cooling. Our results are in agreement with the proposition of a tonic cortical facilitatory action upon the spontaneous and noxious-evoked responses recorded in the Cl cells.     

Activity-dependent enhancement in the reliability of correlated spike timings in cultured cortical neurons

To study the use-dependent modification of activity in neural networks, we investigated the spike timing by simultaneously recording activity at multiple sites in a network of cultured cortical neurons. We used dynamical analysis to study the temporal structure of spike trains and the activity-dependent changes in the reliability and reproducibility of spike patterns evoked by a stimulus. We also used cross-correlation analysis to evaluate the interactions of neuron pairs. Our main conclusions are that even when no obvious change in spike numbers can be seen, use-dependent modification occurs, either enhancing or reducing in the reliability and reproducibility of spike trains evoked by a stimulus, and the fine temporal structure of stimulus-evoked spike trains and interactions between neurons are also modified by tetanic stimulation. Received: 25 February 1998 / Accepted in revised form: 24 August 1998     

Homeostatic plasticity of GABA-ergic synaptic transmission in rat hippocampal cell cultures

It has been shown recently that prolonged blockade of neuronal firing activates several homeostatic mechanisms in neocortical networks, including alteration of glutamatergic and GABA-ergic synaptic transmission, and postsynaptic changes are involved in both cases. We studied whether such treatment also affects GABA-ergic synaptic transmission in hippocampal cell cultures. Using whole-cell voltage-clamp recording and local extracellular stimulation, we investigated evoked inhibitory postsynaptic currents (IPSC) in cultured rat hippocampal neurons grown with the sodium channel blocker tetrodotoxin (TTX) and under control conditions. We found that chronic TTX treatment significantly decreased the amplitude of evoked IPSC. This decrease was accompanied by an increase in the coefficient of variation of the above parameter, which is suggestive of a presynaptic mechanism. In contrast, no changes in the IPSC reversal potential or paired-pulse depression were observed in TTX-treated cultures. We conclude that alteration of GABA-ergic synaptic transmission contributes to the homeostatic plasticity in hippocampal neuronal networks, and this change is at least in part due to a presynaptic mechanism.Neirofiziologiya/Neurophysiology, Vol. 36, Nos. 5/6, pp. 432–437, September–December, 2004.This revised version was published online in April 2005 with a corrected cover date and copyright year.     

The Effect of Slow Electrical Stimuli to Achieve Learning in Cultured Networks of Rat Cortical Neurons

Learning, or more generally, plasticity may be studied using cultured networks of rat cortical neurons on multi electrode arrays. Several protocols have been proposed to affect connectivity in such networks. One of these protocols, proposed by Shahaf and Marom, aimed to train the input-output relationship of a selected connection in a network using slow electrical stimuli. Although the results were quite promising, the experiments appeared difficult to repeat and the training protocol did not serve as a basis for wider investigation yet. Here, we repeated their protocol, and compared our ‘learning curves’ to the original results. Although in some experiments the protocol did not seem to work, we found that on average, the protocol showed a significantly improved stimulus response indeed. Furthermore, the protocol always induced functional connectivity changes that were much larger than changes that occurred after a comparable period of random or no stimulation. Finally, our data shows that stimulation at a fixed electrode induces functional connectivity changes of similar magnitude as stimulation through randomly varied sites; both larger than spontaneous connectivity fluctuations. We concluded that slow electrical stimulation always induced functional connectivity changes, although uncontrolled. The magnitude of change increased when we applied the adaptive (closed-loop) training protocol. We hypothesize that networks develop an equilibrium between connectivity and activity. Induced connectivity changes depend on the combination of applied stimulus and initial connectivity. Plain stimuli may drive networks to the nearest equilibrium that accommodates this input, whereas adaptive stimulation may direct the space for exploration and force networks to a new balance, at a larger distance from the initial state.     

Microiontophoretic study of the effect of biogenic amines on unit activity in the rabbit visual cortex

The effect of acetylcholine, noradrenalin, and serotonin on spontaneous activity of visual cortical neurons and on their activity evoked by flashes, recorded extracellularly, was studied by microiontophoresis in unanesthetized rabbits. The ability of visual cortical neurons to respond to light does not correlate with their sensitivity to acetylcholine. This substance, which changes the spontaneous firing rate of many of the neurons tested, was less effective against their evoked activity. Noradrenalin had a powerful depressant action on both spontaneous and evoked activity of most neurons studied. Serotonin acted in different ways on the spontaneous and evoked activity of some neurons tested. It is postulated that acetylcholine mediates reticulo-cortical inputs, noradrenalin is a true inhibitory mediator in the cerebral cortex, and serotonin has a presynaptic action by preventing the liberation of natural mediators.     

Low-frequency stimulation induces stable transitions in stereotypical activity in cortical networks

Reverberating spontaneous synchronized brain activity is believed to play an important role in neural information processing. Whether and how external stimuli can influence this spontaneous activity is poorly understood. Because periodic synchronized network activity is also prominent in in vitro neuronal cultures, we used cortical cultures grown on multielectrode arrays to examine how spontaneous activity is affected by external stimuli. Spontaneous network activity before and after low-frequency electrical stimulation was quantified in several ways. Our results show that the initially stable pattern of stereotypical spontaneous activity was transformed into another activity pattern that remained stable for at least 1 h. The transformations consisted of changes in single site and culture-wide network activity as well as in the spatiotemporal dynamics of network bursting. We show for the first time that low-frequency electrical stimulation can induce long-lasting alterations in spontaneous activity of cortical neuronal networks. We discuss whether the observed transformations in network activity could represent a switch in attractor state.     

Epoxy-silane linking of biomolecules is simple and effective for patterning neuronal cultures

Surface chemistry is one of the main factors that contributes to the longevity and compliance of cell patterning. Two to three weeks are required for dissociated, embryonic rat neuronal cultures to mature to the point that they regularly produce spontaneous and evoked responses. Though proper surface chemistry can be achieved through the use of covalent protein attachment, often it is not maintainable for the time periods necessary to study neuronal growth. Here we report a new and effective covalent linking approach using (3-glycidoxypropyl) trimethoxysilane (3-GPS) for creating long term neuronal patterns. Micrometer scale patterns of cell adhesive proteins were formed using microstamping; hippocampal neurons, cultured up to 1 month, followed those patterns. Cells did not grow on unmodified 3-GPS surfaces, producing non-permissive regions for the long-term cell patterning. Patterned neuronal networks were formed on two different types of MEA (polyimide or silicon nitride insulation) and maintained for 3 weeks. Even though the 3-GPS layer increased the impedance of metal electrodes by a factor of 2-3, final impedance levels were low enough that low noise extracellular recordings were achievable. Spontaneous neural activity was recorded as early as 10 days in vitro. Neural recording and stimulation were readily achieved from these networks. Our results showed that 3-GPS could be used on surfaces to immobilize biomolecules for a variety of neural engineering applications.     

6-Hydroxydopamine treatment enhances excitation of cultured cerebellar neurons by glutamate

Explant cultures containing identifiable cerebellar cortical neurons and locus coeruleus neurons were treated with 500 microM 6-hydroxydopamine. At this concentration, locus coeruleus neurons were usually degenerated after 48 h, while the cerebellar cortical neurons had a normal appearance. Extracellular recording and iontophoresis of noradrenaline and glutamate were used to test for changes in electrical activity or neurotransmitter responsiveness of the cerebellar neurons. At 4-5 days following the toxin exposure, spontaneous spiking activity appeared similar to that in control cultures. Mean iontophoretic currents required to give noradrenaline-induced depressions of activity were somewhat lower for the toxin-treated cultures than for controls but not significantly so. The mean currents for glutamate excitations, however, were markedly lower in the treated cultures. Noradrenaline potentiations of glutamate responses were observed in both treated and control cultures. The greatly increased sensitivity of cerebellar neurons to glutamate does not seem related to degeneration of granule cells in the treated cultures but might be explained by disruption of astrocytic uptake mechanisms for glutamate.     

Interaction between influences of functionally different cortical areas on caudate neurons in cats

Unit activity in the caudate nucleus evoked by paired stimulation of the anterior sigmoid and middle suprasylvian gyri was studied in acute experiments on cats. Responses in most neurons to testing stimulation of the anterior sigmoid gyrus during the period of inhibition of spontaneous activity evoked by conditioning stimulation of the suprasylvian gyrus were preserved, but in isolated cases they were actually facilitated. Meanwhile conditioning stimulation of the anterior sigmoid gyrus in the period of inhibition depressed responses to testing stimulation of the suprasylvian gyrus. Similar results were obtained in experiments on animals with deep transcortical sections between the sensomotor and parietal regions, ruling out the possibility of interaction between the stimulated zones at the cortical level.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 13, No. 2, pp. 142–148, March–April, 1981.     

Cortical influences on neurons of the lateral reticular nucleus responding to noxious stimuli

The effect of frontoparietal sensorimotor (FPSM) cortex stimulation on both the spontaneous and the noxious evoked activity of neurons in the lateral reticular nucleus (LRN) was tested in barbiturate-anesthetized rats. Ninety-three LRN neurons that responded to a noxious heat stimulus (HS) were recorded (72% antidromically fired from the cerebellum). Of these, 66 neurons altered their spontaneous firing rates in response to cortical stimulation. Two patterns of responses were found: either an excitation followed by a suppression of spontaneous activity (52 neurons), or a pure suppression of spontaneous activity lasting 50-400 msec (14 neurons). In 46 of these neurons, it was found that cortical stimulation reduced HS-evoked activity to near the baseline level. Furthermore, it was found that when applied after a prolonged cortical stimulation, the HS was ineffective. It is concluded that FPSM cortex can influence nociceptive information in LRN neurons that respond to its stimulation, possibly interfering with the mechanisms underlying stimulation-produced analgesia (SPA). In this context, it is proposed that the cortex can modulate the activity of LRN neurons that activate, through local loops, a descending antinociceptive system and also a separate projection system to the cerebellum.     

Cortical Influences on Neurons of the Lateral Reticular Nucleus Responding to Noxious Stimuli

The effect of frontoparietal sensorimotor (FPSM) cortex stimulation on both the spontaneous and the noxious evoked activity of neurons in the lateral reticular nucleus (LRN) was tested in barbiturate-anesthetized rats. Ninety-three LRN neurons that responded to a noxious heat stimulus (HS) were recorded (72% antidromically fired from the cerebellum). Of these, 66 neurons altered their spontaneous firing rates in response to cortical stimulation. Two patterns of responses were found: either an excitation followed by a suppression of spontaneous activity (52 neurons), or a pure suppression of spontaneous activity lasting 50-400 msec (14 neurons). In 46 of these neurons, it was found that cortical stimulation reduced HS-evoked activity to near the baseline level. Furthermore, it was found that when applied after a prolonged cortical stimulation, the HS was ineffective. It is concluded that FPSM cortex can influence nociceptive information in LRN neurons that respond to its stimulation, possibly interfering with the mechanisms underlying stimulation-produced analgesia (SPA). In this context, it is proposed that the cortex can modulate the activity of LRN neurons that activate, through local loops, a descending antinociceptive system and also a separate projection system to the cerebellum.     

Emergence of a small-world functional network in cultured neurons

The functional networks of cultured neurons exhibit complex network properties similar to those found in vivo. Starting from random seeding, cultures undergo significant reorganization during the initial period in vitro, yet despite providing an ideal platform for observing developmental changes in neuronal connectivity, little is known about how a complex functional network evolves from isolated neurons. In the present study, evolution of functional connectivity was estimated from correlations of spontaneous activity. Network properties were quantified using complex measures from graph theory and used to compare cultures at different stages of development during the first 5 weeks in vitro. Networks obtained from young cultures (14 days in vitro) exhibited a random topology, which evolved to a small-world topology during maturation. The topology change was accompanied by an increased presence of highly connected areas (hubs) and network efficiency increased with age. The small-world topology balances integration of network areas with segregation of specialized processing units. The emergence of such network structure in cultured neurons, despite a lack of external input, points to complex intrinsic biological mechanisms. Moreover, the functional network of cultures at mature ages is efficient and highly suited to complex processing tasks.     

Role of Acetylcholine in the Modulation of Activity of Neocortical Neurons in Awake Animals Performing an Instrumental Conditioned Reflex

We studied modulatory effects of the cholinergic system on the activity of sensorimotor cortex neurons related to realization of an instrumental conditioned placing reflex. Experiments were carried out on awake cats; multibarrel glass microelectrodes were used for extracellular recording of impulse activity of neurons in the sensorimotor cortex and iontophoretic application of synaptically active agents within the recording region. The background and reflex-related activity was recorded in the course of realization of conditioned movements, and then changes of spiking induced by applications of the testing substances were examined. Applications of acetylcholine and carbachol resulted in increases in the intensity of impulse reactions of neocortical neurons evoked by presentation of an acoustic signal and in simultaneous shortening of the response latencies. An agonist of muscarinic receptors, pylocarpine, exerted a similar effect on the evoked activity of sensorimotor cortex neurons. Blockers of muscarinic receptors, atropine and scopolamine, vice versa, sharply suppressed impulse reactions of cortical neurons to afferent stimulation and simultaneously increased latencies of these responses. Applications of an agonist of nicotinic receptors, nicotine, was accompanied by suppression of impulse neuronal responses, an increase in the latency of spike reactions to presentation of a sound signal, and a corresponding increase in the latency of a conditioned motor reaction. In contrast, application of an antagonist of nicotinic receptors, tubocurarine, significantly intensified neuronal spike responses and shortened their latency. The mechanisms underlying the effects of antagonists of membrane muscarinic and nicotinic cholinoreceptors and the role of activation of these receptors in the modulation of activity of pyramidal and non-pyramidal neocortical neurons related to realization of the instrumental motor reflex are discussed.     

The dynamics of a neuronal culture of dissociated cortical neurons of neonatal rats

Neuronal networks of dissociated cortical neurons from neonatal rats were cultured over a multielectrode dish with 64 active sites, which were used both for recording the electrical activity and for stimulation. After about 4 weeks of culture, a dense network of neurons had developed and their electrical activity was studied. When a brief voltage pulse was applied to one extracellular electrode, a clear electrical response was evoked over almost the entire network. When a strong voltage pulse was used, the response was composed of an early phase, terminating within 25 ms, and a late phase which could last several hundreds of milliseconds. Action potentials evoked during the early phase occurred with a precise timing with a small jitter and the electrical activity initiated by a localized stimulation diffused significantly over the network. In contrast, the late phase was characterized by the occurrence of clusters of electrical activity with significant spatio-temporal fluctuations. The late phase was suppressed by adding small amounts of d(−)-2-amino-5-phosphonovaleric acid to the extracellular medium, or by increasing the amount of extracellular Mg²⁺. The electrical activity of the network was substantially increased by the addition of bicuculline to the extracellular medium. The results presented here show that the neuronal network may exist in two different dynamical states: one state in which the neuronal network behaves as a non-chaotic deterministic system and another state where the system exhibits large spatio-temporal fluctuations, characteristic of stochastic or chaotic systems. Received: 8 June 1999 / Accepted in revised form: 10 January 2000     

Spontaneous and evoked activities of interpositus nucleus neurons before and after lesion of the cerebellar cortex

Spontaneous and evoked activities of nucleus interpositus neurons (IN) of the cerebellum were examined before and after cerebellar paravermal cortex lesions in cats anesthetized with alpha-chloralose. It was found that spontaneous activity increased dramatically following cortical ablation: before the lesion only 4% of cells encountered fired at a rate exceeding 80 impulses/sec., whereas up to 40% discharged at this rate postoperatively. Responses to paw stimulation were also altered: the initial excitation was lengthened from 8.5 to 15.8 msec; narrow; trough causing segmentation in this excitation, which seems to result from Purkinje cell inhibition, was absent; and the succeeding inhibitory period was reduced in duration by 50%. Also after the lesion there was a strong tendency for the neurons to discharge in bursts. It is suggested that changes in cell activity in the IN following cortical lesion unveil neural mechanisms of motor disturbances in lesioned cats.     

Membrane potential synchrony in primary visual cortex during sensory stimulation

When the primary visual cortex (V1) is activated by sensory stimulation, what is the temporal correlation between the synaptic inputs to nearby neurons? This question underlies the origin of correlated activity, the mechanism of how visually evoked activity emerges and propagates in cortical circuits, and the relationship between spontaneous and evoked activity. Here, we have recorded membrane potential from pairs of V1 neurons in anesthetized cats and found that visual stimulation suppressed low-frequency membrane potential synchrony (0-10 Hz), and often increased synchrony at high frequencies (20-80 Hz). The increase in high-frequency synchrony occurred for neurons with similar orientation preferences and for neurons with different orientation preferences and occurred for a wide range of stimulus orientations. Thus, while only a subset of neurons spike in response to visual stimulation, a far larger proportion of the circuit is correlated with spiking activity through subthreshold, high-frequency synchronous activity that crosses functional domains.     

Responses of caudate neurons to clicks in chronic experiments on cats

Spontaneous and evoked unit activity in response to repeated application of clicks at a frequency of 0.3–2.0 Hz in the caudate nucleus was studied by an extracellular recording technique in chronic experiments on cats. Four types of spontaneous unit activity in the caudate nucleus were distinguished. Altogether 44% of neurons tested responded by changes in spontaneous activity to clicks. Five types of responses of caudate neurons to clicks were discovered: phasic excitation, phasic inhibition, tonic activation, tonic inhibition, and mixed tonic responses; the commonest type was tonic activation. During prolonged stimulation by clicks extinction of the phasic responses was not observed. Complete or partial extinction of tonic responses in the course of frequent repetition of stimulation was observed in 33% of responding neurons. The question of possible convergence of specific and nonspecific influences on caudate neurons is discussed.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 12, No. 1, pp. 28–35, January–February, 1980.     

视觉编码的快速学习

活动依赖的神经可塑性在视觉皮层信息处理过程中起着很重要的作用。该文将讲述几个关于视觉刺激引起皮层反应发生快速变化的研究工作。在体膜片钳的实验结果表明,将视觉刺激与能够诱发孽个视皮层神经元发放动作电位的电刺激相偶联可以改变神经元的感受野特性。单电极和多电极胞外记录的实验结果显示,反复地给予自然图形电影刺激,不仅能增加视皮层神经元反应的可靠性,而且能造成之后的自发活动中存在“记忆的痕迹”。最后,用电压敏感染料成像的方法对群体细胞活动进行考察,结果提示视觉活动之后的皮层回放可能是由皮层波介导的。